Staphylococcus aureus Enterotoxin A Gene Isolated From Raw Red Meat and Poultry in Tehran, Iran

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Mohammad Hossein Sarrafzadeh Zargar

2014-07-01

Full Text Available Background: Staphylococcus aureus is the most prevalent infectious agent of food materials. Enterotoxin producing types of S. aureus cause well-known food-borne disease. Staphylococcal Enterotoxin A (SEA is the most important agent of gastroenteritis. Objectives: The present study aimed to screen the raw meat samples collected from different regions of Tehran for S. aureus infection and type of encoding enterotoxin. Materials and Methods: Hundred and eighty six meat samples were collected randomly from city dealers and transferred to laboratory within screw cap containers. The samples were first cultured according to the standard bacteriological methods and then S. aureus isolates were identified using standard bacteriological tests. The isolates were subjected to Polymerase Chain Reaction (PCR to detect gene encoding SEA. Results: Staphylococcus aureus isolated from 29 (15.6% meat samples including beef 14.8%, raw lamb 15%, raw chicken 15.7% and raw turkey 16.6%. Using special primer sets proved that the species isolated from five samples (two raw chicken, two raw beef and one raw turkey encoded enterotoxin A. Conclusions: Although staphylococcal contamination within food material is more or less a routine, but detection of enterotoxin encoding species from raw meat samples is alarming for health authorities. These data highlight the importance of periodic surveillance of raw meat distributed among ordinary consumers.

Staphylococcal superantigens; toxic shock syndrome toxin-1 and enterotoxins in pediatric otitis media effusion: a brief report

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Noorbakhsh S

2013-03-01

Full Text Available Background: Staphylococcal superantigens (SAg's may have some role in otitis media with effusion (OME. The aim of this study was the search of staphylococcal SAg's in middle ear effusion of children with OME.  Methods: This cross sectional-analytic study was done in ENT & pediatric wards upon 64 children with otitis media with effusion (OME between 1-15 years, (mean age=7.42+4 years of Rasoul Akram University Hospital, Tehran, Iran in 2009-2011. Fifty six percent (36 of cases were male, 43.8% (28 were female. Staphylococcal SAg's; Toxic Shock Syndrome Toxin-1 (TSST-1, Staphylococcal enterotoxin A, B, C, D (Enzyme immune assay, AB Cam, USA were detected in middle ear effusion samples after conventional culture.Results: None type of SAg's found in 39% of OME cases, enterotoxin B found in: 22%; enterotoxin A: 17%, enterotoxin C: 15.6%, enterotoxin D: 12.5%, Toxic Shock Syndrome Toxin-1 (TSST-1: 7.8% Mean age of cases with positive TSST-1, enterotoxin A, B, C, and D was: 1, 5, 8.6, 9.6 and 9.6 years respectively. Positive TSST had no agreement with positive enterotoxin A and C but had weak agreement with type B and D. Mean age of cases with positive TSST was one years which had significant difference with (7.9 years in cases with negative TSST test (P<0.0001.Conclusion: At least one or more type of staphylococcal toxins had found in middle ear effusion of 70% of OME cases with negative culture for Staphylococcus aureus. Even in culture negative cases, staphylococcal toxins might have some immunologic role in middle ear effusion forming. Finding the SAg's (at least one type are important for treatment of immunosuppressive or corticosteroid in cases with resistant OME.

Production of enterotoxins of Staphylococcus spp. isolated from samples of sheep milk

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František Zigo

2014-02-01

Full Text Available In our study was followed occurrence of mastitis in herd of 430 sheep of breed zoslachtena valaska with hand milking technology examined two times during one lactation season. Individual examination consisted from clinical examination of udder and microbiological examination of milk samples. By PCR was determined presence of genes coding production of enterotoxins, and by ELISA methods production individual types of enterotoxins. From individual forms of mastitis were frequently detected subacute (6.7%, subclinical (5.7% and acute (2.9%. The coagulase-negative staphylococci (CNS were identified in 102 (65.4% from all 156 positive isolates. The CNS and S. aureus caused subacute (5.1%, subclinical (3.9% and acute (2.4% forms of mastitis. The most frequently isolated were S. epidermidis, followed by S. chromogenes and S. xylosus from ewes with subacute and subclinical mastitis. From acute and chronical forms of mastitis were  predominantly isolated S. aureus, S. uberis and S. epidermidis. The production of staphylococcal enterotoxins (SE - SEA, SEB, SEC, SED and the presence of genes sec (3, sea (2, seb (2 and sed (2 were determined in S. aureus, S. epidermidis, S. schleiferi and S. chromogenes, respectively. The results suggested on the high occurrence (12.4% of subacute and subclinical forms. Confirmed production of enterotoxins and presence of genes coding their production present a risk for human health and decreased a quality of milk and products from sheep´s milk.

Prevalence of enterotoxigenic Bacillus Cereus and Its enterotoxins ...

African Journals Online (AJOL)

Objectives: To determine the prevalence of enterotoxigenic Bacillus cereus (B. cereus) and enterotoxins in milk and milk products. Design: A random sampling of milk products was carried out. Setting: Market milk and milk products were collected from retail shops in Nairobi and analysed for contamination with ...

Cloning and Expression of Nano Body Gene against Enterotoxin B of Staphylococcus Aureus

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Zahra Tavassoli

2017-02-01

Full Text Available Background & Objectives: Staphylococcus aureus bacteria causes many different diseases by secretion of various enterotoxins. Therefore, it is necessary to develop ways that facilitate the detection of enterotoxins. Nowadays, immunochemical methods which are based on monoclonal antibody technology are used. The heavy chain antibodies that are called VHH or Nano body were found in blood serum of the Camelidae family. The unique properties of this antibody such as their binding to small molecules like toxins make them attractive candidates for the development of immunodiagnostic tests. The present study was done to achieve a VHH molecules against Staphylococcus enterotoxin B. Materials & Methods: Freighting phage library for isolate private Nano bodies against enterotoxin B was done in previous works. Next, pCANTAB 5E vector that consists VHH, extracted from E.coli bacteria strain xl1blue, and after doing PCR process with relative primers, sub cloning in pET21a(+ as an expression vector with cut sites NdeI and XhoI was done. Transformation in E.coli bacteria strain BL21(DE3 was done. Then, the cells effected with IPTG and producing time, and other terms were optimized. Finally, the expression of the protein with SDS-PAGE and western blot techniques was evaluated. Result: For proving cloning of nano body gene in pET21a (+ vector, nucleotide sequence of gene was analyzed, and transforming to E.coli bacteria strain BL21(DE3 was successful. After inspiration, active protein in cell was seen by SDS-PAGE technique and proved by western blot. Conclusion: cloning, sub cloning, and nonabody expression were surveyed in this research. Production of this protein can help to develop new therapeutic methods and produce vaccine against enterotoxin B of Staphylococcus aureus

Stimulation of Clostridium perfringens enterotoxin formation by caffeine and theobromine.

Science.gov (United States)

Labbe, R G; Nolan, L L

1981-01-01

In the presence of 100 micrograms of caffeine per ml or 200 micrograms of theobromine per ml, sporulation of Clostridium perfringens NCTC 8679 rose from less than 1 to 80 or 85%. Enterotoxin concentration increased from undetectable levels to 450 micrograms/mg of cell extract protein. Heat-resistant spore levels increased from less than 1,000 to between 1 X 10(7) and 2 X 10(7)/ml. These effects were partially reversible by the addition of adenosine or thymidine. In the case of NCTC 8238, caffeine and theobromine caused a three- to fourfold increase in the percentages of cells possessing refractile spores and a similar increase in enterotoxin concentration. Heat-resistant spore levels, however, were unaffected. Inosine was ineffective in promoting sporulation in NCTC 8679. PMID:6271685

Inhibitory effect of totarol on exotoxin proteins hemolysin and enterotoxins secreted by Staphylococcus aureus.

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Shi, Ce; Zhao, Xingchen; Li, Wenli; Meng, Rizeng; Liu, Zonghui; Liu, Mingyuan; Guo, Na; Yu, Lu

2015-10-01

Staphylococcus aureus (S. aureus) causes a wide variety of infections, which are of major concern worldwide. S. aureus produces multiple virulence factors, resulting in food infection and poisoning. These virulence factors include hyaluronidases, proteases, coagulases, lipases, deoxyribonucleases and enterotoxins. Among the extracellular proteins produced by S. aureus that contribute to pathogenicity, the exotoxins α-hemolysin, staphylococcal enterotoxin A (SEA) and staphylococcal enterotoxin B (SEB) are thought to be of major significance. Totarol, a plant extract, has been revealed to inhibit the proliferation of several pathogens effectively. However, there are no reports on the effects of totarol on the production of α-hemolysin, SEA or SEB secreted by S. aureus. The aim of this study was to evaluate the effects of totarol on these three exotoxins. Hemolysis assay, western blotting and real-time reverse transcriptase-PCR assay were performed to identify the influence of graded subinhibitory concentrations of totarol on the production of α-hemolysin and the two major enterotoxins, SEA and SEB, by S. aureus in a dose-dependent manner. Moreover, an enzyme linked immunosorbent assay showed that the TNF-α production of RAW264.7 cells stimulated by S. aureus supernatants was inhibited by subinhibitory concentrations of totarol. Form the data, we propose that totarol could potentially be used as a promising natural compound in the food and pharmaceutical industries.

[Enterotoxin genes occurance among S. aureus strains isolated from inpatients and carriers].

Science.gov (United States)

Lawrynowicz-Paciorek, Maja; Kochman, Maria; Piekarska, Katarzyna; Wyrebiak, Agata; Potracka, Ewa; Leniak-Chmiel, Urszula; Magdziak, Agnieszka

2006-01-01

We examined 44 inpatients and 66 carriers Staphylococcus aureus strains, isolated in years 2002-2005, for the presence of 18 enterotoxin genes (se/sel) (by PCR), the ability for A-D enterotoxin production (by SET-RPLA) and antibiotic resistance distribution (by disc diffusion method). se/sel genes were detected in 90,9% of all strains, sea (70,5%) and selk and selq (52,3%) - among inpatients strains and egc (65,2%) - among carriers strains were the most frequently se/sel genes found. Positive results of SET-RPLA were consistent with PCR results. There was no correlation observed between antibiotic resistance and se/sel genes distribution among tested S. aureus strains.

Construction of PVX virus-expression vector to express enterotoxin ...

African Journals Online (AJOL)

Potato X potyvirus (PVX)-based vector has been comprehensively applied in transient expression system. In order to produce the heterologous proteins more quickly and stably, the ClaI and NotI enzyme sites were introduced into the Enterotoxin fusion gene LTB-ST by polymerase chain reaction (PCR) and the LTB-ST ...

Characterization of Enterotoxigenic Bacillus cereus sensu lato and Staphylococcus aureus Isolates and Associated Enterotoxin Production Dynamics in Milk or Meat-Based Broth.

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Walker-York-Moore, Laura; Moore, Sean C; Fox, Edward M

2017-07-15

Bacillus cereus sensu lato species, as well as Staphylococcus aureus , are important pathogenic bacteria which can cause foodborne illness through the production of enterotoxins. This study characterised enterotoxin genes of these species and examined growth and enterotoxin production dynamics of isolates when grown in milk or meat-based broth. All B. cereus s. l. isolates harboured nheA , hblA and entFM toxin genes, with lower prevalence of bceT and hlyII . When grown at 16 °C, toxin production by individual B. cereus s. l. isolates varied depending on the food matrix; toxin was detected at cell densities below 5 log 10 (CFU/mL). At 16 °C no staphylococcal enterotoxin C (SEC) production was detected by S. aureus isolates, although low levels of SED production was noted. At 30 °C all S. aureus isolates produced detectable enterotoxin in the simulated meat matrix, whereas SEC production was significantly reduced in milk. Relative to B. cereus s. l. toxin production, S. aureus typically required reaching higher cell numbers to produce detectable levels of enterotoxin. Phylogenetic analysis of the sec and sel genes suggested population evolution which correlated with animal host adaptation, with subgroups of bovine isolates or caprine/ovine isolates noted, which were distinct from human isolates. Taken together, this study highlights the marked differences in the production of enterotoxins both associated with different growth matrices themselves, but also in the behaviour of individual strains when exposed to different food matrices.

Organ Culture as a Model System for Studies on Enterotoxin Interactions with the Intestinal Epithelium

DEFF Research Database (Denmark)

Lorenzen, Ulver Spangsberg; Hansen, Gert H; Danielsen, E Michael

2015-01-01

Studies on bacterial enterotoxin-epithelium interactions require model systems capable of mimicking the events occurring at the molecular and cellular levels during intoxication. In this chapter, we describe organ culture as an often neglected alternative to whole-animal experiments or enterocyte......-like cell lines. Like cell culture, organ culture is versatile and suitable for studying rapidly occurring events, such as enterotoxin binding and uptake. In addition, it is advantageous in offering an epithelium with more authentic permeability/barrier properties than any cell line, as well...

The Isolation and Detection of Staphylococcus aureus Enterotoxins A-E and TSST-1 Genes from Different Sources by PCR Method

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Norouzi J

2012-09-01

Full Text Available Background and Objectives: Enterotoxins and toxic shock syndrome toxin–1 (TSST-1 are an important virulence factor of S. aureus. The purpose of this study was to analyze the presence of S. aureus enterotoxin (sea-see and tst genes in the samples that collected from different sources with PCR method. Methods: During 5 months from 150 collected samples, 80 strains were identified as S. aureus. PCR reaction was used for investigation on the presence of genes for staphylococcal enterotoxins (A-E and toxic shock syndrome toxin-1 (TSST-1. Results: Fifty three samples (66.25% out of 80 samples were positive for one or more ETs and TSST-1 genes. of these positive strains, 17 (32.07% were positive for sea 39 (73.58% for seb, 30 (56.6% for sec, 2 (3.7% for sed, 21 (39.62% for see, and 14 (26.41% for tst. Enterotoxins and tst in 40 samples (75.47% observed.Conclusion: In this study, high prevalence of S. aureus, its enterotoxin, and tst genes were observed in clinical samples, food samples, and healthy people. This fact emphasizes on the role of human as original source and carrier of S. aureus. Also, use of PCR reaction for detection of these genes in S. aureus that isolated from various sources is recommended.

Cloning in Escherichia coli of the enterotoxin gene from Clostridium perfringens type A.

Science.gov (United States)

Iwanejko, L A; Routledge, M N; Stewart, G S

1989-04-01

A 26 bp DNA probe has been constructed with minimal degeneracy to the protein sequence for Clostridium perfringens enterotoxin. The probe has been hybridized against a 6-10 kb chromosomal bank from C. perfringens 8239, prepared as a HindIII partial digest in pHG165. From this survey a clone has been identified containing a 6.8 kb DNA insert with strong hybridization to the probe. Direct plasmid sequencing has identified a translational reading frame within this clone which correlates with the known protein sequence for the type A enterotoxin. DNA sequences 5' to this open reading frame and containing the putative transcriptional control regions show areas of significant homology with regions upstream from the ATG codon of the tetanus toxin gene.

Detection of enterotoxins produced by B. cereus through PCR analysis of ground and roasted coffee samples in Rio de Janeiro, Brazil

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Cyllene de Matos Ornelas da Cunha Corrêa de Souza

2011-06-01

Full Text Available Coffee is one of the most appreciated drinks in the world. Coffee ground is obtained from the fruit of a small plant that belongs to the genus Coffea. Coffea arabica and Coffea canephora robusta are the two most commercially important species. They are more commonly known as arabica and robusta, respectively. Two-thirds of Coffea arabica plants are grown in South and Central America, and Eastern Africa - the place of origin for this coffee species. Contamination by microorganisms has been a major matter affecting coffee quality in Brazil, mainly due to the harvesting method adopted. Brazilian harvests are based on fruits collected from the ground mixed with those that fall on collection cloths. As the Bacillus cereus bacterium frequently uses the soil as its environmental reservoir, it is easily capable of becoming a contaminant. This study aimed to evaluate the contamination and potential of B. cereus enterotoxin genes encoding the HBL and NHE complexes, which were observed in strains of ground and roasted coffee samples sold in Rio de Janeiro. The PCR (Polymerase Chain Reaction results revealed high potential of enterotoxin production in the samples. The method described by Speck (1984 was used for the isolation of contaminants. The investigation of the potential production of enterotoxins through isolates of the microorganism was performed using the B. cereus enterotoxin Reverse Passive Latex Agglutination test-kit (BCET-RPLA, Oxoid, according to the manufacturer's instructions. The potential of enterotoxin production was investigated using polymerase chain reaction (PCR methods for hblA, hblD and hblC genes (encoding hemolysin HBL and for nheA, nheB and nheC genes (encoding non-hemolytic enterotoxin - NHE. Of all the 17 strains, 100% were positive for at least 1 enterotoxin gene; 52.9% (9/17 were positive for the 3 genes encoding the HBL complex; 35.3% (6/17 were positive for the three NHE encoding genes; and 29.4% (5/17 were positive for

Effects of frozen storage on survival of Staphylococcus aureus and enterotoxin production in precooked tuna meat.

Science.gov (United States)

Wu, Xulei; Su, Yi-Cheng

2014-08-01

This study investigated the survival of Staphylococcus aureus in precooked tuna meat for producing canned products during frozen storage (-20 ± 2 °C) as well as its growth and enterotoxin production at 35 to 37 °C after the storage. Samples (50 ± 5 g) of precooked albacore (loin, chunk, and flake) and skipjack (chunk and flake) tuna were inoculated with 5 enterotoxin-producing strains of S. aureus at a level of approximately 3.5 log CFU/g and individually packed in a vacuum bag after 3 h incubation at 35 to 37 °C. Vacuum-packed samples were stored in a freezer (-20 ± 2 °C) for 4 wk. The frozen samples were then thawed in 37 °C circulating water for 2 h and incubated at 35 to 37 °C for 22 h. Populations of S. aureus in all precooked tuna samples decreased slightly (canned tuna within 6 to 8 h of thawing to avoid product spoilage and potential enterotoxin production by S. aureus in contaminated precooked tuna meat. © 2014 Institute of Food Technologists®

Prevalence of Staphylococcal Enterotoxins in Ready-to-Eat Foods Sold in Istanbul.

Science.gov (United States)

Ulusoy, Beyza H; Çakmak Sancar, Burcu; Öztürk, Muhsin

2017-10-01

The aim of this study was to investigate the prevalence of staphylococcal enterotoxins (SEs) in ready-to-eat (RTE) foods sold in Istanbul, Turkey. A total of 5,241 samples were randomly collected from various caterers, hotels, and restaurants from 2014 to 2016. The samples were classified into four groups: (i) various cooked RTE meat and vegetable meals, (ii) various RTE salads, charcuterie, and cold appetizers, (iii) various cooked RTE bakery products (pasta, pastries, pizza, pita, ravioli, etc.), and (iv) any cooked RTE sweets and desserts (pudding, custard, cream, ashura, etc.). The samples were examined for the presence of SEs by 3M Tecra Staph Enterotoxin Visual Immunoassay method, which is a manual enzyme-linked immunosorbent assay method. Among all samples, only 1 (0.019%) RTE meal (vegetable meal with meat) was found to be contaminated with SEs, a good result in terms of staphylococcal food poisoning risk and public health.

Selection and characterization of DNA aptamers against Staphylococcus aureus enterotoxin C1.

Science.gov (United States)

Huang, Yukun; Chen, Xiujuan; Duan, Nuo; Wu, Shijia; Wang, Zhouping; Wei, Xinlin; Wang, Yuanfeng

2015-01-01

Enterotoxins from pathogenic bacteria are known as the main reason that can cause the bacterial foodborne diseases. In this study, aptamers that bound to Staphylococcus aureus enterotoxin C1 (SEC1) with high affinity and selectivity were generated in vitro by twelve rounds of selection based on magnetic separation technology, with a low-level dissociation constant (Kd) value of 65.14 ± 11.64 nmol/L of aptamer C10. Aptamer-based quantification of SEC1 in the food sample by a graphene oxide (GO)-based method was implemented to investigate the potential of the aptamer against SEC1 with a limit of detection of 6 ng/mL. On the basis of this work, biosensors using the selected SEC1 aptamers as new molecular recognition elements could be applied for innovative determinations of SEC1. Copyright © 2014 Elsevier Ltd. All rights reserved.

Detection of coding genes for enterotoxins in Bacillus cereus by PCR and their products by BCET-RPLA and ELISA Assay

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Marcela Vyletělová

2010-01-01

Full Text Available Determination of enterotoxin production, diarrhoeal and emetic gene identification was studied in 41 Bacillus cereus strains isolated from raw cows’ and raw goats’ milk, pasteurized milk, dairy products during technological processing and from dairy plant equipment. Presence of enterotoxins was detected by BCET-RPLA (HBL and ELISA immunoassay (NHE. Gene identification (nheA, nheB, nheC, hblA, hblC, hblD, bceT, cytK-1, cytK-2, entFM and ces was achieved by means of PCR. Enterotoxin HBL was detected in 32 strains, enterotoxin NHE in all 41 strains. Presence of all three genes nheA, nheB and nheC was confirmed in 40 strains and genes hblA, hblC and hblD in 29 strains. Comparison of used methods was as follow: 1 BCET-RPLA (which detects L2 component and PCR (positive or negative all three hblA, hblC and hblD gene detection were identical in 30 (73%; 2 ELISA (NheA and PCR (all three nheC, nheB and nheA gene expression were identical in 40 (98% cases isolated strains.

Inhibition of Binding of the AB5-Type Enterotoxins LT-I and Cholera Toxin to Ganglioside GM1 by Galactose-Rich Dietary Components

NARCIS (Netherlands)

Becker, P.M.; Widjaja-Greefkes, H.C.A.; Wikselaar, van P.G.

2010-01-01

Cholera, travelers' diarrhea, or colibacillosis in pigs can possibly be prevented or attenuated by dietary provision of competitive inhibitors that react with the GM1-binding sites of the enterotoxins cholera toxin (CT), human Escherichia coli heat-labile enterotoxin of serogroup I (LTh-I), and

Role of Enterotoxin-Producing Staphylococci in Zoonotic Infections in Iran

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Abdolmajid Ghasemian

2018-05-01

Full Text Available Context: Staphylococci spp, are enterotoxin-producing zoonotic agents causing a variety of infections such as mastitis in animals and wound bite infections in humans. This review was conducted to determine the prevalence of Staphylococci infections especially to uncover enterotoxin-producing species in Iran. Evidence acquisition: for this review, words of "Staphylococcus", "zoonotic", "prevalence", "animals", "human" and "Iran" were searched in the internet engines such as Google Scholar, PubMed, Google, Science Direct and so on. Patients with no history of contact with animals were also included in the study for comparison aims. Both veterinary and human coagulase positive isolates were included. Data was analyzed with Graph Pad Prism 6, meta-analysis section. A total of 20 previous studies (450 clinical samples were found. S. intermedius was the predominant isolate identified in veterinary sources. Other coagulase positive spp such as S. hycus and S. simulans were isolated with lower prevalence, but S. delphini has not been detected. Conclusion: S. intermedius was the most isolate identified in veterinary sources with potential of causing infections in humans. Other coagulase positive spp such as S. hycus and S. simulans were isolated with lower prevalence, but S. delphini was not detected.

The influence of radiation on the enterotoxin and thermoresistant deoxyribo-nuclease production by Staphylococcus sp

International Nuclear Information System (INIS)

Szulc, M.; Pliszka, A.; Peconek, J.

1980-01-01

Six strains of Staph. aureus present in the environment with and without protein were exposed to X rays in the doses range from 1 to 400 Gy. The production of enterotoxin and thermoresistant deoxyribonuclease by enterotoxic strains No 262 and 100 was determined. All the strains showed similar radiosensitivity to X rays. About 1 per cent of protein in the environment exposed to radiation increased the resistance of Staphylococcus strains. The doses of radiation from 1 to 400 Gy did not influence the enterotoxin A and B production by bacteria multiplied from those which had survived the radiation. The irradiation of the strains present in the environment without protein brought about a temporary inhibition of thermoresistant deoxyribonuclease production. (author)

Estrogen inhibits chloride secretion caused by cholera and Escherichia coli enterotoxins in female rat distal colon.

LENUS (Irish Health Repository)

Alzamora, Rodrigo

2011-05-08

Excessive Cl(-) secretion is the driving force for secretory diarrhea. 17β-Estradiol has been shown to inhibit Cl(-) secretion in rat distal colon through a nongenomic pathway. We examined whether 17β-estradiol inhibits Cl(-) secretion in an animal model of secretory diarrhea and the downstream effectors involved. The effect of 17β-estradiol on cholera toxin and heat-stable enterotoxin induced Cl(-) secretion in rat colonic mucosal sheets was studied by current-voltage clamping. Selective permeabilization of apical or basolateral membranes with amphotericin B or nystatin was used to isolate basolateral K(+) channel and apical Cl(-) channel activity, respectively. 17β-Estradiol dose-dependently inhibited secretory responses to both toxins with IC(50) values of approximately 1nM. This effect was female-gender specific, with no inhibition observed in male tissues. 17β-Estradiol responses were insensitive to the pure anti-estrogen ICI 182,720. 17β-Estradiol exerted its effects downstream of enterotoxin-induced production of second messengers (cAMP and cGMP) but was dependent on PKCδ activation. In nystatin-permeabilized tissues, apical Cl(-) currents were unaffected by 17β-estradiol treatment while basolateral K(+) current was profoundly inhibited by the hormone. This current was sensitive to the specific KCNQ1 channel inhibitors chromanol 293B and HMR-1556. In conclusion, 17β-estradiol inhibits enterotoxin-induced Cl(-) secretion via a PKCδ-dependent mechanism involving inhibition of basolateral KCNQ1 channels. These data elucidate mechanisms of 17β-estradiol inhibition of Cl(-) secretion induced by enterotoxins in intestinal epithelia, which may be relevant for the treatment of diarrheal diseases.

Estrogen inhibits chloride secretion caused by cholera and Escherichia coli enterotoxins in female rat distal colon.

LENUS (Irish Health Repository)

Alzamora, Rodrigo

2012-02-01

Excessive Cl(-) secretion is the driving force for secretory diarrhea. 17beta-Estradiol has been shown to inhibit Cl(-) secretion in rat distal colon through a nongenomic pathway. We examined whether 17beta-estradiol inhibits Cl(-) secretion in an animal model of secretory diarrhea and the downstream effectors involved. The effect of 17beta-estradiol on cholera toxin and heat-stable enterotoxin induced Cl(-) secretion in rat colonic mucosal sheets was studied by current-voltage clamping. Selective permeabilization of apical or basolateral membranes with amphotericin B or nystatin was used to isolate basolateral K(+) channel and apical Cl(-) channel activity, respectively. 17beta-Estradiol dose-dependently inhibited secretory responses to both toxins with IC(50) values of approximately 1nM. This effect was female-gender specific, with no inhibition observed in male tissues. 17beta-Estradiol responses were insensitive to the pure anti-estrogen ICI 182,720. 17beta-Estradiol exerted its effects downstream of enterotoxin-induced production of second messengers (cAMP and cGMP) but was dependent on PKCdelta activation. In nystatin-permeabilized tissues, apical Cl(-) currents were unaffected by 17beta-estradiol treatment while basolateral K(+) current was profoundly inhibited by the hormone. This current was sensitive to the specific KCNQ1 channel inhibitors chromanol 293B and HMR-1556. In conclusion, 17beta-estradiol inhibits enterotoxin-induced Cl(-) secretion via a PKCdelta-dependent mechanism involving inhibition of basolateral KCNQ1 channels. These data elucidate mechanisms of 17beta-estradiol inhibition of Cl(-) secretion induced by enterotoxins in intestinal epithelia, which may be relevant for the treatment of diarrheal diseases.

PCR detection of staphylococcal enterotoxin genes in Staphylococcus aureus strains isolated from raw and pasteurized milk.

Science.gov (United States)

Rall, V L M; Vieira, F P; Rall, R; Vieitis, R L; Fernandes, A; Candeias, J M G; Cardoso, K F G; Araújo, J P

2008-12-10

Milk is considered a nutritious food because it contains several important nutrients including proteins and vitamins. Conversely, it can be a vehicle for several pathogenic bacteria such as Staphylococcus aureus. This study aimed to analyze the frequency of genes encoding the staphylococcal enterotoxins (SEs) SEA, SEB, SEC, SED, SEE, SEG, SEH, SEI and SEJ in S. aureus strains isolated from raw or pasteurized bovine milk. S. aureus was found in 38 (70.4%) out of 54 raw milk samples at concentrations of up to 8.9 x 10(5) CFU/ml. This microorganism was present in eight samples of pasteurized milk before the expiration date and in 11 samples analyzed on the expiration date. Of the 57 strains studied, 68.4% were positive for one or more genes encoding the enterotoxins, and 12 different genotypes were identified. The gene coding for enterotoxin A, sea, was the most frequent (16 strains, 41%), followed by sec (8 strains, 20.5%), sed (5 strains, 12.8%), seb (3 strains, 7.7%) and see (2 strains, 5.1%). Among the genes encoding the other enterotoxins, seg was the most frequently observed (11 strains, 28.2%), followed by sei (10 strains) and seh and sej (3 strains each). With the recent identification of new SEs, the perceived frequency of enterotoxigenic strains has increased, suggesting that the pathogenic potential of staphylococci may be higher than previously thought; however, further studies are required to assess the expression of these new SEs by S. aureus, and their impact in foodborne disease. The quality of Brazilian milk is still low, and efforts from the government and the entire productive chain are required to attain consumer safety.

Enterotoxin Producing Ability And Antimicrobial Susceptibility Of Coagulase-Negative Staphylococci Isolated From Goat Milk Cheese And Salted Yoghurt In Turkey

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S. Pehlivanlar Onen

2017-10-01

Full Text Available The aim of this study was to determine enterotoxin producing ability and antimicrobial susceptibility of coagulase negative staphylococci CNS in goats bulk milk cheese and salted yoghurt. CNS strains were identified by using GP card in VITEK 2 system. The presence of enterotoxins was determined by enzyme immunoassay test by using RIDASCREEN test kit. Antibiotic susceptibility in CNS strains was detected by using AST-P640 card in VITEK 2 system. A total of 100 CNS strains were isolated in 22 55 bulk milk samples and in 23 57.5 cheese samples. Staphylococcus spp. could not be isolated from salted yoghurt samples. The most encountered species were S. caprae 51.9 S. chromogenes 11.5 and S. xylosus 9.6 from milk samples and S. saprophyticus 60.4 S. xylosus 12.5 and S. haemolyticus 8.3 from cheese samples. Four CNS strains 4 isolated from samples were capable of producing enterotoxin. While all isolates were resistant to at least one antibiotic 74 of CNS strains showed resistance to two or more antibiotics. Enterotoxin production ability and high antibiotic resistance of the CNS strains isolated from goat bulk milk and cheese can lead to a risk for public health.

An Enterotoxin-Like Binary Protein from Pseudomonas protegens with Potent Nematicidal Activity.

Science.gov (United States)

Wei, Jun-Zhi; Siehl, Daniel L; Hou, Zhenglin; Rosen, Barbara; Oral, Jarred; Taylor, Christopher G; Wu, Gusui

2017-10-01

Soil microbes are a major food source for free-living soil nematodes. It is known that certain soil bacteria have evolved systems to combat predation. We identified the nematode-antagonistic Pseudomonas protegens strain 15G2 from screening of microbes. Through protein purification we identified a binary protein, designated Pp-ANP, which is responsible for the nematicidal activity. This binary protein inhibits Caenorhabditis elegans growth and development by arresting larvae at the L1 stage and killing older-staged worms. The two subunits, Pp-ANP1a and Pp-ANP2a, are active when reconstituted from separate expression in Escherichia coli The binary toxin also shows strong nematicidal activity against three other free-living nematodes ( Pristionchus pacificus , Panagrellus redivivus , and Acrobeloides sp.), but we did not find any activity against insects and fungi under test conditions, indicating specificity for nematodes. Pp-ANP1a has no significant identity to any known proteins, while Pp-ANP2a shows ∼30% identity to E. coli heat-labile enterotoxin (LT) subunit A and cholera toxin (CT) subunit A. Protein modeling indicates that Pp-ANP2a is structurally similar to CT/LT and likely acts as an ADP-ribosyltransferase. Despite the similarity, Pp-ANP shows several characteristics distinct from CT/LT toxins. Our results indicate that Pp-ANP is a new enterotoxin-like binary toxin with potent and specific activity to nematodes. The potency and specificity of Pp-ANP suggest applications in controlling parasitic nematodes and open an avenue for further research on its mechanism of action and role in bacterium-nematode interaction. IMPORTANCE This study reports the discovery of a new enterotoxin-like binary protein, Pp-ANP, from a Pseudomonas protegens strain. Pp-ANP shows strong nematicidal activity against Caenorhabditis elegans larvae and older-staged worms. It also shows strong activity on other free-living nematodes ( Pristionchus pacificus , Panagrellus redivivus , and

A complex array of Hpr consensus DNA recognition sequences proximal to the enterotoxin gene in Clostridium perfringens type A.

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Brynestad, S; Iwanejko, L A; Stewart, G S; Granum, P E

1994-01-01

Enterotoxin production in Clostridium perfringens is both strain dependent and sporulation associated. Underlying these phenotypic observations must lie a genetic and molecular explanation and the principal keys will be held within the DNA sequence both upstream and downstream of the structural gene cpe. In accordance with the above we have sequenced 4.1 kbp of DNA upstream of cpe in the type strain NCTC 8239. A region of DNA extending up to 1.5 kb 5' to cpe is conserved in all enterotoxin-positive strains. This region contains a putative ORF with substantial homology to an ORF in the Salmonella typhimurium IS200 insertion element and, in addition, contains multiple perfect consensus DNA-binding sequences for the Bacillus subtilis transition state regulator Hpr. The detailed structural elements revealed by the sequence analysis are presented and used to develop a new perspective on the molecular basis of enterotoxin production in this important food-poisoning bacterium.

Cloning the enterotoxin gene from Clostridium perfringens type A

OpenAIRE

Iwanejko, Lesley Ann.

1991-01-01

A C. perfringens type A genomic library was constructed in E. coli by banking overlapping 6-10 kbp Hind III fragments of chromosomal DNA from the enterotoxin (CPE) positive strain NCTC 8239 into the pUC derived vector pHG165. The library was screened by colony hybridization with a degenerate 26 bp oligonucleotide probe, derived from the amino acid sequence CPE9_17A. complex mixture of plasmid DNA was isolated from the only hybridization positive clone. A second round of screening picked out a...

Growth and enterotoxin production of Staphylococcus aureus during the manufacture and ripening of Camembert-type cheeses from raw goats' milk.

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Meyrand, A; Boutrand-Loei, S; Ray-Gueniot, S; Mazuy, C; Gaspard, C E; Jaubert, G; Perrin, G; Lapeyre, C; Vernozy-Rozand, C

1998-09-01

Tests were carried out to determine the effect of manufacturing procedures for a Camembert-type cheese from raw goats' milk on the growth and survival of Staphylococcus aureus organisms added to milk at the start of the process, and to study the possible presence of staphylococcal enterotoxin A in these cheeses. The initial staphylococcal counts were, respectively, 2, 3, 4, 5 and 6 log cfu ml-1. Cheese was prepared following the industrial specifications and ripened for 41 d. Detection of enterotoxins was done by the Vidas SET test and by an indirect double-sandwich ELISA technique using antienterotoxin monoclonal antibodies. Generally, numbers of microbes increased at a similar rate during manufacture in all cheeses until salting. During the ripening period, the aerobic plate count population and Staph. aureus levels remained stable and high. There was an approximately 1 log reduction of Staph. aureus in cheeses made with an initial inoculum of Staph. aureus greater than 10(3) cfu ml-1 at the end of the ripening period (41 d) compared with the count at 22 h. The level of staphylococcal enterotoxin A recovered varied from 1 to 3.2 ng g-1 of cheese made with an initial population of 10(3)-10(6) cfu ml-1. No trace of enterotoxin A was detected in cheeses made with the lowest Staph. aureus inoculum used in this study.

How do the rotavirus NSP4 and bacterial enterotoxins lead differently to diarrhea?

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Vasseur Monique

2007-03-01

Full Text Available Abstract Rotavirus is the major cause of infantile gastroenteritis and each year causes 611 000 deaths worldwide. The virus infects the mature enterocytes of the villus tip of the small intestine and induces a watery diarrhea. Diarrhea can occur with no visible tissue damage and, conversely, the histological lesions can be asymptomatic. Rotavirus impairs activities of intestinal disaccharidases and Na+-solute symports coupled with water transport. Maldigestion of carbohydrates and their accumulation in the intestinal lumen as well as malabsorption of nutrients and a concomitant inhibition of water reabsorption can lead to a malabsorption component of diarrhea. Since the discovery of the NSP4 enterotoxin, diverse hypotheses have been proposed in favor of an additional secretion component in the pathogenesis of diarrhea. Rotavirus induces a moderate net chloride secretion at the onset of diarrhea, but the mechanisms appear to be quite different from those used by bacterial enterotoxins that cause pure secretory diarrhea. Rotavirus failed to stimulate Cl- secretion in crypt, whereas it stimulated Cl- reabsorption in villi, questioning, therefore, the origin of net Cl- secretion. A solution to this riddle was that intestinal villi do in fact secrete chloride as a result of rotavirus infection. Also, the overall chloride secretory response is regulated by a phospholipase C-dependent calcium signaling pathway induced by NSP4. However, the overall response is weak, suggesting that NSP4 may exert both secretory and subsequent anti-secretory actions, as did carbachol, hence limiting Cl- secretion. All these characteristics provide the means to make the necessary functional distinction between viral NSP4 and bacterial enterotoxins.

Lab-on-a-chip for label free biological semiconductor analysis of Staphylococcal Enterotoxin B

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Yang, Minghui; Sun, Steven; Bruck, Hugh Alan; Kostov, Yordan; Rasooly, Avraham

2010-01-01

We describe a new lab-on-a-chip (LOC) which utilizes a biological semiconductor (BSC) transducer for label free analysis of Staphylococcal Enterotoxin B (SEB) (or other biological interactions) directly and electronically. BSCs are new transducers based on electrical percolation through a

Molecular Characterization of Enterotoxin-Producing Escherichia coli Collected in 2011-2012, Russia.

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Nikolay N Kartsev

Full Text Available Enterotoxin-producing Escherichia coli (ETEC are one of the main causative agents of diarrhea in children especially in developing countries and travel diarrhoea in adults. Pathogenic properties of ETEC associated with their ability to produce a heat-stable (ST and/or heat-labile (LT enterotoxins, as well as adhesins providing bacterial adhesion to intestinal epithelial cells. This study presents the molecular characterization of the ETEC isolates collected from the Central and Far-Eastern regions of Russia in 2011-2012. It was shown that all ETEC under study (n=18 had the heat-labile enterotoxin-coding operon elt, and had no the genes of the heat-stable enterotoxin operon est. DNA sequencing revealed two types of nucleotide exchanges in the eltB gene coding subunit B of LT in isolates collected from Cherepovets city (Central region, Russia and Vladivostok city (Far-East region, Russia. Only one ETEC strain carried genes cfaA, cfaB, cfaC and cfaD coding adhesion factor CFA/I. Expression of LT in four ETEC isolates in the agglutination reaction was detected using a latex test-system. The isolates were assigned to serogroups O142 (n = 6, О6 (n = 4, О25 (n = 5, О26 (n = 2, and O115 (n = 1. Genotyping showed that they belonged to an earlier described sequence-type ST4 (n = 3 as well as to 11 novel sequence-types ST1043, ST1312, ST3697, ST3707, ST3708, ST3709, ST3710, ST3755, ST3756, ST3757 and ST4509. The ETEC isolates displayed different levels of antimicrobial resistance. Eight isolates were resistant to only one drug, three isolates-to two drugs, one isolate-to three drugs, two isolates-to four antibacterials, and only one isolate to each of the five, six and ten antibacterials simultaneously. Genetic determinants of the resistance to beta-lactams and other classes of antibacterials on the ETEC genomes were identified. There are blaTEM (n = 10, blaCTX-M-15 (n = 1, class 1 integron (n = 3 carrying resistance cassettes to aminoglycosides and

Molecular Characterization of Enterotoxin-Producing Escherichia coli Collected in 2011-2012, Russia.

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Kartsev, Nikolay N; Fursova, Nadezhda K; Pachkunov, Dmitry M; Bannov, Vasiliy A; Eruslanov, Boris V; Svetoch, Edward A; Dyatlov, Ivan A

2015-01-01

Enterotoxin-producing Escherichia coli (ETEC) are one of the main causative agents of diarrhea in children especially in developing countries and travel diarrhoea in adults. Pathogenic properties of ETEC associated with their ability to produce a heat-stable (ST) and/or heat-labile (LT) enterotoxins, as well as adhesins providing bacterial adhesion to intestinal epithelial cells. This study presents the molecular characterization of the ETEC isolates collected from the Central and Far-Eastern regions of Russia in 2011-2012. It was shown that all ETEC under study (n=18) had the heat-labile enterotoxin-coding operon elt, and had no the genes of the heat-stable enterotoxin operon est. DNA sequencing revealed two types of nucleotide exchanges in the eltB gene coding subunit B of LT in isolates collected from Cherepovets city (Central region, Russia) and Vladivostok city (Far-East region, Russia). Only one ETEC strain carried genes cfaA, cfaB, cfaC and cfaD coding adhesion factor CFA/I. Expression of LT in four ETEC isolates in the agglutination reaction was detected using a latex test-system. The isolates were assigned to serogroups O142 (n = 6), О6 (n = 4), О25 (n = 5), О26 (n = 2), and O115 (n = 1). Genotyping showed that they belonged to an earlier described sequence-type ST4 (n = 3) as well as to 11 novel sequence-types ST1043, ST1312, ST3697, ST3707, ST3708, ST3709, ST3710, ST3755, ST3756, ST3757 and ST4509. The ETEC isolates displayed different levels of antimicrobial resistance. Eight isolates were resistant to only one drug, three isolates-to two drugs, one isolate-to three drugs, two isolates-to four antibacterials, and only one isolate to each of the five, six and ten antibacterials simultaneously. Genetic determinants of the resistance to beta-lactams and other classes of antibacterials on the ETEC genomes were identified. There are blaTEM (n = 10), blaCTX-M-15 (n = 1), class 1 integron (n = 3) carrying resistance cassettes to aminoglycosides and

Characterization of Staphylococcus aureus strains and evidence for the involvement of non-classical enterotoxin genes in food poisoning outbreaks.

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Ciupescu, Laurentiu-Mihai; Auvray, Frederic; Nicorescu, Isabela Madalina; Meheut, Thomas; Ciupescu, Veronica; Lardeux, Anne-Laure; Tanasuica, Rodica; Hennekinne, Jacques-Antoine

2018-06-05

To an increasing extent, molecular and genetic characterization is now used to investigate foodborne outbreaks. The aim of this study was to seek molecular links among coagulase-positive staphylococci (CPS) isolated from three recent food poisoning outbreaks in Romania using polymerase chain reaction and pulsed-field gel electrophoresis (PFGE) techniques. Nineteen CPS isolates were identified as Staphylococcus aureus by detection of the 23S rDNA gene. Among them, 15 carried at least one staphylococcal enterotoxin-encoding gene (se). The Calarași outbreak strains grouped in pulsotype 2 and were sed/sej/ser-positive, whereas the Arad outbreak strains clustered in pulsotype 17 and were either sed/seg/sei/sej/ser- or seg/sei-positive. The Pitești outbreak strains clustered in pulsotype 1 and, surprisingly, possessed only one enterotoxin gene, i.e. seh. Similar to other European countries, the seh gene has been identified with increasing frequency in Romanian outbreaks; this highlights the importance of considering the application of methods recommended for staphylococcal enterotoxin regulation in Europe.

Heat-Labile Enterotoxin: Beyond G M1 Binding

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Benjamin Mudrak

2010-06-01

Full Text Available Enterotoxigenic Escherichia coli (ETEC is a significant source of morbidity and mortality worldwide. One major virulence factor released by ETEC is the heat-labile enterotoxin LT, which is structurally and functionally similar to cholera toxin. LT consists of five B subunits carrying a single catalytically active A subunit. LTB binds the monosialoganglioside GM1, the toxin’s host receptor, but interactions with A-type blood sugars and E. coli lipopolysaccharide have also been identified within the past decade. Here, we review the regulation, assembly, and binding properties of the LT B-subunit pentamer and discuss the possible roles of its numerous molecular interactions.

Monkey Feeding Assay for Testing Emetic Activity of Staphylococcal Enterotoxin.

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Seo, Keun Seok

2016-01-01

Staphylococcal enterotoxins (SEs) are unique bacterial toxins that cause gastrointestinal toxicity as well as superantigenic activity. Since systemic administration of SEs induces superantigenic activity leading to toxic shock syndrome that may mimic enterotoxic activity of SEs such as vomiting and diarrhea, oral administration of SEs in the monkey feeding assay is considered as a standard method to evaluate emetic activity of SEs. This chapter summarizes and discusses practical considerations of the monkey feeding assay used in studies characterizing classical and newly identified SEs.

Uncoupling of T Cell Receptor Zeta Chain Function during the Induction of Anergy by the Superantigen, Staphylococcal Enterotoxin A

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William D. Cornwell

2010-06-01

Full Text Available Staphylococcus aureus enterotoxins have immunomodulatory properties. In this study, we show that Staphylococcal enterotoxin A (SEA induces a strong proliferative response in a murine T cell clone independent of MHC class II bearing cells. SEA stimulation also induces a state of hypo-responsiveness (anergy. We characterized the components of the T cell receptor (TCR during induction of anergy by SEA. Most interestingly, TCR zeta chain phosphorylation was absent under SEA anergizing conditions, which suggests an uncoupling of zeta chain function. We characterize here a model system for studying anergy in the absence of confounding costimulatory signals.

The olive compound 4-hydroxytyrosol inactivates Staphyloccoccus aureus bacteria and Staphylococcal enterotoxin A (SEA)

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The foodborne pathogen Staphylococcus aureus produces the virulent staphylococcal enterotoxin A (SEA), a single chain protein which consists of 233 amino acid residues with a molecular weight of 27,078 Da. SEA is a superantigen that is reported to contribute to animal (mastitis) and human (emesis, ...

Staphylococcal enterotoxins in the Etiopathogenesis of Mucosal Autoimmunity within the Gastrointestinal Tract

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MaryAnn Principato

2014-04-01

Full Text Available The staphylococcal enterotoxins (SEs are the products of Staphylococcus aureus and are recognized as the causative agents of classical food poisoning in humans following the consumption of contaminated food. While illness evoked by ingestion of the SE or its producer organism in tainted food are often self-limited, our current understanding regarding the evolution of S. aureus provokes the utmost concern. The organism and its associated toxins, has been implicated in a wide variety of disease states including infections of the skin, heart, sinuses, inflammatory gastrointestinal disease, toxic shock, and Sudden Infant Death Syndrome. The intricate relationship between the various subsets of immunocompetent T cells and accessory cells and the ingested material found within the gastrointestinal tract present daunting challenges to the maintenance of immunologic homeostasis. Dysregulation of the intricate balances within this environment has the potential for extreme consequences within the host, some of which are long-lived. The focus of this review is to evaluate the relevance of staphylococcal enterotoxin in the context of mucosal immunity, and the underlying mechanisms that contribute to the pathogenesis of gastrointestinal autoimmune disease.

Lung and pharyngeal abscess caused by enterotoxin G- and I-producing Staphylococcus aureus.

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Barnett, S Y; Hattotuwa, K L; Teare, L

2012-05-01

We report a particularly serious case of extensive meticillin sensitive Staphylococcal lung and pharyngeal abscess. Our patient had no significant risk factors for severe infection. The detection of enterotoxin G and I here suggest that when present together, these toxins work synergistically to produce a more virulent strain of Staphylococcus aureus. Copyright © 2011. Published by Elsevier Ltd.

Effects of nisin and temperature on survival, growth, and enterotoxin production characteristics of psychrotrophic Bacillus cereus in beef gravy.

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Beuchat, L R; Clavero, M R; Jaquette, C B

1997-05-01

The presence of psychrotrophic enterotoxigenic Bacillus cereus in ready-to-serve meats and meat products that have not been subjected to sterilization treatment is a public health concern. A study was undertaken to determine the survival, growth, and diarrheal enterotoxin production characteristics of four strains of psychrotrophic B. cereus in brain heart infusion (BHI) broth and beef gravy as affected by temperature and supplementation with nisin. A portion of unheated vegetative cells from 24-h BHI broth cultures was sensitive to nisin as evidenced by an inability to form colonies on BHI agar containing 10 micrograms of nisin/ml. Heat-stressed cells exhibited increased sensitivity to nisin. At concentrations as low as 1 microgram/ml, nisin was lethal to B. cereus, the effect being more pronounced in BHI broth than in beef gravy. The inhibitory effect of nisin (1 microgram/ml) was greater on vegetative cells than on spores inoculated into beef gravy and was more pronounced at 8 degrees C than at 15 degrees C. Nisin, at a concentration of 5 or 50 micrograms/ml, inhibited growth in gravy inoculated with vegetative cells and stored at 8 or 15 degrees C, respectively, for 14 days. Growth of vegetative cells and spores of B. cereus after an initial period of inhibition is attributed to loss of activity of nisin. One of two test strains produced diarrheal enterotoxin in gravy stored at 8 or 15 degrees C within 9 or 3 days, respectively. Enterotoxin production was inhibited in gravy supplemented with 1 microgram of nisin/ml and stored at 8 degrees C for 14 days; 5 micrograms of nisin/ml was required for inhibition at 15 degrees C. Enterotoxin was not detected in gravy in which less than 5.85 log10 CFU of B. cereus/ml had grown. Results indicate that as little as 1 microgram of nisin/ml may be effective in inhibiting or retarding growth of and diarrheal enterotoxin production by vegetative cells and spores of psychrotrophic B. cereus in beef gravy at 8 degrees C, a

Prevalence of enterotoxin-encoding genes and antimicrobial resistance in coagulase-negative and coagulase-positive Staphylococcus isolates from black pudding

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Tiane Martin de Moura

2012-10-01

Full Text Available INTRODUCTION: Staphylococcal species are pathogens that are responsible for outbreaks of foodborne diseases. The aim of this study was to investigate the prevalence of enterotoxin-genes and the antimicrobial resistance profile in staphylococcus coagulase-negative (CoNS and coagulasepositive (CoPS isolates from black pudding in southern Brazil. METHODS: Two hundred typical and atypical colonies from Baird-Parker agar were inoculated on mannitol salt agar. Eighty-two mannitol-positive staphylococci were submitted to conventional biochemical tests and antimicrobial susceptibility profiling. The presence of coagulase (coa and enterotoxin (se genes was investigated by polymerase chain reaction. RESULTS: The isolates were divided into 2 groups: 75.6% (62/82 were CoNS and 24.4% (20/82 were CoPS. The biochemical tests identified 9 species, of which Staphylococcus saprophyticus (37.8% and Staphylococcus carnosus (15.9% were the most prevalent. Antimicrobial susceptibility tests showed resistance phenotypes to antibiotics widely administered in humans, such as gentamicin, tetracycline, chloramphenicol, and erythromycin. The coa gene was detected in 19.5% (16/82 of the strains and 4 polymorphic DNA fragments were observed. Five CoNS isolates carrying the coa gene were submitted for 16S rRNA sequencing and 3 showed similarity with CoNS. Forty strains were positive for at least 1 enterotoxin-encoding gene, the genes most frequently detected were sea (28.6% and seb (27.5%. CONCLUSIONS: The presence of antimicrobial resistant and enterotoxin-encoding genes in staphylococci isolates from black pudding indicated that this fermented food may represent a potential health risk, since staphylococci present in food could cause foodborne diseases or be a possible route for the transfer of antimicrobial resistance to humans.

Inhibitory effects of food additives derived from polyphenols on staphylococcal enterotoxin A production and biofilm formation by Staphylococcus aureus.

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Shimamura, Yuko; Hirai, Chikako; Sugiyama, Yuka; Shibata, Masaharu; Ozaki, Junya; Murata, Masatsune; Ohashi, Norio; Masuda, Shuichi

2017-12-01

In this study, we examined the inhibitory effects of 14 food additives derived from polyphenol samples on staphylococcal enterotoxin A (SEA) production and biofilm formation by Staphylococcus aureus. Tannic acid AL (TA), Purephenon 50 W (PP) and Polyphenon 70A (POP) at 0.25 mg/mL and Gravinol®-N (GN), Blackcurrant polyphenol AC10 (BP), and Resveratrol-P5 (RT) at 1.0 mg/mL significantly decreased SEA production by S. aureus C-29 (p Food additives derived from polyphenols have viability to be used as a means to inhibit the enterotoxin production and control the biofilm formation of foodborne pathogens.

Use of inactivated E.Coli enterotoxins to enhance respiratory mucosal adjuvanticity during vaccination in swine

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In order to augment responses to respiratory vaccines in swine, various adjuvants were intranasally co-administered with an antigen to pigs. Detoxified E. coli enterotoxins LTK63 and LTR72 enhanced mucosal and systemic immunity to the model peptide, exhibiting their efficacy as mucosal adjuvants for...

A Single-Stranded DNA Aptamer That Selectively Binds to Staphylococcus aureus Enterotoxin B

OpenAIRE

DeGrasse, Jeffrey A.

2012-01-01

The bacterium Staphylococcus aureus is a common foodborne pathogen capable of secreting a cocktail of small, stable, and strain-specific, staphylococcal enterotoxins (SEs). Staphylococcal food poisoning (SFP) results when improperly handled food contaminated with SEs is consumed. Gastrointestinal symptoms of SFP include emesis, diarrhea and severe abdominal pain, which manifest within hours of ingesting contaminated food. Immuno-affinity based methods directly detect, identify, and quantify s...

Plant-Derived Polyphenols Interact with Staphylococcal Enterotoxin A and Inhibit Toxin Activity

OpenAIRE

Shimamura, Yuko; Aoki, Natsumi; Sugiyama, Yuka; Tanaka, Takashi; Murata, Masatsune; Masuda, Shuichi

2016-01-01

This study was performed to investigate the inhibitory effects of 16 different plant-derived polyphenols on the toxicity of staphylococcal enterotoxin A (SEA). Plant-derived polyphenols were incubated with the cultured Staphylococcus aureus C-29 to investigate the effects of these samples on SEA produced from C-29 using Western blot analysis. Twelve polyphenols (0.1-0.5 mg/mL) inhibited the interaction between the anti-SEA antibody and SEA. We examined whether the polyphenols could directly i...

Contamination of beef products with staphylococcal classical enterotoxins in Egypt and Saudi Arabia

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Shawish, Reyad R.

2016-04-01

Full Text Available Food-borne pathogens are of high concern for public health and food safety. food poisoning is one of the most economically devastating types of food poisoning globally. The purpose of this study was to detect staphylococcal classical enterotoxins (SEs in processed beef from Kingdom of Saudi Arabia (KSA and Egypt. In the present investigation a total of 250 random processed meat samples (50 each of minced meat, beef burger, beef sausage, beef kofta and beef luncheon were collected from different super markets in the study area. Using conventional cultural methods, samples were cultured for isolation and identification of . Multiplex PCR was used to detect SEs of the classical type SEA, SEB, SEC and SED from isolates.The percentage presence of in minced meat, beef burger, beef sausage, beef kofta and beef luncheon was 38%, 22%, 30%, 32% and 12%, respectively. Multiplex PCR indicated that all examined samples contain different types of classical staphylococcal enterotoxins and only minced meat samples contained all four types of toxins. Multiplex PCR is efficient in detection of SEs from food and may be used in tracing of toxins to promote food hygiene. Implications of contamination of processed meat to food hygiene in the study area are highlighted.

Suppression of colorectal tumorigenesis by recombinant Bacteroides fragilis enterotoxin-2 in vivo

OpenAIRE

Lv, You; Ye, Tao; Wang, Hui-Peng; Zhao, Jia-Ying; Chen, Wen-Jie; Wang, Xin; Shen, Chen-Xia; Wu, Yi-Bin; Cai, Yuan-Kun

2017-01-01

AIM To evaluate the impact of recombinant Bacteroides fragilis enterotoxin-2 (BFT-2, or Fragilysin) on colorectal tumorigenesis in mice induced by azoxymethane/dextran sulfate sodium (AOM/DSS). METHODS Recombinant proBFT-2 was expressed in Escherichia coli strain Rosetta (DE3) and BFT-2 was obtained and tested for its biological activity via colorectal adenocarcinoma cell strains SW-480. Seventy C57BL/6J mice were randomly divided into a blank (BC; n = 10), model (AD; n = 20), model + low-dos...

GM1 erythroimmunoassay for detection and titration of Escherichia coli heat-labile enterotoxin.

OpenAIRE

Germani, Y; Bégaud, E; Guesdon, J L; Moreau, J P

1986-01-01

A GM1 ganglioside erythroimmunoassay for the detection of heat-labile Escherichia coli enterotoxin (LT) was developed for use in poorly equipped laboratories in developing countries. This assay is based on the immunological similarity between Vibrio cholerae toxin and LT and uses cholera toxin antiserum and sheep anti-rabbit immunoglobulin covalently coupled to sheep erythrocytes as conjugate. This assay has the following advantages over other currently available techniques: the reagents it u...

Lactobacillus zeae protects Caenorhabditis elegans from enterotoxigenic Escherichia coli-caused death by inhibiting enterotoxin gene expression of the pathogen.

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Mengzhou Zhou

Full Text Available BACKGROUND: The nematode Caenorhabditis elegans has become increasingly used for screening antimicrobials and probiotics for pathogen control. It also provides a useful tool for studying microbe-host interactions. This study has established a C. elegans life-span assay to preselect probiotic bacteria for controlling K88(+ enterotoxigenic Escherichia coli (ETEC, a pathogen causing pig diarrhea, and has determined a potential mechanism underlying the protection provided by Lactobacillus. METHODOLOGY/PRINCIPAL FINDINGS: Life-span of C. elegans was used to measure the response of worms to ETEC infection and protection provided by lactic acid-producing bacteria (LAB. Among 13 LAB isolates that varied in their ability to protect C. elegans from death induced by ETEC strain JG280, Lactobacillus zeae LB1 offered the highest level of protection (86%. The treatment with Lactobacillus did not reduce ETEC JG280 colonization in the nematode intestine. Feeding E. coli strain JFF4 (K88(+ but lacking enterotoxin genes of estA, estB, and elt did not cause death of worms. There was a significant increase in gene expression of estA, estB, and elt during ETEC JG280 infection, which was remarkably inhibited by isolate LB1. The clone with either estA or estB expressed in E. coli DH5α was as effective as ETEC JG280 in killing the nematode. However, the elt clone killed only approximately 40% of worms. The killing by the clones could also be prevented by isolate LB1. The same isolate only partially inhibited the gene expression of enterotoxins in both ETEC JG280 and E. coli DH5α in-vitro. CONCLUSIONS/SIGNIFICANCE: The established life-span assay can be used for studies of probiotics to control ETEC (for effective selection and mechanistic studies. Heat-stable enterotoxins appeared to be the main factors responsible for the death of C. elegans. Inhibition of ETEC enterotoxin production, rather than interference of its intestinal colonization, appears to be the

A probability model for enterotoxin production of Bacillus cereus as a function of pH and temperature

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Bacillus cereus is frequently isolated from a variety of foods including vegetables, dairy products, meat, and other raw and processed foods. The bacterium is capable of producing enterotoxin and emetic toxin that can cause severe nausea, vomiting and diarrhea. The objectives of this study were to a...

Presence of Classical Enterotoxin Genes, agr Typing, Antimicrobial Resistance, and Genetic Diversity of Staphylococcus aureus from Milk of Cows with Mastitis in Southern Brazil.

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Kroning, Isabela S; Iglesias, Mariana A; Mendonça, Karla S; Lopes, Graciela V; Silva, Wladimir P

2018-05-01

Staphylococcus aureus is a common causative agent of bovine mastitis in dairy cows and commonly associated with foodborne disease outbreaks. The aim of this study was to evaluate the presence of enterotoxin genes, agr typing, antimicrobial resistance, and genetic diversity of S. aureus isolated from milk of cows with mastitis in dairy farms from southern Brazil. Results showed that 7 (22.6%) of 31 S. aureus isolates were positive for enterotoxin genes. Specifically, the genes encoding for enterotoxins A ( n = 4), C ( n = 2), and B ( n = 1) were detected. Isolates belonging to the agr group III (10 of 31, 32.2%) and agr group I (7 of 31, 22.5%) were the most common. To our knowledge, this is the first report of both agr I and III in the same S. aureus isolate from milk of cows with bovine mastitis. The antimicrobial resistance test showed that 54% of the isolates were multiresistant to antimicrobial agents. The macrorestriction analysis produced 16 different major SmaI pulsed-field gel electrophoresis patterns, with up to two subpatterns. Moreover, the presence of some S. aureus clones in a distinct area was observed. Although this study characterized a limited number of S. aureus isolates, the presence of classical enterotoxin genes and resistance to multiple antimicrobial agents reinforces the importance of this microorganism to animal and human health. In addition, similar genetic profiles have been identified in distinct geographic areas, suggesting clonal dissemination of S. aureus in dairy herds from southern Brazil.

The staphylococcal enterotoxin (SE) family: SEB and siblings.

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Krakauer, Teresa; Stiles, Bradley G

2013-11-15

Staphylococcus aureus plays an important role in numerous human cases of food poisoning, soft tissue, and bone infections, as well as potentially lethal toxic shock. This common bacterium synthesizes various virulence factors that include staphylococcal enterotoxins (SEs). These protein toxins bind directly to major histocompatibility complex class II on antigen-presenting cells and specific Vβ regions of T-cell receptors, resulting in potentially life-threatening stimulation of the immune system. Picomolar concentrations of SEs ultimately elicit proinflammatory cytokines that can induce fever, hypotension, multi-organ failure, and lethal shock. Various in vitro and in vivo models have provided important tools for studying the biological effects of, as well as potential vaccines/therapeutics against, the SEs. This review succinctly presents known physical and biological properties of the SEs, including various intervention strategies. In particular, SEB will often be portrayed as per biodefense concerns dating back to the 1960s.

Magnetic Bead and Fluorescent Silica Nanoparticles Based Optical Immunodetection of Staphylococcal Enterotoxin B (SEB in Bottled Water

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Shiva K. RASTOGI

2009-10-01

Full Text Available Staphylococcal enterotoxins (SEs are a major cause of food-borne diseases, most commonly SEs assayed immunologically with ELISA. An immunoassay based on fluorescein dye doped silica dioxide nanoparticles (F-SiNPs and magnetic bead (MB is described here for the detection of staphylococcal enterotoxin B (SEB. F-SiNPs have unique optical properties which make them attractive for biosensing. The water-in-oil (W/O reverse microemulsion method was used for the synthesis of F-SiNPs (~ 95 nm of diameter. The F-SiNPs were characterized using SEM, TEM and FTIR spectroscopy. The detection of SEB is preformed in PBS buffer, and bottled drinking water using sandwich immunoassay format. Target analytes were captured using MBs modified with the antigen-specific “capture” antibody, and detected using F-SiNP labeled secondary antigen-specific antibody. We report a limit of detection down to 1 ng/mL SEB spiked sample in less than 2 hr assay time using fluorocount method. This study demonstrates the bio warfare agent SEB capture by magnetic beads and detection using F-SiNPs.

Sulfasalazine Attenuates Staphylococcal Enterotoxin B-Induced Immune Responses

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Teresa Krakauer

2015-02-01

Full Text Available Staphylococcal enterotoxin B (SEB and related exotoxins are important virulence factors produced by Staphylococcus aureus as they cause human diseases such as food poisoning and toxic shock. These toxins bind directly to cells of the immune system resulting in hyperactivation of both T lymphocytes and monocytes/macrophages. The excessive release of proinflammatory cytokines from these cells mediates the toxic effects of SEB. This study examined the inhibitory activities of an anti-inflammatory drug, sulfasalazine, on SEB-stimulated human peripheral blood mononuclear cells (PBMC. Sulfasalazine dose-dependently inhibited tumor necrosis factor α, interleukin 1 (IL-1 β, IL-2, IL-6, interferon γ (IFNγ, and various chemotactic cytokines from SEB-stimulated human PBMC. Sulfasalazine also potently blocked SEB-induced T cell proliferation and NFκB activation. These results suggest that sulfasalazine might be useful in mitigating the toxic effects of SEB by blocking SEB-induced host inflammatory cascade and signaling pathways.

Antibiotic resistance and enterotoxin genes in Staphylococcus sp. isolates from polluted water in Southern Brazil.

Science.gov (United States)

Basso, Ana P; Martins, Paula D; Nachtigall, Gisele; Van Der Sand, Sueli; De Moura, Tiane M; Frazzon, Ana Paula G

2014-12-01

The aim of this study was to evaluate the species distribution, antibiotic-resistance profile and presence of enterotoxin (SE) genes in staphylococci isolated from the Dilúvio stream in South Brazil. Eighty-eight staphylococci were identified, 93.18% were identified as coagulase-negative (CNS) and 6.82% coagulase-positive (CPS). Fourteen Staphylococcus species were detected and the most frequently were Staphylococcus cohnii (30.48%) and S. haemolyticus (21.95%). Resistance to erythromycin was verified in 37.50% of the strains, followed by 27.27% to penicillin, 12.50% to clindamycin, 6.81% to trimethoprim-sulfamethoxazole, 5.68% to chloramphenicol and 2.27% to norfloxacin. None of the investigated strains showed gentamicin and ciprofloxacin resistance. The strains were tested for the presence of sea, seb, sec, sed and see genes by PCR and only CNS strains (43.18%) showed positive results to one or more SE genes. The scientific importance of our results is due to the lack of data about these topics in polluted waters in Brazil. In conclusion, polluted waters from the Dilúvio stream may constitute a reservoir for disseminating antibiotic-resistance and enterotoxin into the community. In addition, the detection of staphylococci in the polluted waters of the Dilúvio stream indicated a situation of environmental contamination and poor sanitation conditions.

Antibiotic resistance and enterotoxin genes in Staphylococcus sp. isolates from polluted water in Southern Brazil

OpenAIRE

BASSO, ANA P.; MARTINS, PAULA D.; NACHTIGALL, GISELE; SAND, SUELI VAN DER; MOURA, TIANE M. DE; FRAZZON, ANA PAULA G.

2014-01-01

The aim of this study was to evaluate the species distribution, antibiotic-resistance profile and presence of enterotoxin (SE) genes in staphylococci isolated from the Dilúvio stream in South Brazil. Eighty-eight staphylococci were identified, 93.18% were identified as coagulase-negative (CNS) and 6.82% coagulase-positive (CPS). Fourteen Staphylococcus species were detected and the most frequently were Staphylococcus cohnii (30.48%) and S. haemolyticus (21.95%). Resistance to erythromycin was...

Comparison of toxicity neutralization-, ELISA- and PCR tests for typing of Clostridium perfringens and detection of the enterotoxin gene by PCR

DEFF Research Database (Denmark)

Møller, Kristian; Ahrens, Peter

1996-01-01

A polymerase chain reaction (PCR) was developed for the specific amplification of a part of each of the five Clostridium perfringens toxin genes: alpha (alpha), beta (beta), epsilon (epsilon), iota (iota), and enterotoxin (CPE). While the toxicity neutralization test (TNT) only showed limited...

Detection of enterotoxins and genotyping of Staphylococcus aureus strains isolated from Isfahan Educational Hospital, Iran

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Seyed Asghar Havaei

2017-10-01

Full Text Available Background and aims: Staphylococcus aureus is known as one of the most important nosocomial pathogens, which may lead to several infections. The aim of this study was determining the enterotoxins A, C, and TSST-1 and molecular characterization of S. aureus strains with PFGE and MLST typing methods. Materials and methods: In the present study during the sixmonths sampling, fifty S. aureus strains were isolated from patients admitted to Al-Zahra university hospital. Antimicrobial susceptibility testing, Multiplex PCR for detection of enterotoxin A, C and TSST-1, pulse field gel electrophoresis (PFGE and multilocus sequence typing (MLST were used for molecular typing. Results: In antibiogram the highest and lowest percentage of resistance was belonged to tetracycline and rifampin respectively. Multiplex PCR indicated that 30% of the strains harbored sea and 34% harbored sec genes. However, only 4% of our collected isolates had tsst gene. In PFGE method analysis on all S. aureus strains, a total of 19 different patterns were identified. Nine various sequence types in 27 selected S. aureus isolates were identified by MLST. Conclusions: Present study indicates a possible higher variability among our S. aureus strains by two different molecular typing methods; nevertheless four main common types (CT1, CT7, CT9, and CT11 with at least one toxin genes were determined.

Antibiotic resistance and enterotoxin genes in Staphylococcus sp. isolates from polluted water in Southern Brazil

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ANA P. BASSO

2014-12-01

Full Text Available The aim of this study was to evaluate the species distribution, antibiotic-resistance profile and presence of enterotoxin (SE genes in staphylococci isolated from the Dilúvio stream in South Brazil. Eighty-eight staphylococci were identified, 93.18% were identified as coagulase-negative (CNS and 6.82% coagulase-positive (CPS. Fourteen Staphylococcus species were detected and the most frequently were Staphylococcus cohnii (30.48% and S. haemolyticus (21.95%. Resistance to erythromycin was verified in 37.50% of the strains, followed by 27.27% to penicillin, 12.50% to clindamycin, 6.81% to trimethoprim-sulfamethoxazole, 5.68% to chloramphenicol and 2.27% to norfloxacin. None of the investigated strains showed gentamicin and ciprofloxacin resistance. The strains were tested for the presence of sea, seb, sec, sed and see genes by PCR and only CNS strains (43.18% showed positive results to one or more SE genes. The scientific importance of our results is due to the lack of data about these topics in polluted waters in Brazil. In conclusion, polluted waters from the Dilúvio stream may constitute a reservoir for disseminating antibiotic-resistance and enterotoxin into the community. In addition, the detection of staphylococci in the polluted waters of the Dilúvio stream indicated a situation of environmental contamination and poor sanitation conditions.

Elevated Enterotoxin A Expression and Formation in Staphylococcus aureus and Its Association with Prophage Induction

OpenAIRE

Cao, Rong; Zeaki, Nikoleta; Wallin-Carlquist, Nina; Skandamis, Panagiotis N.; Schelin, Jenny; Rådström, Peter

2012-01-01

Staphylococcus aureus strains producing the bacteriophage-encoded staphylococcal enterotoxin A (SEA) were divided into two groups, high- and low-SEA-producing strains, based on the amount of SEA produced. After growth under favorable conditions in batch cultures, 10 of the 21 strains tested produced more than 1,000 ng/ml SEA, and 9 strains produced less than 10 ng/ml SEA; two enterotoxigenic strains, MRSA252 and Newman, produced intermediate levels of SEA (around 450 ng/ml). The differences i...

Prevalence and Genetic Characteristics of Staphylococcus aureus and Staphylococcus argenteus Isolates Harboring Panton-Valentine Leukocidin, Enterotoxins, and TSST-1 Genes from Food Handlers in Myanmar.

Science.gov (United States)

Aung, Meiji Soe; San, Thida; Aye, Mya Mya; Mya, San; Maw, Win Win; Zan, Khin Nyein; Htut, Wut Hmone Win; Kawaguchiya, Mitsuyo; Urushibara, Noriko; Kobayashi, Nobumichi

2017-08-04

Asymptomatic carriers of toxigenic Staphylococcus aureus are potential source of diseases, including food poisoning. Toxigenic potential and genetic traits of colonizing S. aureus were investigated for 563 healthy food handlers in Myanmar. Carriage of S. aureus was found in 110 individuals (19.5%), and a total of 144 S. aureus isolates were recovered from nasal cavities (110 isolates) and hands (34 isolates). Panton-Valentine leucocidin genes ( pvl ) were detected in 18 isolates (12.5%), among which 11 isolates were classified into coa -VIa, agr type III, and ST1930 (CC96) that had been also detected in pvl -positive clinical isolates in Myanmar. A pvl -positive, ST2250 nasal isolate was identified as S. argenteus , a novel coagulase-positive staphylococcus species. Toxic shock syndrome toxin-1 (TSST-1) gene was detected in five pvl -negative isolates. All of the 144 isolates harbored at least one of the 21 enterotoxin(-like) gene(s). The most prevalent enterotoxin(-like) gene was selw (98%), followed by selx (97%), sei (28%), sely (28%), sem (26%), sel (24%), and sea and sec (22% each). Considerable genetic diversity with five groups was detected for selw . The present study revealed the relatively high rate of pvl , as well as the wide distribution of enterotoxin(-like) genes among colonizing S. aureus in Myanmar.

Estrogen inhibits chloride secretion caused by cholera and Escherichia coli enterotoxins in female rat distal colon.

OpenAIRE

Alzamora, Rodrigo; O'Mahony, Fiona; Harvey, Brian J

2011-01-01

Excessive Cl(-) secretion is the driving force for secretory diarrhea. 17β-Estradiol has been shown to inhibit Cl(-) secretion in rat distal colon through a nongenomic pathway. We examined whether 17β-estradiol inhibits Cl(-) secretion in an animal model of secretory diarrhea and the downstream effectors involved. The effect of 17β-estradiol on cholera toxin and heat-stable enterotoxin induced Cl(-) secretion in rat colonic mucosal sheets was studied by current-voltage clamping. Selective per...

Type II heat-labile enterotoxins from 50 diverse Escherichia coli isolates belong almost exclusively to the LT-IIc family and may be prophage encoded.

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Michael G Jobling

Full Text Available Some enterotoxigenic Escherichia coli (ETEC produce a type II heat-labile enterotoxin (LT-II that activates adenylate cyclase in susceptible cells but is not neutralized by antisera against cholera toxin or type I heat-labile enterotoxin (LT-I. LT-I variants encoded by plasmids in ETEC from humans and pigs have amino acid sequences that are ≥ 95% identical. In contrast, LT-II toxins are chromosomally encoded and are much more diverse. Early studies characterized LT-IIa and LT-IIb variants, but a novel LT-IIc was reported recently. Here we characterized the LT-II encoding loci from 48 additional ETEC isolates. Two encoded LT-IIa, none encoded LT-IIb, and 46 encoded highly related variants of LT-IIc. Phylogenetic analysis indicated that the predicted LT-IIc toxins encoded by these loci could be assigned to 6 subgroups. The loci corresponding to individual toxins within each subgroup had DNA sequences that were more than 99% identical. The LT-IIc subgroups appear to have arisen by multiple recombinational events between progenitor loci encoding LT-IIc1- and LT-IIc3-like variants. All loci from representative isolates encoding the LT-IIa, LT-IIb, and each subgroup of LT-IIc enterotoxins are preceded by highly-related genes that are between 80 and 93% identical to predicted phage lysozyme genes. DNA sequences immediately following the B genes differ considerably between toxin subgroups, but all are most closely related to genomic sequences found in predicted prophages. Together these data suggest that the LT-II loci are inserted into lambdoid type prophages that may or may not be infectious. These findings raise the possibility that production of LT-II enterotoxins by ETEC may be determined by phage conversion and may be activated by induction of prophage, in a manner similar to control of production of Shiga-like toxins by converting phages in isolates of enterohemmorhagic E. coli.

Cloning Sequencing and Structural Manipulation of the Enterotoxin D and E Genes from Staphylococcus aureus

Science.gov (United States)

1990-07-01

Iandolo, J.J. and R.K. Tweten. 1988. Purification of staphylococcal enterotoxin. In. Methods of Enzymology, (N.O. Kaplan , S. Harshman, Eds.) volume 165...N C V 9 L G 0 K 1 S P A K I C T S N 0 149 "T7AACTGG&&C&hICG&clhht:7caflCATGTAATAa~aI ACca CAACaCAA 942 L K D G D K L Z L I G T P r D H K V N 0 H L..L

Studies on the mechanism of action of enterotoxin-induced fluid secretion in the gut

International Nuclear Information System (INIS)

Schirgi-Degen, A.

1992-12-01

The mechanism of action of Clostridium difficile enterotoxin A (CA), of Escherichia coli enterotoxin (STa) and of cholera toxin (CT), which are known to cause severe diarrhea, were studied in a preparation of ligated jejunal loops of anesthetized rats in vivo. The toxins were administered intraluminally. Pharmacological agents, which were tested for their potency to influence toxin-related effects, were administered subcutaneously. Net fluid transport was determined gravimetrically, prostaglandin (PG) E 2 -output into the lumen, cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP) contents in the mucosa were measured by radioimmunoassay, serotonin-(5-HT)-output into the lumen was determined by high performance liquid chromatography. The histopathological effects of CA and CT were examined by light- and scanning electron microscopy. All three toxins caused net fluid secretion (FS). 5-HT 2 -(ketanserin) and 5-HT 3 -receptor antagonists (tropisetron, ondansetron, granisetron) dose-dependently reduced or abolished CT- and STa-induced net FS, CA-induced net FS was not influenced. Indomethacin reduced CA-, CT- and STa-induced net FS. Elevation of PGE 2 -output occurred after exposure to CA and CT and was reduced by indomethacin. CA caused severe histopathological lesions and also CT time-dependently caused morphological changes, which may take part in the secretory response. It is concluded that 5-HT, using both 5-HT 2 - and 5-HT 3 -receptors, mediates CT- and STa, but not CA-induced FS. PGE 2 is involved in FS caused by all three toxins. CAMP and cGMP are presumedly no causative mediators of toxin-induced FS

Influence of holding temperature on the growth and survival of Salmonella spp. and Staphylococcus aureus and the production of staphylococcal enterotoxin in egg products.

Science.gov (United States)

Yang, S E; Yu, R C; Chou, C C

2001-01-22

In this study, growth and survival of Salmonella spp. and Staphylococcus aureus in steamed egg and scrambled egg held at 5, 18, 22, 37, 55 and 60 degrees C are investigated. The production of staphylococcal enterotoxin in steamed egg is also examined. Results reveal that Salmonella spp. and Staph. aureus in the egg products multiply best at 37 degrees C, followed closely by 22 and 18 degrees C. Neither pathogen showed growth in the egg products held at 5 degrees C. Initial inoculation dose, holding temperature and holding time affected the population of both organisms found in the egg products. Staphylococcal enterotoxin A (SEA) and B (SEB) are detected only in the egg products held at 37 or 22 degrees C. After holding at 37 degrees C for 36 h, scrambled egg inoculated with ca. 5.0 log cfu/g Staph. aureus contains the highest levels of SEA (> 64 ng/g) and SEB (> 64 ng/g). Although Salmonella spp. and Staph. aureus grow better in steamed eggs than in scrambled eggs, production of staphylococcal enterotoxin, in general, was higher in scrambled eggs than in steamed eggs. On the other hand, a repaid destruction of the test organisms in steamed eggs held at 60 degrees C was observed. Holding the steamed eggs at 60 degrees C, Salmonella spp. and Staph. aureus with an initial population of ca. 5.9 and 5.6 log cfu/g, respectively, reduced to a non-detectable level in 1 h.

Noncontiguous Finished Genome Sequence of Staphylococcus aureus KLT6, a Staphylococcal Enterotoxin B-Positive Strain Involved in a Food Poisoning Outbreak in Switzerland

Science.gov (United States)

Tobes, Raquel; Manrique, Marina; Brozynska, Marta; Stephan, Roger; Pareja, Eduardo

2013-01-01

We present the first complete genome sequence of a Staphylococcus aureus strain assigned to clonal complex 12. The strain was isolated in a food poisoning outbreak due to contaminated potato salad in Switzerland in 2009, and it produces staphylococcal enterotoxin B. PMID:23704175

Comparison of Genotypes and Enterotoxin Genes Between Staphylococcus aureus Isolates from Blood and Nasal Colonizers in a Korean Hospital

Science.gov (United States)

Peck, Kyong Ran; Baek, Jin Yang; Song, Jae-Hoon

2009-01-01

In this study, we investigated the genetic background of 70 Staphylococcus aureus isolates (36 methicillin-resistant S. aureus [MRSA] and 34 methicillin-susceptible S. aureus [MSSA]) obtained from blood at a Korean tertiary-care hospital, using spa typing, multilocus sequence typing, and SCCmec typing. In addition, the prevalence of enterotoxin (sea, seb, sec, sed, see, seg, seh, sei, and sek), tst, and pvl genes among the samples was assessed via polymerase chain reaction, and the results were compared with those of 95 isolates of S. aureus obtained from nasal swabs. All MRSA isolates from blood, except one, belonged to three major clones: sequence type (ST)5-MRSA-II, ST72-MRSA-II (or IVA), and ST239-MRSA-III, among which ST5-MRSA-II was the predominant clone. The prevalence of enterotoxin genes in the S. aureus isolates obtained from blood differed significantly from those from the nasal swabs for the sea, seb, sec, and seh gene. In particular, the seb and sec genes were detected exclusively in the MRSA isolates of ST5 or spa-CC002, thereby suggesting the co-adaptation of virulence genes with the genetic background and their contribution to biological fitness. PMID:19654937

A single-stranded DNA aptamer that selectively binds to Staphylococcus aureus enterotoxin B.

Science.gov (United States)

DeGrasse, Jeffrey A

2012-01-01

The bacterium Staphylococcus aureus is a common foodborne pathogen capable of secreting a cocktail of small, stable, and strain-specific, staphylococcal enterotoxins (SEs). Staphylococcal food poisoning (SFP) results when improperly handled food contaminated with SEs is consumed. Gastrointestinal symptoms of SFP include emesis, diarrhea and severe abdominal pain, which manifest within hours of ingesting contaminated food. Immuno-affinity based methods directly detect, identify, and quantify several SEs within a food or clinical sample. However, the success of these assays depends upon the availability of a monoclonal antibody, the development of which is non-trivial and costly. The current scope of the available immuno-affinity based methods is limited to the classical SEs and does not encompass all of the known or emergent SEs. In contrast to antibodies, aptamers are short nucleic acids that exhibit high affinity and specificity for their targets without the high-costs and ethical concerns of animal husbandry. Further, researchers may choose to freely distribute aptamers and develop assays without the proprietary issues that increase the per-sample cost of immuno-affinity assays. This study describes a novel aptamer, selected in vitro, with affinity to staphylococcal enterotoxin B (SEB) that may be used in lieu of antibodies in SE detection assays. The aptamer, designated APT(SEB1), successfully isolates SEB from a complex mixture of SEs with extremely high discrimination. This work sets the foundation for future aptamer and assay development towards the entire family of SEs. The rapid, robust, and low-cost identification and quantification of all of the SEs in S. aureus contaminated food is essential for food safety and epidemiological efforts. An in vitro generated library of SE aptamers could potentially allow for the comprehensive and cost-effective analysis of food samples that immuno-affinity assays currently cannot provide.

A single-stranded DNA aptamer that selectively binds to Staphylococcus aureus enterotoxin B.

Directory of Open Access Journals (Sweden)

Jeffrey A DeGrasse

Full Text Available The bacterium Staphylococcus aureus is a common foodborne pathogen capable of secreting a cocktail of small, stable, and strain-specific, staphylococcal enterotoxins (SEs. Staphylococcal food poisoning (SFP results when improperly handled food contaminated with SEs is consumed. Gastrointestinal symptoms of SFP include emesis, diarrhea and severe abdominal pain, which manifest within hours of ingesting contaminated food. Immuno-affinity based methods directly detect, identify, and quantify several SEs within a food or clinical sample. However, the success of these assays depends upon the availability of a monoclonal antibody, the development of which is non-trivial and costly. The current scope of the available immuno-affinity based methods is limited to the classical SEs and does not encompass all of the known or emergent SEs. In contrast to antibodies, aptamers are short nucleic acids that exhibit high affinity and specificity for their targets without the high-costs and ethical concerns of animal husbandry. Further, researchers may choose to freely distribute aptamers and develop assays without the proprietary issues that increase the per-sample cost of immuno-affinity assays. This study describes a novel aptamer, selected in vitro, with affinity to staphylococcal enterotoxin B (SEB that may be used in lieu of antibodies in SE detection assays. The aptamer, designated APT(SEB1, successfully isolates SEB from a complex mixture of SEs with extremely high discrimination. This work sets the foundation for future aptamer and assay development towards the entire family of SEs. The rapid, robust, and low-cost identification and quantification of all of the SEs in S. aureus contaminated food is essential for food safety and epidemiological efforts. An in vitro generated library of SE aptamers could potentially allow for the comprehensive and cost-effective analysis of food samples that immuno-affinity assays currently cannot provide.

Sensitive chemiluminescence immunoassay for staphylococcal enterotoxin C1 based on the use of dye-encapsulated mesoporous silica nanoparticles

International Nuclear Information System (INIS)

Tao, Liang; Zhang, Chunmei; Sun, Yuanjie; Jin, Boquan; Yang, Kun; Li, Xiaohua; Zhang, Zhujun; Zhang, Jinpeng; Yan, Kuocheng

2016-01-01

A chemiluminescent immunoassay for the staphylococcal enterotoxin C1 (SEC1) based on the use of dye-encapsulated mesoporous silica nanoparticles (m-SiNPs) as a label is described. The dyes are retained in the m-SiNPs via strong hydrophobic interactions. The assay comprises the following steps: (a) Microplates coated with antibody against SEC1 are filled with sample upon which the SEC antigen will be bound to the surface; (b) following a washing step, secondary antibody linked to m-SiNPs (that were covalently labeled with rhodamine 6G and fluorescein) were added to form the sandwich complex; (c) after another washing step, bis(2,4,6-trichlorophenyl) oxalate, H_2O_2 and imidazole are added to generate chemiluminescence whose intensity is proportional to the number of m-SiNPs and thus to the number of antigen (SEC) molecules. It is found that the use of functionalized m-SiNPs strongly amplifies the signal. Enterotoxin SEC1 can be detected by this method in the 0.025 to 2 ng⋅mL"-"1 concentration range, the detection limit is 19 pg⋅mL"-"1 (at 3σ), and the relative standard deviation (for 11 parallel measurements at a 1 ng⋅mL"-"1 level) is 4.6 %. The use of an automated chemiluminescence analyzer further improves detection. (author)

Effects of nisin and temperature on survival, growth, and enterotoxin production characteristics of psychrotrophic Bacillus cereus in beef gravy.

OpenAIRE

Beuchat, L R; Clavero, M R; Jaquette, C B

1997-01-01

The presence of psychrotrophic enterotoxigenic Bacillus cereus in ready-to-serve meats and meat products that have not been subjected to sterilization treatment is a public health concern. A study was undertaken to determine the survival, growth, and diarrheal enterotoxin production characteristics of four strains of psychrotrophic B. cereus in brain heart infusion (BHI) broth and beef gravy as affected by temperature and supplementation with nisin. A portion of unheated vegetative cells from...

Detection of Staphylococcus aureus enterotoxins in sheep cheese and dairy desserts by multiplex PCR technique.

Science.gov (United States)

Ertas, Nurhan; Gonulalan, Zafer; Yildirim, Yeliz; Kum, Erhan

2010-08-15

The aim of this study was to investigate the presence of Staphylococcus aureus (S. aureus) and staphylococcal enterotoxins (SEs) genes in sheep cheese and dairy dessert samples by multiplex PCR (mPCR) technique. A total of 150 samples were analyzed consisting of 50 dairy dessert samples and 100 sheep cheese. Coagulase positive staphylococci (CPS) were found in 86 (57.3%) out of 150 analyzed samples. S. aureus were isolated from 60 (60%), 26 (52%) of sheep cheese and from of dairy desserts, respectively. Five suspected colonies were tested from each sheep cheese and dairy dessert samples for phenotypic and genotypic characterizations. A total of 430 isolates from the 86 positive samples were investigated in this study. Eighty (18.6%) isolates were characterized as S. aureus. The enterotoxin genes (sea, seb, sec, sed) were found in 13 (3.02%) out of 80 isolates. From cheese isolates, sea, seb and sed were detected in 5 (1.6%), 2 (0.6%), 1 (0.3%), respectively. From dairy dessert isolates, sea, sec and sed were detected in 3 (2.3%), 1 (0.76%), 1 (0.76%), respectively. The presence of SEs was identified in 12 (2.8%) out of 80 isolates by using ELISA technique. It was determined that these SEs had a distribution of 7 (1.6%) SEA, 2 (0.46%) SEB, 1 (0.23%) SEC, and 2 (0.46%) SED. SEs were found in 7 (2.3%) cheese and 5 (3.8%) dairy dessert isolates. In conclusion, S.aureus and their SEs were found to be present in sheep cheese and dairy desserts in this study. It is emphasized that the presence of S. aureus and their SEs genes in sheep cheese and dairy desserts may be regarded as a potential risk for human health. Copyright 2010 Elsevier B.V. All rights reserved.

Comparative Effects of Food Preservatives on the Production of Staphylococcal Enterotoxin I from Staphylococcus aureus Isolate

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Yanying Zhao

2017-01-01

Full Text Available Staphylococcal enterotoxin I (SEI is associated with staphylococcal food poisoning, but little is known about different food preservatives on the production of SEI. In this study, the effect of different food preservatives (sodium nitrite, polylysine, chitosan, and tea catechin on the bacteria growth, sei gene expression, and extracellular SEI production of Staphylococcus aureus isolate H4 was detected in tryptone soya broth (TSB culture. Our results showed that all of these preservatives depressed S. aureus H4 growth and the order of inhibitory effect was 0.8 g/L tea catechin > 6 g/L chitosan > 0.25 g/L polylysine > 0.4 g/L tea catechin > 0.15 g/L sodium nitrite. Furthermore, 0.25 g/L polylysine or 0.15 g/L sodium nitrite did not significantly alter sei gene transcription, while 6 g/L chitosan obviously increased the relative mRNA level of sei gene expression. 0.4 g/L tea catechin remarkably inhibited sei gene transcription. In addition, 0.15 g/L sodium nitrite and 6 g/L chitosan significantly enhanced SEI secretion. 0.25 g/L polylysine, especially 0.4 g/L tea catechin, sharply inhibited the level of SEI secretion. The results indicated that tea catechin not only suppressed Staphylococcus aureus growth, but also inhibited SEI production and secretion, suggesting that tea catechin may be better than sodium nitrite, polylysine, or chitosan for keeping the food from the contamination of SEI. These investigations would be useful for food industry to provide safer food products due to S. aureus enterotoxins-related control strategy.

Coupling immunomagnetic separation on magnetic beads with matrix-assisted laser desorption ionization-time of flight mass spectrometry for detection of Staphylococcal Enterotoxin B

Czech Academy of Sciences Publication Activity Database

Schlosser, G.; Kačer, P.; Kuzma, Marek; Szilágyi, Z.; Sorrentino, Z.; Manzo, C.; Pizzano, R.; Malorni, L.; Poscfalvi, G.

2007-01-01

Roč. 73, č. 21 (2007), s. 6945-6952 ISSN 0099-2240 Grant - others:IT(IT) project SIFORTI (MIUR-CNR, Legge 449/97); XE(XE) 94.05.09.103, ARINCO 94.IT.16.028 Institutional research plan: CEZ:AV0Z50200510 Keywords : mass spectrometry * staphylococcal enterotoxin b * bacterial protein toxins Subject RIV: EE - Microbiology, Virology Impact factor: 4.004, year: 2007

Influence of carvacrol and thymol on the physiological attributes, enterotoxin production and surface characteristics of Staphylococcus aureus strains isolated from foods

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E.L. Souza

2013-01-01

Full Text Available This study evaluated the influence of the phenolic compounds carvacrol (CAR and thymol (THY on some physiological characteristics and on the modulation of the secretion of some staphylococcal virulence factors, that is, coagulase and enterotoxin. This study also investigated possible mechanisms for the establishment of the anti-staphylococcal activity of these compounds. Sublethal concentrations (0.3 and 0.15 µL/mL of CAR and THY inhibited the activity of the enzymes coagulase and lipase and led to a decrease in salt tolerance. At the tested sublethal concentrations, both CAR and THY led to a total suppression of enterotoxin production. The loss of a 260-nm-absorbing material and an efflux of potassium ions occurred immediately after the addition of CAR and THY at 0.6 and 1.2 µL/mL and increased up to 120 min of exposure. Electron microscopy of cells exposed to CAR and THY (0.6 µL/mL revealed that individual cells appeared to be deformed, with projections of cellular material. The observations of leakage of cellular material and an altered cell surface suggest that gross damage to a cell's cytoplasmic membrane, which results in a disruption in protein secretion, could be responsible for the anti-staphylococcal properties of CAR and THY.

Effect of different ionizing radiation doses and dose rates, using Cobalt-60 and electrons beam sources, on the staphylococcal enterotoxin inoculated in mechanically deboned chicken meat

International Nuclear Information System (INIS)

Pomarico Neto, Walter; Brito, Poliana de Paula; Azevedo, Heliana de; Roque, Claudio Vitor; Fukuma, Henrique Takuji; Kodama, Yasko; Miya, Norma Terugo Nago; Pereira, Jose Luiz

2011-01-01

The purpose of food irradiation is the destruction of present pathogenic microorganisms and the increase of shelf life of foods. To achieve this process, the source of cobalt-60 and the electron accelerator can be used. The mechanically deboned chicken meat (MDCM) is used for the production of traditional meat products, and it may come to present pathogenic microorganisms such as staphylococcus aureus, a bacterium that produces enterotoxin, which causes food poisoning. The objective of this study is to analyze the effect of ionizing irradiation with different doses and dose rates, deriving from different radiation sources, on staphylococcal enterotoxin type B (SEB) in the MDCM. 50 g samples of MDCM were prepared in a batch of 6 kg of MDCM. The samples were contaminated, with the exception of the control, with SEB in amounts of about 100 ng. Then they were conditioned in a transparent bag made of low density polyethylene, frozen at -18±1 deg C overnight and irradiated in these conditions with doses of 0.0 kGy (control), 1.5 kGy and 3.0 kGy, and with three different dose rates, both in the Cobalt-60 and the electron accelerator. The experiments were conducted in quintuplicate. The SEB extraction from the MDCM was performed according to the protocol recommended by the manufacturer of the kit VIDAS Staph Enterotoxin II (bioMerrieux). The principle of mass balance was used to determine the actual amount of SEB removed by irradiation. The treatment that presented the best results was the one with a dose of 1.5 kGy, high dose rate of the electron accelerator. (author)

Effect of different ionizing radiation doses and dose rates, using Cobalt-60 and electrons beam sources, on the staphylococcal enterotoxin inoculated in mechanically deboned chicken meat

Energy Technology Data Exchange (ETDEWEB)

Pomarico Neto, Walter; Brito, Poliana de Paula; Azevedo, Heliana de; Roque, Claudio Vitor; Fukuma, Henrique Takuji, E-mail: pbrito@cnen.gov.br, E-mail: hazevedo@cnen.gov.br, E-mail: cvroque@cnen.gov.br, E-mail: htfukuma@cnen.gov.br [Brazilian Nuclear Energy Commission (LAPOC/CNEN), Pocos de Caldas, MG (Brazil); Kodama, Yasko, E-mail: ykodama@ipen.br [Nuclear and Energy Research Institute (IPEN/CNEN-SP), Sao Paulo, SP (Brazil); Miya, Norma Terugo Nago; Pereira, Jose Luiz, E-mail: miya@fea.unicamp.br, E-mail: pereira@fea.unicamp.br [Campinas State University (UNICAMP), SP (Brazil). Dept. of Food Sciences

2011-07-01

The purpose of food irradiation is the destruction of present pathogenic microorganisms and the increase of shelf life of foods. To achieve this process, the source of cobalt-60 and the electron accelerator can be used. The mechanically deboned chicken meat (MDCM) is used for the production of traditional meat products, and it may come to present pathogenic microorganisms such as staphylococcus aureus, a bacterium that produces enterotoxin, which causes food poisoning. The objective of this study is to analyze the effect of ionizing irradiation with different doses and dose rates, deriving from different radiation sources, on staphylococcal enterotoxin type B (SEB) in the MDCM. 50 g samples of MDCM were prepared in a batch of 6 kg of MDCM. The samples were contaminated, with the exception of the control, with SEB in amounts of about 100 ng. Then they were conditioned in a transparent bag made of low density polyethylene, frozen at -18{+-}1 deg C overnight and irradiated in these conditions with doses of 0.0 kGy (control), 1.5 kGy and 3.0 kGy, and with three different dose rates, both in the Cobalt-60 and the electron accelerator. The experiments were conducted in quintuplicate. The SEB extraction from the MDCM was performed according to the protocol recommended by the manufacturer of the kit VIDAS Staph Enterotoxin II (bioMerrieux). The principle of mass balance was used to determine the actual amount of SEB removed by irradiation. The treatment that presented the best results was the one with a dose of 1.5 kGy, high dose rate of the electron accelerator. (author)

From Escherichia coli heat-stable enterotoxin to mammalian endogenous guanylin hormones

International Nuclear Information System (INIS)

Lima, A.A.M.; Fonteles, M.C.

2014-01-01

The isolation of heat-stable enterotoxin (STa) from Escherichia coli and cholera toxin from Vibrio cholerae has increased our knowledge of specific mechanisms of action that could be used as pharmacological tools to understand the guanylyl cyclase-C and the adenylyl cyclase enzymatic systems. These discoveries have also been instrumental in increasing our understanding of the basic mechanisms that control the electrolyte and water balance in the gut, kidney, and urinary tracts under normal conditions and in disease. Herein, we review the evolution of genes of the guanylin family and STa genes from bacteria to fish and mammals. We also describe new developments and perspectives regarding these novel bacterial compounds and peptide hormones that act in electrolyte and water balance. The available data point toward new therapeutic perspectives for pathological features such as functional gastrointestinal disorders associated with constipation, colorectal cancer, cystic fibrosis, asthma, hypertension, gastrointestinal barrier function damage associated with enteropathy, enteric infection, malnutrition, satiety, food preferences, obesity, metabolic syndrome, and effects on behavior and brain disorders such as attention deficit, hyperactivity disorder, and schizophrenia

From Escherichia coli heat-stable enterotoxin to mammalian endogenous guanylin hormones

Energy Technology Data Exchange (ETDEWEB)

Lima, A.A.M. [Universidade Federal do Ceará, Departamento de Fisiologia e Farmacologia, Escola de Medicina, Instituto de Biomedicina, Unidade de Pesquisas Clínicas, Fortaleza, CE, Brasil, Unidade de Pesquisas Clínicas, Instituto de Biomedicina, Departamento de Fisiologia e Farmacologia, Escola de Medicina, Universidade Federal do Ceará, Fortaleza, CE (Brazil); Fonteles, M.C. [Universidade Federal do Ceará, Departamento de Fisiologia e Farmacologia, Escola de Medicina, Instituto de Biomedicina, Unidade de Pesquisas Clínicas, Fortaleza, CE, Brasil, Unidade de Pesquisas Clínicas, Instituto de Biomedicina, Departamento de Fisiologia e Farmacologia, Escola de Medicina, Universidade Federal do Ceará, Fortaleza, CE (Brazil); Universidade Estadual do Ceará, Instituto de Ciências Biomédicas, Fortaleza, CE, Brasil, Instituto de Ciências Biomédicas, Universidade Estadual do Ceará, Fortaleza, CE (Brazil)

2014-03-03

The isolation of heat-stable enterotoxin (STa) from Escherichia coli and cholera toxin from Vibrio cholerae has increased our knowledge of specific mechanisms of action that could be used as pharmacological tools to understand the guanylyl cyclase-C and the adenylyl cyclase enzymatic systems. These discoveries have also been instrumental in increasing our understanding of the basic mechanisms that control the electrolyte and water balance in the gut, kidney, and urinary tracts under normal conditions and in disease. Herein, we review the evolution of genes of the guanylin family and STa genes from bacteria to fish and mammals. We also describe new developments and perspectives regarding these novel bacterial compounds and peptide hormones that act in electrolyte and water balance. The available data point toward new therapeutic perspectives for pathological features such as functional gastrointestinal disorders associated with constipation, colorectal cancer, cystic fibrosis, asthma, hypertension, gastrointestinal barrier function damage associated with enteropathy, enteric infection, malnutrition, satiety, food preferences, obesity, metabolic syndrome, and effects on behavior and brain disorders such as attention deficit, hyperactivity disorder, and schizophrenia.

Identification and Characterization of a New Enterotoxin Produced by Clostridium perfringens Isolated from Food Poisoning Outbreaks.

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Daisuke Irikura

Full Text Available There is a strain of Clostridium perfringens, W5052, which does not produce a known enterotoxin. We herein report that the strain W5052 expressed a homologue of the iota-like toxin components sa and sb of C. spiroforme, named Clostridium perfringens iota-like enterotoxin, CPILE-a and CPILE-b, respectively, based on the results of a genome sequencing analysis and a systematic protein screening. In the nicotinamide glyco-hydrolase (NADase assay the hydrolysis activity was dose-dependently increased by the concentration of rCPILE-a, as judged by the mass spectrometry analysis. In addition, the actin monomer of the lysates of Vero and L929 cells were radiolabeled in the presence of [32P]NAD and rCPILE-a. These findings indicated that CPILE-a possesses ADP-ribosylation activity. The culture supernatant of W5052 facilitated the rounding and killing of Vero and L929 cells, but the rCPILE-a or a non-proteolyzed rCPILE-b did not. However, a trypsin-treated rCPILE-b did. Moreover, a mixture of rCPILE-a and the trypsin-treated rCPILE-b enhanced the cell rounding and killing activities, compared with that induced by the trypsin-treated rCPILE-b alone. The injection of the mixture of rCPILE-a and the trypsin-treated rCPILE-b into an ileum loop of rabbits evoked the swelling of the loop and accumulation of the fluid dose-dependently, suggesting that CPILE possesses enterotoxic activity. The evidence presented in this communication will facilitate the epidemiological, etiological, and toxicological studies of C. perfringens food poisoning, and also stimulate studies on the transfer of the toxins' gene(s among the Genus Clostridium.

Short communication: Pasteurization as a means of inactivating staphylococcal enterotoxins A, B, and C in milk.

Science.gov (United States)

Necidova, Lenka; Bogdanovicova, Katerina; Harustiakova, Danka; Bartova, Katerina

2016-11-01

Our aim was to assess the effect of pasteurization temperature on inactivation of staphylococcal enterotoxins (SE). Milk samples were inoculated with log 4.38 to 5.18cfu/mL of 40 different Staphylococcus aureus strains having the ability to produce types A, B, or C SE and incubated at 37°C for 24h to develop SE. This incubation was followed by heat treatment for 15 s at 72, 85, and 92°C. Samples were analyzed for Staph. aureus count by plate method and, specifically, for SE presence. An enzyme-linked immunofluorescent assay on a MiniVIDAS analyzer (bioMérieux, Marcy l'Étoile, France) was used to detect SE, which were determined semiquantitatively based on test values. The Staph. aureus count in milk before pasteurization did not affect the amount of SE. Before pasteurization, SEB was detected in the lowest amount compared with other SE types. Staphylococcal enterotoxins were markedly reduced with pasteurization and inactivated at pasteurization temperatures to an extent depending on the amount in the sample before pasteurization. After pasteurization at 72°C, SE were detected in 87.5% of samples (35/40), after pasteurization at 85°C in 52.5% of samples (21/40), and after pasteurization at 92°C in 45.0% of samples (18/40). We determined that SE may still persist in milk even when Staph. aureus bacteria are inactivated through pasteurization. Although pasteurization may partially inactivate SE in milk, a key measure in the prevention of staphylococcal enterotoxicosis linked to pasteurized milk consumption is to avoid any cold chain disruption during milk production and processing. Copyright © 2016 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Staphylococcus aureus and staphylococcal enterotoxin A in breaded chicken products: detection and behavior during the cooking process.

Science.gov (United States)

Pepe, Olimpia; Blaiotta, Giuseppe; Bucci, Francesca; Anastasio, Marilena; Aponte, Maria; Villani, Francesco

2006-11-01

In this study we examined the presence of Staphylococcus aureus and staphylococcal enterotoxin A (SEA) in 20 industrial breaded chicken products obtained from different retail butchers and supermarket stores in Italy. The levels of contamination in the products analyzed were quite low, although the pH values and water activities (a(w)) in the samples considered were in ranges favorable for S. aureus growth. As demonstrated by phenotypic and molecular characterization, in spite of the high percentage of coagulase-positive Staphylococcus strains, only three strains could be referred to the species S. aureus. Moreover, all the strains were negative in PCR assays targeting staphylococcal enterotoxin genes (seA to seE, seG to seJ, and seM to seO), as well as the toxic shock syndrome toxin 1 gene, and no SEA was detected in the retail breaded chicken samples analyzed by a reversed passive latex agglutination assay or by Western blotting. Hence, we evaluated the thermal resistance of two strains of SEA-producing S. aureus in a laboratory-scale preparation of precooked breaded chicken cutlets. The heat treatment employed in the manufacture determined the inactivation of S. aureus cells, but the preformed SEA remained active during product storage at 4 degrees C. The presence of the staphylococci and, in particular, of S. aureus in the retail breaded chicken products analyzed is a potential health risk for consumers since the pH and a(w) values of these kinds of products are favorable for S. aureus growth. The thermal process used during their manufacture can limit staphylococcal contamination but cannot eliminate preformed toxins.

Subinhibitory concentrations of thymol reduce enterotoxins A and B and alpha-hemolysin production in Staphylococcus aureus isolates.

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Jiazhang Qiu

Full Text Available BACKGROUND: Targeting bacterial virulence factors is now gaining interest as an alternative strategy to develop new types of anti-infective agents. It has been shown that thymol, when used at low concentrations, can inhibit the TSST-1 secretion in Staphylococcus aureus. However, there are no data on the effect of thymol on the production of other exotoxins (e.g., alpha-hemolysin and enterotoxins by S. aureus. METHODOLOGY/PRINCIPAL FINDINGS: Secretion of alpha-hemolysin, SEA and SEB in both methicillin-sensitive and methicillin-resistant S. aureus isolates cultured with graded subinhibitory concentrations of thymol was detected by immunoblot analysis. Hemolysin and tumor necrosis factor (TNF release assays were performed to elucidate the biological relevance of changes in alpha-hemolysin, SEA and SEB secretion induced by thymol. In addition, the influence of thymol on the transcription of hla, sea, and seb (the genes encoding alpha-hemolysin, SEA and SEB, respectively was analyzed by quantitative RT-PCR. Thymol inhibited transcription of hla, sea and seb in S. aureus, resulting in a reduction of alpha-hemolysin, SEA and SEB secretion and, thus, a reduction in hemolytic and TNF-inducing activities. CONCLUSIONS/SIGNIFICANCE: Subinhibitory concentrations of thymol decreased the production of alpha-hemolysin, SEA and SEB in both MSSA and MRSA in a dose-dependent manner. These data suggest that thymol may be useful for the treatment of S. aureus infections when used in combination with beta-lactams and glycopeptide antibiotics, which induce expression of alpha-hemolysin and enterotoxins at subinhibitory concentrations. Furthermore, the structure of thymol may potentially be used as a basic structure for development of drugs aimed against these bacterial virulence factors.

CodY Promotes Sporulation and Enterotoxin Production by Clostridium perfringens Type A Strain SM101.

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Li, Jihong; Freedman, John C; Evans, Daniel R; McClane, Bruce A

2017-03-01

Clostridium perfringens type D strains cause enterotoxemia and enteritis in livestock via epsilon toxin production. In type D strain CN3718, CodY was previously shown to increase the level of epsilon toxin production and repress sporulation. C. perfringens type A strains producing C. perfringens enterotoxin (CPE) cause human food poisoning and antibiotic-associated diarrhea. Sporulation is critical for C. perfringens type A food poisoning since spores contribute to transmission and resistance in the harsh food environment and sporulation is essential for CPE production. Therefore, the current study asked whether CodY also regulates sporulation and CPE production in SM101, a derivative of C. perfringens type A food-poisoning strain NCTC8798. An isogenic codY -null mutant of SM101 showed decreased levels of spore formation, along with lower levels of CPE production. A complemented strain recovered wild-type levels of both sporulation and CPE production. When this result was coupled with the earlier results obtained with CN3718, it became apparent that CodY regulation of sporulation varies among different C. perfringens strains. Results from quantitative reverse transcriptase PCR analysis clearly demonstrated that, during sporulation, codY transcript levels remained high in SM101 but rapidly declined in CN3718. In addition, abrB gene expression patterns varied significantly between codY -null mutants of SM101 and CN3718. Compared to the levels in their wild-type parents, the level of abrB gene expression decreased in the CN3718 codY -null mutant strain but significantly increased in the SM101 codY -null mutant strain, demonstrating CodY-dependent regulation differences in abrB expression between these two strains. This difference appears to be important since overexpression of the abrB gene in SM101 reduced the levels of sporulation and enterotoxin production, supporting the involvement of AbrB repression in regulating C. perfringens sporulation. Copyright © 2017

Formation of cereulide and enterotoxins by Bacillus cereus in fermented African locust beans

DEFF Research Database (Denmark)

Thorsen, Line; Azokpota, Paulin; Munk Hansen, Bjarne

2011-01-01

Afitin, iru and sonru are three spontaneously fermented African locust bean Benin condiments. The fermentation processes are exothermic, with temperatures mostly being above 40 °C. A total of 19 predominant Bacillus cereus isolates from afitin, iru and sonru, were investigated. The enterotoxin...... genes nhe (A, B, C) were present in all 19 isolates, the hbl (A, C, D) in one (afitin), and the cytK gene in three isolates (afitin). Levels of cytotoxicity to Vero cells and NheA production in BHI-broth was within the range of known diarrheal outbreak strains. Autoclaved cooked African locust beans...... inoculated with emetic (cereulide producing) B. cereus Ba18H2/RIF supported growth at 25, 30 and 40 °C with highly different maximum cereulide productions of 6 ± 5, 97 ± 3 and 0.04 ± 0.02 μg/g beans, respectively (48 h). For non-autoclaved cooked beans inoculated with 2, 4 and 6 log10 B. cereus Ba18H2/RIF...

A regenerable piezoelectric immunosensor on the basis of electropolymerized polypyrrole for highly selective detection of Staphylococcal Enterotoxin A in foodstuffs

International Nuclear Information System (INIS)

Karaseva, Nadezhda; Ermolaeva, Tatyana

2015-01-01

Piezoelectric immunosensors have been developed for the determination of trace concentrations of Staphylococcal enterotoxin A (SEA) in foodstuff. Antibodies against SEA were covalently immobilized on an electropolymerized polypyrrole substrate on a gold electrode which warrants increased stability of the biolayer and sensitive detection. The calibration plot is linear in the 1–80 ng∙mL -1 SEA concentration range, the limit of detection is 0.4 ng∙mL -1 , and the time of analysis is <30 min. The sensor was successfully applied to quantify SEA in chicken meat and milk. (author)

Selection of staphylococcal enterotoxin B (SEB)-binding peptide using phage display technology

International Nuclear Information System (INIS)

Soykut, Esra Acar; Dudak, Fahriye Ceyda; Boyaci, Ismail Hakki

2008-01-01

In this study, peptides were selected to recognize staphylococcal enterotoxin B (SEB) which cause food intoxication and can be used as a biological war agent. By using commercial M13 phage library, single plaque isolation of 38 phages was done and binding affinities were investigated with phage-ELISA. The specificities of the selected phage clones showing high affinity to SEB were checked by using different protein molecules which can be found in food samples. Furthermore, the affinities of three selected phage clones were determined by using surface plasmon resonance (SPR) sensors. Sequence analysis was realized for three peptides showing high binding affinity to SEB and WWRPLTPESPPA, MNLHDYHRLFWY, and QHPQINQTLYRM amino acid sequences were obtained. The peptide sequence with highest affinity to SEB was synthesized with solid phase peptide synthesis technique and thermodynamic constants of the peptide-SEB interaction were determined by using isothermal titration calorimetry (ITC) and compared with those of antibody-SEB interaction. The binding constant of the peptide was determined as 4.2 ± 0.7 x 10 5 M -1 which indicates a strong binding close to that of antibody

Staphylococcal enterotoxin C2 promotes osteogenesis and suppresses osteoclastogenesis of human mesenchymal stem cells.

Science.gov (United States)

Fu, Wei-ming; Zhu, Xiao; Wang, Hua; Wei-Mao Wang; Chen, Ju-yu; Liang, Yan; Zhang, Jin-fang; Kung, Hsiang-fu

2014-03-10

As a super-antigen, staphylococcal enterotoxin C2 (SEC2) stimulates the release of massive inflammatory cytokines such as interferon-gamma (IFN-γ), interleukin-1 (IL-1) and interleukin-2 (IL-2) which are documented to implicate osteoblast differentiation. In the present study, SEC2 was found to significantly improve the osteoblast differentiation by up-regulating BMP2 and Runx2/Cbfa1 expression. Interferon (IFN)-inducible gene IFI16, a co-activator of Runx2/Cbfa1, was also activated by SEC2 in the osteoblast differentiation. In addition, exogenous introduction of SEC2 stimulated OPG expression and suppressed RANKL, suggesting suppression of osteoclastogenesis in hMSCs. Therefore, our results displayed that SEC2 plays an important role in the commitment of MSC to the osteoblast and it might be a potential new therapeutic candidate for bone regeneration. Copyright © 2013 Elsevier Inc. All rights reserved.

Association and dissociation of Escherichia coli heat-stable enterotoxin from rat brush border membrane receptors

International Nuclear Information System (INIS)

Cohen, M.B.; Thompson, M.R.; Overmann, G.J.; Giannella, R.A.

1987-01-01

Escherichia coli heat-stable enterotoxin (ST) binds to receptors on rat intestinal cells and brush border membranes (BBM). We devised experiments to examine the reversibility of ST binding. We found that both 125 I-labeled ST and native ST were spontaneously dissociable from the BBM receptor. Radiolabeled ST bound to BBM was also dissociated by the addition of avid goat anti-ST antiserum. Furthermore, using a computer program for analysis of ligand binding, we calculated an apparent Ka of 10(8) liters/mol from competitive inhibition and saturation-binding data. This is significantly lower than the value previously reported by others. Our findings, of a lower Ka and a reversible ST-binding process, suggest that a therapeutic strategy of removing bound ST from its receptor or competing with the enterocyte receptor for unbound ST might be successful in terminating ST-induced secretion

Comparative Genomics and Identification of an Enterotoxin-Bearing Pathogenicity Island, SEPI-1/SECI-1, in Staphylococcus epidermidis Pathogenic Strains.

Science.gov (United States)

Argemi, Xavier; Nanoukon, Chimène; Affolabi, Dissou; Keller, Daniel; Hansmann, Yves; Riegel, Philippe; Baba-Moussa, Lamine; Prévost, Gilles

2018-02-25

Staphylococcus epidermidis is a leading cause of nosocomial infections, majorly resistant to beta-lactam antibiotics, and may transfer several mobile genetic elements among the members of its own species, as well as to Staphylococcus aureus ; however, a genetic exchange from S. aureus to S. epidermidis remains controversial. We recently identified two pathogenic clinical strains of S. epidermidis that produce a staphylococcal enterotoxin C3-like (SEC) similar to that by S. aureus pathogenicity islands. This study aimed to determine the genetic environment of the SEC-coding sequence and to identify the mobile genetic elements. Whole-genome sequencing and annotation of the S. epidermidis strains were performed using Illumina technology and a bioinformatics pipeline for assembly, which provided evidence that the SEC-coding sequences were located in a composite pathogenicity island that was previously described in the S. epidermidis strain FRI909, called SePI-1/SeCI-1, with 83.8-89.7% nucleotide similarity. Various other plasmids were identified, particularly p_3_95 and p_4_95, which carry antibiotic resistance genes ( hsrA and dfrG , respectively), and share homologies with SAP085A and pUSA04-2-SUR11, two plasmids described in S. aureus . Eventually, one complete prophage was identified, ΦSE90, sharing 30 out of 52 coding sequences with the Acinetobacter phage vB_AbaM_IME200. Thus, the SePI-1/SeCI-1 pathogenicity island was identified in two pathogenic strains of S. epidermidis that produced a SEC enterotoxin causing septic shock. These findings suggest the existence of in vivo genetic exchange from S. aureus to S. epidermidis .

Comparative Genomics and Identification of an Enterotoxin-Bearing Pathogenicity Island, SEPI-1/SECI-1, in Staphylococcus epidermidis Pathogenic Strains

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Xavier Argemi

2018-02-01

Full Text Available Staphylococcus epidermidis is a leading cause of nosocomial infections, majorly resistant to beta-lactam antibiotics, and may transfer several mobile genetic elements among the members of its own species, as well as to Staphylococcus aureus; however, a genetic exchange from S. aureus to S. epidermidis remains controversial. We recently identified two pathogenic clinical strains of S. epidermidis that produce a staphylococcal enterotoxin C3-like (SEC similar to that by S. aureus pathogenicity islands. This study aimed to determine the genetic environment of the SEC-coding sequence and to identify the mobile genetic elements. Whole-genome sequencing and annotation of the S. epidermidis strains were performed using Illumina technology and a bioinformatics pipeline for assembly, which provided evidence that the SEC-coding sequences were located in a composite pathogenicity island that was previously described in the S. epidermidis strain FRI909, called SePI-1/SeCI-1, with 83.8–89.7% nucleotide similarity. Various other plasmids were identified, particularly p_3_95 and p_4_95, which carry antibiotic resistance genes (hsrA and dfrG, respectively, and share homologies with SAP085A and pUSA04-2-SUR11, two plasmids described in S. aureus. Eventually, one complete prophage was identified, ΦSE90, sharing 30 out of 52 coding sequences with the Acinetobacter phage vB_AbaM_IME200. Thus, the SePI-1/SeCI-1 pathogenicity island was identified in two pathogenic strains of S. epidermidis that produced a SEC enterotoxin causing septic shock. These findings suggest the existence of in vivo genetic exchange from S. aureus to S. epidermidis.

Multiple-Strain Approach and Probabilistic Modeling of Consumer Habits in Quantitative Microbial Risk Assessment: A Quantitative Assessment of Exposure to Staphylococcal Enterotoxin A in Raw Milk.

Science.gov (United States)

Crotta, Matteo; Rizzi, Rita; Varisco, Giorgio; Daminelli, Paolo; Cunico, Elena Cosciani; Luini, Mario; Graber, Hans Ulrich; Paterlini, Franco; Guitian, Javier

2016-03-01

Quantitative microbial risk assessment (QMRA) models are extensively applied to inform management of a broad range of food safety risks. Inevitably, QMRA modeling involves an element of simplification of the biological process of interest. Two features that are frequently simplified or disregarded are the pathogenicity of multiple strains of a single pathogen and consumer behavior at the household level. In this study, we developed a QMRA model with a multiple-strain approach and a consumer phase module (CPM) based on uncertainty distributions fitted from field data. We modeled exposure to staphylococcal enterotoxin A in raw milk in Lombardy; a specific enterotoxin production module was thus included. The model is adaptable and could be used to assess the risk related to other pathogens in raw milk as well as other staphylococcal enterotoxins. The multiplestrain approach, implemented as a multinomial process, allowed the inclusion of variability and uncertainty with regard to pathogenicity at the bacterial level. Data from 301 questionnaires submitted to raw milk consumers were used to obtain uncertainty distributions for the CPM. The distributions were modeled to be easily updatable with further data or evidence. The sources of uncertainty due to the multiple-strain approach and the CPM were identified, and their impact on the output was assessed by comparing specific scenarios to the baseline. When the distributions reflecting the uncertainty in consumer behavior were fixed to the 95th percentile, the risk of exposure increased up to 160 times. This reflects the importance of taking into consideration the diversity of consumers' habits at the household level and the impact that the lack of knowledge about variables in the CPM can have on the final QMRA estimates. The multiple-strain approach lends itself to use in other food matrices besides raw milk and allows the model to better capture the complexity of the real world and to be capable of geographical

Inhibition of the superantigenic activities of Staphylococcal enterotoxin A by an aptamer antagonist.

Science.gov (United States)

Wang, Kaiyu; Wu, Dong; Chen, Zhuang; Zhang, Xianhui; Yang, Xiangyue; Yang, Chaoyong James; Lan, Xiaopeng

2016-09-01

Staphylococcal enterotoxin A (SEA) is an important component of Staphylococcus aureus pathogenesis. SEA induces T lymphocytes activation and proliferation, resulting in the release of a large number of inflammatory cytokines. Blocking the toxic cascade triggered by SEA may be an effective strategy for the treatment of SEA-induced diseases. Through a systematic evolution of ligands by exponential enrichment process, we obtained an aptamer (S3) that could bind SEA with both high affinity and specificity, with a Kd value 36.93 ± 7.29 nM (n = 3). This aptamer antagonist effectively inhibited SEA-mediated human peripheral blood mononuclear cells proliferation and inflammatory cytokines (IFN-γ, TNF-α, IL-2 and IL-6) secretion. Moreover, PEGylated S3 significantly reduced mortality in murine lethal toxic shock models established by lipopolysaccharide-potentiated SEA. Therefore, this novel aptamer antagonist has the potential to become a new strategy for treating S. aureus infections and SEA-induced diseases. Copyright © 2016 Elsevier Ltd. All rights reserved.

Plant-Derived Polyphenols Interact with Staphylococcal Enterotoxin A and Inhibit Toxin Activity.

Science.gov (United States)

Shimamura, Yuko; Aoki, Natsumi; Sugiyama, Yuka; Tanaka, Takashi; Murata, Masatsune; Masuda, Shuichi

2016-01-01

This study was performed to investigate the inhibitory effects of 16 different plant-derived polyphenols on the toxicity of staphylococcal enterotoxin A (SEA). Plant-derived polyphenols were incubated with the cultured Staphylococcus aureus C-29 to investigate the effects of these samples on SEA produced from C-29 using Western blot analysis. Twelve polyphenols (0.1-0.5 mg/mL) inhibited the interaction between the anti-SEA antibody and SEA. We examined whether the polyphenols could directly interact with SEA after incubation of these test samples with SEA. As a result, 8 polyphenols (0.25 mg/mL) significantly decreased SEA protein levels. In addition, the polyphenols that interacted with SEA inactivated the toxin activity of splenocyte proliferation induced by SEA. Polyphenols that exerted inhibitory effects on SEA toxic activity had a tendency to interact with SEA. In particular, polyphenol compounds with 1 or 2 hexahydroxydiphenoyl groups and/or a galloyl group, such as eugeniin, castalagin, punicalagin, pedunculagin, corilagin and geraniin, strongly interacted with SEA and inhibited toxin activity at a low concentration. These polyphenols may be used to prevent S. aureus infection and staphylococcal food poisoning.

Plant-Derived Polyphenols Interact with Staphylococcal Enterotoxin A and Inhibit Toxin Activity.

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Yuko Shimamura

Full Text Available This study was performed to investigate the inhibitory effects of 16 different plant-derived polyphenols on the toxicity of staphylococcal enterotoxin A (SEA. Plant-derived polyphenols were incubated with the cultured Staphylococcus aureus C-29 to investigate the effects of these samples on SEA produced from C-29 using Western blot analysis. Twelve polyphenols (0.1-0.5 mg/mL inhibited the interaction between the anti-SEA antibody and SEA. We examined whether the polyphenols could directly interact with SEA after incubation of these test samples with SEA. As a result, 8 polyphenols (0.25 mg/mL significantly decreased SEA protein levels. In addition, the polyphenols that interacted with SEA inactivated the toxin activity of splenocyte proliferation induced by SEA. Polyphenols that exerted inhibitory effects on SEA toxic activity had a tendency to interact with SEA. In particular, polyphenol compounds with 1 or 2 hexahydroxydiphenoyl groups and/or a galloyl group, such as eugeniin, castalagin, punicalagin, pedunculagin, corilagin and geraniin, strongly interacted with SEA and inhibited toxin activity at a low concentration. These polyphenols may be used to prevent S. aureus infection and staphylococcal food poisoning.

Concerted action of sphingomyelinase and non-hemolytic enterotoxin in pathogenic Bacillus cereus.

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Viktoria M Doll

Full Text Available Bacillus cereus causes food poisoning and serious non-gastrointestinal-tract infections. Non-hemolytic enterotoxin (Nhe, which is present in most B. cereus strains, is considered to be one of the main virulence factors. However, a B. cereus ΔnheBC mutant strain lacking Nhe is still cytotoxic to intestinal epithelial cells. In a screen for additional cytotoxic factors using an in vitro model for polarized colon epithelial cells we identified B. cereus sphingomyelinase (SMase as a strong inducer of epithelial cell death. Using single and double deletion mutants of sph, the gene encoding for SMase, and nheBC in B. cereus we demonstrated that SMase is an important factor for B. cereus cytotoxicity in vitro and pathogenicity in vivo. SMase substantially complemented Nhe induced cytotoxicity in vitro. In addition, SMase but not Nhe contributed significantly to the mortality rate of larvae in vivo in the insect model Galleria mellonella. Our study suggests that the role of B. cereus SMase as a secreted virulence factor for in vivo pathogenesis has been underestimated and that Nhe and SMase complement each other significantly to cause full B. cereus virulence hence disease formation.

Claudins Overexpression in Ovarian Cancer: Potential Targets for Clostridium Perfringens Enterotoxin (CPE Based Diagnosis and Therapy

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Diana P. English

2013-05-01

Full Text Available Claudins are a family of tight junction proteins regulating paracellular permeability and cell polarity with different patterns of expression in benign and malignant human tissues. There are approximately 27 members of the claudin family identified to date with varying cell and tissue-specific expression. Claudins-3, -4 and -7 represent the most highly differentially expressed claudins in ovarian cancer. While their exact role in ovarian tumors is still being elucidated, these proteins are thought to be critical for ovarian cancer cell invasion/dissemination and resistance to chemotherapy. Claudin-3 and claudin-4 are the natural receptors for the Clostridium perfringens enterotoxin (CPE, a potent cytolytic toxin. These surface proteins may therefore represent attractive targets for the detection and treatment of chemotherapy-resistant ovarian cancer and other aggressive solid tumors overexpressing claudin-3 and -4 using CPE-based theranostic agents.

The detection of K88, K99 fimbrial antigen and enterotoxin genes of Escherichia coli isolated from piglets and calves with diarrhoea in Indonesia

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Supar

1996-03-01

Full Text Available Enterotoxigenic Escherichia coli (ETEC strains cause diarrhoeal disease in piglets and calves in Indonesia. These strains possess two virulence factors namely attachment and enterotoxin antigens . These factors could be detected phenotypically and genetically. Haemolytic Escherichia coli (E coli isolates possessing K88 fimbrial antigen associated with 0-group 108 and 149. They were positive for K88 gene and demonstrated their ability to produce heat labile enterotoxin (LT and genetically were all positive for LT gene . Seventeen isolates ofE coli K88 which associated with 0-group 149 were positive forSTb gene, other O-serotypes were negative . Ten isolates of Ecoli K88 which associated with 0-group 108 possessed K88, K99, LT and STa genes, but negative for STb gene . However, phenotypically the K99 antigen and STa toxin were not expressed under laboratory conditions, the reason was not well understood . E. coli K99 strains isolated from calves wit h diarrhoea were all associated with 0-group 9 and produced STa toxin when tested by suckling mousse bioassay. The E. coli K99 calf isolates were all hybridized with K99 and STa gene only . It is likely that K99 gene is associated with STa gene . The DNA hybridization technique is more convenience to be used for confirmation diagnosis of colibacillosis, however, not all veterinary laboratories could perform these tests .

Immunogenicity of recombinant Lactobacillus casei-expressing F4 (K88) fimbrial adhesin FaeG in conjunction with a heat-labile enterotoxin A (LTAK63) and heat-labile enterotoxin B (LTB) of enterotoxigenic Escherichia coli as an oral adjuvant in mice.

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Yu, M; Qi, R; Chen, C; Yin, J; Ma, S; Shi, W; Wu, Y; Ge, J; Jiang, Y; Tang, L; Xu, Y; Li, Y

2017-02-01

The aims of this study were to develop an effective oral vaccine against enterotoxigenic Escherichia coli (ETEC) infection and to design new and more versatile mucosal adjuvants. Genetically engineered Lactobacillus casei strains expressing F4 (K88) fimbrial adhesin FaeG (rLpPG-2-FaeG) and either co-expressing heat-labile enterotoxin A (LTA) subunit with an amino acid mutation associated with reduced virulence (LTAK63) and a heat-labile enterotoxin B (LTB) subunit of E. coli (rLpPG-2-LTAK63-co-LTB) or fused-expressing LTAK63 and LTB (rLpPG-2-LTAK63-fu-LTB) were constructed. The immunogenicity of rLpPG-2-FaeG in conjunction with rLpPG-2-LTAK63-co-LTB or rLpPG-2-LTAK63-fu-LTB as an orally administered mucosal adjuvant in mice was evaluated. Results showed that the levels of FaeG-specific serum IgG and mucosal sIgA, as well as the proliferation of lymphocytes, were significantly higher in mice orally co-administered rLpPG-2-FaeG and rLpPG-2-LTAK63-fu-LTB compared with those administered rLpPG-2-FaeG alone, and were lower than those co-administered rLpPG-2-FaeG and rLpPG-2-LTAK63-co-LTB. Moreover, effective protection was observed after challenge with F4+ ETEC strain CVCC 230 in mice co-administered rLpPG-2-FaeG and rLpPG-2-LTAK63-co-LTB or rLpPG-2-FaeG and rLpPG-2-LTAK63-fu-LTB group compared with those that received rLpPG-2-FaeG alone. rLpPG-2-FaeG showed greater immunogenicity in combination with LTAK63 and LTB as molecular adjuvants. Recombinant Lactobacillus provides a promising platform for the development of vaccines against F4+ ETEC infection. © 2016 The Society for Applied Microbiology.

Bystander Host Cell Killing Effects of Clostridium perfringens Enterotoxin

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Archana Shrestha

2016-12-01

Full Text Available Clostridium perfringens enterotoxin (CPE binds to claudin receptors, e.g., claudin-4, and then forms a pore that triggers cell death. Pure cultures of host cells that do not express claudin receptors, e.g., fibroblasts, are unaffected by pathophysiologically relevant CPE concentrations in vitro. However, both CPE-insensitive and CPE-sensitive host cells are present in vivo. Therefore, this study tested whether CPE treatment might affect fibroblasts when cocultured with CPE-sensitive claudin-4 fibroblast transfectants or Caco-2 cells. Under these conditions, immunofluorescence microscopy detected increased death of fibroblasts. This cytotoxic effect involved release of a toxic factor from the dying CPE-sensitive cells, since it could be reproduced using culture supernatants from CPE-treated sensitive cells. Supernatants from CPE-treated sensitive cells, particularly Caco-2 cells, were found to contain high levels of membrane vesicles, often containing a CPE species. However, most cytotoxic activity remained in those supernatants even after membrane vesicle depletion, and CPE was not detected in fibroblasts treated with supernatants from CPE-treated sensitive cells. Instead, characterization studies suggest that a major cytotoxic factor present in supernatants from CPE-treated sensitive cells may be a 10- to 30-kDa host serine protease or require the action of that host serine protease. Induction of caspase-3-mediated apoptosis was found to be important for triggering release of the cytotoxic factor(s from CPE-treated sensitive host cells. Furthermore, the cytotoxic factor(s in these supernatants was shown to induce a caspase-3-mediated killing of fibroblasts. This bystander killing effect due to release of cytotoxic factors from CPE-treated sensitive cells could contribute to CPE-mediated disease.

Antimicrobial resistance and detection of the mecA gene besides enterotoxin-encoding genes among coagulase-negative Staphylococci isolated from clam meat of Anomalocardia brasiliana.

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Batista, Jacqueline Ellen Camelo; Ferreira, Ewerton Lucena; Nascimento, Danielle Cristina de Oliveira; Ventura, Roberta Ferreira; de Oliveira, Wagner Luis Mendes; Leal, Nilma Cintra; Lima-Filho, José Vitor

2013-12-01

The marine clam Anomalocardia brasiliana is a candidate as a sentinel animal to monitor the contamination levels of coliforms in shellfish-harvesting areas of Brazil's northeastern region. The aim of the present study was to search enterotoxin-encoding genes plus the mecA gene among coagulase-negative staphylococci (CNS) isolates from shellfish meats of A. brasiliana. The specimen clam (n=48; 40 clams per sample) was collected during low tide in the bay area of Mangue Seco from April through June 2009, and random samples of chilled and frozen shelled clam meat (n=33; 250 g per sample) were obtained from retail shops from January through March 2012. Seventy-nine CNS isolates were identified, including Staphylococcus xylosus, S. cohnii spp. urealyticus, S. sciuri, and S. lentus. A high percentage of isolates resistant to erythromycin (58.5%), penicillin (51.2%), and tetracycline (43.9%), and the fluoroquinolones levofloxacin (39%) and ciprofloxacin (34.1%) were recorded from those environmental samples. Isolates from retail shops were particularly resistant to oxacillin (55.3%) and penicillin (36.8%). All CNS resistant to oxacillin and/or cefoxitin were positive for the presence of the mecA gene, but phenotypically susceptible to vancomycin. Also, the enterotoxin-encoding genes seg and seh were detected through multiplex-polymerase chain reaction in 77.7% and 88.8% of the isolates from environmental samples, versus 90.5% and 100% of the isolates from retail shops, respectively. The data reveal the risk to public health due to consuming raw or undercooked shellfish containing enterotoxigenic plus methicillin-resistant CNS.

Structure of the superantigen staphylococcal enterotoxin B in complex with TCR and peptide-MHC demonstrates absence of TCR-peptide contacts.

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Rödström, Karin E J; Elbing, Karin; Lindkvist-Petersson, Karin

2014-08-15

Superantigens are immune-stimulatory toxins produced by Staphylococcus aureus, which are able to interact with host immune receptors to induce a massive release of cytokines, causing toxic shock syndrome and possibly death. In this article, we present the x-ray structure of staphylococcal enterotoxin B (SEB) in complex with its receptors, the TCR and MHC class II, forming a ternary complex. The structure, in combination with functional analyses, clearly shows how SEB adopts a wedge-like position when binding to the β-chain of TCR, allowing for an interaction between the α-chain of TCR and MHC. Furthermore, the binding mode also circumvents contact between TCR and the peptide presented by MHC, which enables SEB to initiate a peptide-independent activation of T cells. Copyright © 2014 by The American Association of Immunologists, Inc.

Passive therapy with humanized anti-staphylococcal enterotoxin B antibodies attenuates systemic inflammatory response and protects from lethal pneumonia caused by staphylococcal enterotoxin B-producing Staphylococcus aureus.

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Karau, Melissa J; Tilahun, Mulualem E; Krogman, Ashton; Osborne, Barbara A; Goldsby, Richard A; David, Chella S; Mandrekar, Jayawant N; Patel, Robin; Rajagopalan, Govindarajan

2017-10-03

Drugs such as linezolid that inhibit bacterial protein synthesis may be beneficial in treating infections caused by toxigenic Staphylococcus aureus. As protein synthesis inhibitors have no effect on preformed toxins, neutralization of pathogenic exotoxins with anti-toxin antibodies may be beneficial in conjunction with antibacterial therapy. Herein, we evaluated the efficacy of human-mouse chimeric high-affinity neutralizing anti-staphylococcal enterotoxin B (SEB) antibodies in the treatment of experimental pneumonia caused by SEB-producing S. aureus. Since HLA class II transgenic mice mount a stronger systemic immune response following challenge with SEB and are more susceptible to SEB-induced lethal toxic shock than conventional mice strains, HLA-DR3 transgenic mice were used. Lethal pneumonia caused by SEB-producing S. aureus in HLA-DR3 transgenic mice was characterized by robust T cell activation and elevated systemic levels of several pro-inflammatory cytokines and chemokines. Prophylactic administration of a single dose of linezolid 30 min prior to the onset of infection attenuated the systemic inflammatory response and protected from mortality whereas linezolid administered 60 min after the onset of infection failed to confer significant protection. Human-mouse chimeric high-affinity neutralizing anti-SEB antibodies alone, but not polyclonal human IgG, mitigated this response and protected from death when administered immediately after initiation of infection. Further, anti-SEB antibodies as well as intact polyclonal human IgG, but not its Fab or Fc fragments, protected from lethal pneumonia when followed with linezolid therapy 60 min later. In conclusion, neutralization of superantigens with high-affinity antibodies may have beneficial effects in pneumonia.

Staphylococcal enterotoxin-specific IgE antibodies in atopic dermatitis.

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Ide, Fumihito; Matsubara, Tomoyo; Kaneko, Miho; Ichiyama, Takashi; Mukouyama, Tokuko; Furukawa, Susumu

2004-06-01

The authors clarified the clinical significance of the measurement of serum concentrations of specific IgE antibodies to staphylococcal enterotoxin (SE) A- and SEB in atopic dermatitis (AD). The serum concentrations of SEA- and SEB-specific IgE antibodies in 140 pediatric patients with AD were measured with an immuno CAP -radioallergosorbent test system (RAST). To check the cross-reaction of specific IgE antibodies to SEA/SEB and other allergens, the CAP RAST fluorescent enzyme immunoassay inhibition test was performed. Forty-seven patients (33.6%) tested positive for either SEA- or SEB-specific IgE antibodies. School children showed higher positive rates of SEA/SEB-specific IgE antibodies than infants or young children. The patients with severe AD and those with exacerbation of symptoms in summer, had higher positive rates of SEA/SEB-specific IgE antibodies than patients with mild AD or those with exacerbation in winter. In addition, the positive rates of specific IgE antibodies to both dog-dander and cat-dander were higher in patients with positive SEA/SEB-specific IgE antibodies than in patients with negative ones. No cross-reactions occurred among specific IgE antibodies to SEA/SEB and dog/cat dander with one patient's serum, which had positive IgE-specific antibodies against cat/dog dander and SEA/SEB. The positive rate of SEA/SEB-specific IgE antibodies in the patients with dogs and/or cats as pets was 48.4%, which was higher than in those with no pets. Atopic dermatitis patients who exhibit high positive rates of SEA/SEB-specific IgE antibodies were found to be school children, severe cases, cases with high serum concentrations of total IgE, cases with exacerbation in summer, and cases with dogs and/or cats as pets. The measurement of serum concentrations of specific IgE antibodies to SEA and SEB, thus has some value for evaluating AD patients.

Colorimetric DNAzyme Biosensor for Convenience Detection of Enterotoxin B Harboring Staphylococcus aureus from Food Samples.

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Mondal, Bhairab; N, Bhavanashri; Ramlal, Shylaja; Kingston, Joseph

2018-02-14

In the present study, a colorimetric DNAzymes biosensor strategy was devised in combination with immunomagnetic separation for rapid and easy detection of enterotoxin B harboring Staphylococcus aureus from food and clinical samples. The method employs immunocapture of S. aureus and amplification of seb gene by DNAzyme complementary sequence integrated forward primer and with specific reverse primer. The DNAzyme sequence integrated dsDNA PCR products when treated with hemin and TMB (3,3',5,5'-tetramethylbenzidine) in the presence of H 2 O 2 produce colorimetric signal. A linear relationship of optical signal with the initial template of seb was obtained which could be monitored by visually or spectrophotrometrically for qualitative and quantitative detection. The limit of detection for the assay was approximately 10 2 CFU/mL of seb gene harboring target. This method is convenient compared to gel based and ELISA systems. Further, spiking studies and analysis on natural samples emphasized the robustness and applicability of developed method. Altogether, the established assay could be a reliable alternative, low-cost, viable detection tool for the routine investigation of seb from food and clinical sources.

Assessment of Enterotoxin Production and Cross-Contamination of Staphylococcus aureus between Food Processing Materials and Ready-To-Eat Cooked Fish Paste.

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Tango, Charles Nkufi; Hong, Sung-Sam; Wang, Jun; Oh, Deog-Hwan

2015-12-01

This study evaluated Staphylococcus aureus growth and subsequent staphylococcal enterotoxin A production in tryptone soy broth and on ready-to-eat cooked fish paste at 12 to 37 °C, as well as cross-contamination between stainless steel, polyethylene, and latex glove at room temperature. A model was developed using Barany and Roberts's growth model, which satisfactorily described the suitable growth of S. aureus with R(2)-adj from 0.94 to 0.99. Except at 12 °C, S. aureus cells in TSB presented a lag time lower (14.64 to 1.65 h), grew faster (0.08 to 0.31 log CFU/h) and produced SEA at lower cell density levels (5.65 to 6.44 log CFU/mL) compare to those inoculated on cooked fish paste with data of 16.920 to 1.985 h, 0.02 to 0.23 log CFU/h, and 6.19 to 7.11 log CFU/g, respectively. Staphylococcal enterotoxin type A (SEA) visual immunoassay test showed that primary SEA detection varied considerably among different storage temperature degrees and media. For example, it occurred only during exponential phase at 30 and 37 °C in TSB, but in cooked fish paste it took place at late exponential phase of S. aureus growth at 20 and 25 °C. The SEA detection test was negative on presence of S. aureus on cooked fish paste stored at 12 and 15 °C, although cell density reached level of 6.12 log CFU/g at 15 °C. Cross-contamination expressed as transfer rate of S. aureus from polyethylene surface to cooked fish paste surface was slower than that observed with steel surface to cooked fish paste under same conditions. These results provide helpful information for controlling S. aureus growth, SEA production and cross-contamination during processing of cooked fish paste. © 2015 Institute of Food Technologists®

The fecal presence of enterotoxin and F4 genes as an indicator of efficacy of treatment with colistin sulfate in pigs.

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Rhouma, Mohamed; Fairbrother, John Morris; Thériault, William; Beaudry, Francis; Bergeron, Nadia; Laurent-Lewandowski, Sylvette; Letellier, Ann

2017-01-05

Enterotoxigenic Escherichia coli (ETEC) strains producing multiple enterotoxins are important causes of post-weaning diarrhea (PWD) in pigs. The aim of the present study was to investigate the fecal presence of ETEC enterotoxin as well as F4 and F18 genes as an indicator of colistin sulfate (CS) efficacy for treatment of PWD in pigs. Forty-eight piglets were weaned at the age of 21 days, and were divided into four groups: challenged treated, challenged untreated, unchallenged treated, and unchallenged untreated. Challenge was performed using 10 9  CFU of an ETEC: F4 strain, and treatment was conducted using oral CS at the dose of 50,000 IU/kg. The fecal presence of genes encoding for STa, STb, LT, F4 and F18 was detected using PCR. The PCR amplification of ETEC virulence genes showed that nearly 100% of pigs excreted genes encoding for STa and STb toxins in the feces before the challenge. These genes, in the absence of the gene encoding F4, were considered as a marker for F4-negative ETEC. One day after ETEC: F4 oral challenge pigs in the two challenged groups excreted the genes encoding LT and F4 in the feces. These genes were considered as a marker for F4-positive ETEC. However, the gene encoding F18 was not detected in any fecal samples of the 4 groups throughout the experiment. After only 3 days of successive oral treatment with CS, a significant reduction in both the F4-positive and negative ETEC populations was observed in the challenged treated group compared to the challenged untreated group (p F4-positive and F4-negative ETEC in pigs. However, CS clinical efficiency was correlated with non-detection of F4-positive ETEC in the feces. Furthermore the fecal presence of F4-negative ETEC was not associated with clinical symptoms of post-weaning diarrhea in pigs.

Colorimetric determination of staphylococcal enterotoxin B via DNAzyme-guided growth of gold nanoparticles

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Zhou, Dandan; Chen, Hui; Xie, Guoming; Cao, Xianqing; Chen, Xueping; Zhang, Xing

2016-01-01

The authors describe a colorimetric method for the determination of the staphylococcal enterotoxin B (SEB) that also allows for visual readout. The assay is based on the growth of gold nanoparticles (AuNPs) mediated by a hemin/G-quadruplex DNAzyme which generates a color change from red to blue in the presence of SEB. The method is enzyme-free and does not require a label. The kinetics of the formation of the AuNPs is controlled by the hemin/G-quadruplex DNAzyme and this is key to the signal generation mechanism. In the presence of SEB, the reactions between aptamer and target modulated the amount of single probe G strands that form DNAzyme capable of consuming hydrogen peroxide. The growth process of AuNPs is influenced by the resulting concentration of H 2 O 2 and leads to the color change. Under optimal conditions, a linear relationship exists between absorbance and SEB concentration in the range from 0.1 to 500 pg·mL -1 which covers the clinically relevant range. In case of visual detection, the lower limit of detection is 1 pg·mL −1 . The assay described here is sensitive, comparably inexpensive and can detect SEB rapidly without the need for sophisticated equipment. In our perception, the method has a wide scope in that it may be adapted to various nucleic acids, proteins and other biomolecules if respective aptamers are available. (author)

[Poisoning by enterotoxin from Staphylococcus aureus associated with mocha pastry. Microbiology and epidemiology].

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Escartín, E F; Saldaña-Lozano, J; Montiel-Falcón, A

1998-01-01

A brief description of a foodborne outbreak due to S. aureus enterotoxin associated with the consumption of mocha cake in the city of Guadalajara is presented. The cake was prepared in a bakery and affected nearly 100 persons. S. aureus was isolated from the nose and skin of one of the pastry cooks. A S. aureus strain isolated from the cake involved in the outbreak was not only unable to grow in the mocha cream, but it actually decreased in numbers by 2 log after 72 h of storage at 30 degrees C. The pH of mocha cream ranged from 6.2 to 6.6, and water activity from 0.833 to 0.859, with a media of 0.841. In preparing mocha cake at the shop, one half of the dough used to be sprayed with a sucrose solution in water (20% w/v); mocha cream was spread on the other half of the dough before overlapping the two halves. When mocha cake was prepared in this manner, and stored at 30 degrees C, S. aureus increased in number by more than 4 log after 48 h. S. aureus did not grow in the cake stored at 4-7 degrees C. Contributory factors in this outbreak were an increase of water activity in the interphase of the mocha and the cake dough, storage of the cake in an unrefrigerated area, and an unusually high ambient temperature (28-32 degrees C) at that time.

An alternative explanation for the occurrence of short circuit current increases in the small intestine following challenge by bacterial enterotoxins.

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Lucas, M L

2013-10-01

Secretory diarrhoeal disease due to enterotoxins is thought to arise from the enhancement to pathologically high rates of normally occurring chloride ion and therefore fluid secretion from enterocytes. In support of this concept, many enterotoxins increase intestinal short-circuit current, regarded now as faithfully reflecting the increased chloride ion secretion. Contradicting this assumption, STa reduces absorption but does not cause secretion in vivo although short-circuit current is increased in vitro. There is therefore a mismatch between an assumed enterocyte mediated secretory event that should but does not cause net fluid secretion and an undoubtedly increased short-circuit current. It is proposed here that short-circuit current increases are not themselves secretory events but result from interrupted fluid absorption. A noteworthy feature of compounds that inhibit the increase in short-circuit current is that the majority are vasoactive, neuroactive or both. In general, vasodilator substances increase current. An alternative hypothesis for the origin of short-circuit current increases is that these result from reflex induction of electrogenic fluid absorption. This reflex enhances a compensatory response that is also present at a cellular level. An intestinal reflex is therefore proposed by which decreases in interstitial and intravascular volume or pressure within the intestine initiate an electrogenic fluid absorption mechanism that compensates for the loss of electrically neutral fluid absorption. This hypothesis would explain the apparently complex pharmacology of short-circuit current increases since many depressor substances have receptors in common with enterocytes and enteric nerves. The proposed alternative view of the origin of short-circuit current increases assumes that these do not represent chloride secretion from the enterocytes. This view may therefore aid the successful development of anti-diarrhoeal drugs to overcome a major cause of

A highly sensitive fluorescence resonance energy transfer aptasensor for staphylococcal enterotoxin B detection based on exonuclease-catalyzed target recycling strategy

International Nuclear Information System (INIS)

Wu, Shijia; Duan, Nuo; Ma, Xiaoyuan; Xia, Yu; Wang, Hongxin; Wang, Zhouping

2013-01-01

Graphical abstract: -- Highlights: •An ultrasensitive FRET aptasensor was developed for staphylococcal enterotoxin B determination. •SEB was recognized by SEB aptamer with high affinity and specificity. •The Mn 2+ doped NaYF 4 :Yb/Er UCNPs used as donor to quencher dye (BHQ 3 ) in new FRET. •The fluorescence intensity was prominently amplified using an exonuclease-catalyzed target recycling strategy. -- Abstract: An ultrasensitive fluorescence resonance energy transfer (FRET) bioassay was developed to detect staphylococcal enterotoxin B (SEB), a low molecular exotoxin, using an aptamer-affinity method coupled with upconversion nanoparticles (UCNPs)-sensing, and the fluorescence intensity was prominently enhanced using an exonuclease-catalyzed target recycling strategy. To construct this aptasensor, both fluorescence donor probes (complementary DNA 1 –UCNPs) and fluorescence quencher probes (complementary DNA 2 –Black Hole Quencher 3 (BHQ 3 )) were hybridized to an SEB aptamer, and double-strand oligonucleotides were fabricated, which quenched the fluorescence of the UCNPs via FRET. The formation of an aptamer–SEB complex in the presence of the SEB analyte resulted in not only the dissociation of aptamer from the double-strand DNA but also both the disruption of the FRET system and the restoration of the UCNPs fluorescence. In addition, the SEB was liberated from the aptamer–SEB complex using exonuclease I, an exonuclease specific to single-stranded DNA, for analyte recycling by selectively digesting a particular DNA (SEB aptamer). Based on this exonuclease-catalyzed target recycling strategy, an amplified fluorescence intensity could be produced using different SEB concentrations. Using optimized experimental conditions produced an ultrasensitive aptasensor for the detection of SEB, with a wide linear range of 0.001–1 ng mL −1 and a lower detection limit (LOD) of 0.3 pg mL −1 SEB (at 3σ). The fabricated aptasensor was used to measure SEB in a

Structure of Staphylococcal Enterotoxin E in Complex with TCR Defines the Role of TCR Loop Positioning in Superantigen Recognition.

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Karin E J Rödström

Full Text Available T cells are crucial players in cell-mediated immunity. The specificity of their receptor, the T cell receptor (TCR, is central for the immune system to distinguish foreign from host antigens. Superantigens are bacterial toxins capable of inducing a toxic immune response by cross-linking the TCR and the major histocompatibility complex (MHC class II and circumventing the antigen specificity. Here, we present the structure of staphylococcal enterotoxin E (SEE in complex with a human T cell receptor, as well as the unligated T cell receptor structure. There are clear structural changes in the TCR loops upon superantigen binding. In particular, the HV4 loop moves to circumvent steric clashes upon complex formation. In addition, a predicted ternary model of SEE in complex with both TCR and MHC class II displays intermolecular contacts between the TCR α-chain and the MHC, suggesting that the TCR α-chain is of importance for complex formation.

Immunochemical and PCR analysis of Staphylococcus aureus Enterotoxin B (SEB in milk and fruit juices collected in Lahore, Pakistan

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Dar Nadia

2013-01-01

Full Text Available Enterotoxins secreted by S. aureus are known as a food-poisoning agent that is associated with various gastro-intestinal pathological conditions. In this study, a one-step immunodetection method was devised for routine checking of SEB in milk and fruit juices available locally. Antibodies against recombinant SEB were raised, purified, and cross reactivity was checked against clinically important bacteria (Shigella flexneri, Streptococci, Salmonella typhi, Klebsiella and Bacillus subtilis. Purified anti-SEB antibodies were conjugated with gold nanoparticles (Ab-GNPs for direct detection of SEB in samples. SEB (33%, 4.76% and 15% was found in non-sterilized milk (118, sterilized milk (42 and juices (60, respectively. Coagulase, MSA tests and PCR amplification of 725 bp of the SEB gene confirmed the presence of S. aureus in the collected samples positive for SEB. Immunoassay is easy, reliable and less time consuming and will be helpful to detect the SEB in food samples at local level.

Inhibition of biological activity of staphylococcal enterotoxin A (SEA) by apple juice and apple polyphenols.

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Rasooly, Reuven; Do, Paula M; Friedman, Mendel

2010-05-12

The foodborne pathogen Staphylococcus aureus produces the virulent staphylococcal enterotoxin A (SEA), a single-chain protein that consists of 233 amino acid residues with a molecular weight of 27 078 Da. SEA is a superantigen that is reported to contribute to animal (mastitis) and human (emesis, diarrhea, atopic dermatitis, arthritis, and toxic shock) syndromes. Changes of the native structural integrity may inactivate the toxin by preventing molecular interaction with cell membrane receptor sites of their host cells. In the present study, we evaluated the ability of one commercial and two freshly prepared apple juices and a commercial apple polyphenol preparation (Apple Poly) to inhibit the biological activity of SEA. Dilutions of freshly prepared apple juices and Apple Poly inhibited the biological activity of SEA without any significant cytotoxic effect on the spleen cells. Additional studies with antibody-coated immunomagnetic beads bearing specific antibodies against the toxin revealed that SEA added to apple juice appears to be largely irreversibly bound to the juice constituents. The results suggest that food-compatible and safe anti-toxin phenolic compounds can be used to inactivate SEA in vitro and possibly also in vivo, even after induction of T-cell proliferation by long-term exposure to SEA. The significance of the results for microbial food safety and human health is discussed.

Evaluation of Potential Effects of NaCl and Sorbic Acid on Staphylococcal Enterotoxin A Formation

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Nikoleta Zeaki

2015-09-01

Full Text Available The prophage-encoded staphylococcal enterotoxin A (SEA is recognized as the main cause of staphylococcal food poisoning (SFP, a common foodborne intoxication disease, caused by Staphylococcus aureus. Studies on the production of SEA suggest that activation of the SOS response and subsequent prophage induction affect the regulation of the sea gene and the SEA produced, increasing the risk for SFP. The present study aims to evaluate the effect of NaCl and sorbic acid, in concentrations relevant to food production, on SOS response activation, prophage induction and SEA production. The impact of stress was initially evaluated on steady state cells for a homogenous cell response. NaCl 2% was found to activate the SOS response, i.e., recA expression, and trigger prophage induction, in a similar way as the phage-inducer mitomycin C. In contrast, sorbic acid decreased the pH of the culture to a level where prophage induction was probably suppressed, even when combined with NaCl stress. The impact of previous physiological state of the bacteria was also addressed on cells pre-exposed to NaCl, and was found to potentially affect cell response upon exposure to further stress. The results obtained highlight the possible SFP-related risks arising from the use of preservatives during food processing.

Attenuated Escherichia coli strains expressing the colonization factor antigen I (CFA/I) and a detoxified heat-labile enterotoxin (LThK63) enhance clearance of ETEC from the lungs of mice and protect mice from intestinal ETEC colonization and LT-induced fluid accumulation.

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Byrd, Wyatt; Boedeker, Edgar C

2013-03-15

Although enterotoxigenic Escherichia coli (ETEC) infections are important causes of infantile and traveler's diarrhea there is no licensed vaccine available for those at-risk. Our goal is to develop a safe, live attenuated ETEC vaccine. We used an attenuated E. coli strain (O157:H7, Δ-intimin, Stx1-neg, Stx2-neg) as a vector (ZCR533) to prepare two vaccine strains, one strain expressing colonization factor antigen I (ZCR533-CFA/I) and one strain expressing CFA/I and a detoxified heat-labile enterotoxin (ZCR533-CFA/I+LThK63) to deliver ETEC antigens to mucosal sites in BALB/c mice. Following intranasal and intragastric immunization with the vaccine strains, serum IgG and IgA antibodies were measured to the CFA/I antigen, however, only serum IgG antibodies were detected to the heat-labile enterotoxin. Intranasal administration of the vaccine strains induced respiratory and intestinal antibody responses to the CFA/I and LT antigens, while intragastric administration induced only intestinal antibody responses with no respiratory antibodies detected to the CFA/I and LT antigens. Mice immunized intranasally with the vaccine strains showed enhanced clearance of wild-type (wt) ETEC bacteria from the lungs. Mice immunized intranasally and intragastrically with the vaccine strains were protected from intestinal colonization following oral challenge with ETEC wt bacteria. Mice immunized intragastrically with the ZCR533-CFA/I+LThK63 vaccine strain had less fluid accumulate in their intestine following challenge with ETEC wt bacteria or with purified LT as compared to the sham mice indicating that the immunized mice were protected from LT-induced intestinal fluid accumulation. Thus, mice intragastrically immunized with the ZCR533-CFA/I+LThK63 vaccine strain were able to effectively neutralize the activity of the LT enterotoxin. However, no difference in intestinal fluid accumulation was detected in the mice immunized intranasally with the vaccine strain as compared to the sham

Molecular and epidemiological characterization of staphylococcal foodborne outbreak of Staphylococcus aureus harboring seg, sei, sem, sen, seo, and selu genes without production of classical enterotoxins.

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Umeda, Kaoru; Nakamura, Hiromi; Yamamoto, Kaori; Nishina, Nobuko; Yasufuku, Kiyoshi; Hirai, Yuki; Hirayama, Teruo; Goto, Kaoru; Hase, Atsushi; Ogasawara, Jun

2017-09-01

Staphylococcal food poisoning is the result of consumption of food contaminated with staphylococcal enterotoxins (SEs) produced by Staphylococcus aureus. To date, 23 SEs and SE-like enterotoxins (SEls) have been described in the literature. They are divided into classical SEs (SEA-SEE) and new SE/SEls (SEG-SElX). Some have proved to be foodborne-inducible, but others remain unidentified. In May 2016, at an elderly group home in Osaka city, Japan, an outbreak from foodborne pathogens occurred among lunch party participants. Within 2h 30min to 4h 40min, 15 of 53 participants presented gastrointestinal symptoms of vomiting, diarrhea, and nausea. A subsequent laboratory investigation detected S. aureus from most stool samples from patients, several left-over food items, a kitchen swab, and hand swabs from two food handlers. Classical SEs was not detected from S. aureus isolates or left-over food items. From examination for the presence of SE/SEl genes of 20 kinds by PCR, seg, sei, sem, sen, seo, and selu genes were detected in almost all isolates. These isolates exhibited identical or closely related types by coagulase type (type VII), Sma I digested pulsed-field gel electrophoresis analysis and multi-locus sequence typing (MLST-CC45 lineage). These results suggest that the foodborne outbreak was caused by S. aureus harboring seg, sei, sem, sen, seo, and selu genes without production of classical SEs. Additionally, some S. aureus isolates from human nasal swabs and healthy human feces harboring seg, sei, sem, sen, seo, and selu genes without production of classical SEs were classified into CC45 lineage using MLST. These findings suggest new SE/SEls as a potential cause of foodborne outbreaks. Copyright © 2017. Published by Elsevier B.V.

Positive Regulation of Staphylococcal Enterotoxin H by Rot (Repressor of Toxin) Protein and Its Importance in Clonal Complex 81 Subtype 1 Lineage-Related Food Poisoning.

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Sato'o, Yusuke; Hisatsune, Junzo; Nagasako, Yuria; Ono, Hisaya K; Omoe, Katsuhiko; Sugai, Motoyuki

2015-11-01

We previously demonstrated the clonal complex 81 (CC81) subtype 1 lineage is the major staphylococcal food poisoning (SFP)-associated lineage in Japan (Y. Sato'o et al., J Clin Microbiol 52:2637-2640, 2014, http://dx.doi.org/10.1128/JCM.00661-14). Strains of this lineage produce staphylococcal enterotoxin H (SEH) in addition to SEA. However, an evaluation of the risk for the recently reported SEH has not been sufficiently conducted. We first searched for staphylococcal enterotoxin (SE) genes and SE proteins in milk samples that caused a large SFP outbreak in Japan. Only SEA and SEH were detected, while there were several SE genes detected in the samples. We next designed an experimental model using a meat product to assess the productivity of SEs and found that only SEA and SEH were detectably produced in situ. Therefore, we investigated the regulation of SEH production using a CC81 subtype 1 isolate. Through mutant analysis of global regulators, we found the repressor of toxin (Rot) functioned oppositely as a stimulator of SEH production. SEA production was not affected by Rot. seh mRNA expression correlated with rot both in media and on the meat product, and the Rot protein was shown to directly bind to the seh promoter. The seh promoter sequence was predicted to form a loop structure and to hide the RNA polymerase binding sequences. We propose Rot binds to the promoter sequence of seh and unfolds the secondary structure that may lead the RNA polymerase to bind the promoter, and then seh mRNA transcription begins. This alternative Rot regulation for SEH may contribute to sufficient toxin production by the CC81 subtype 1 lineage in foods to induce SFP. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Characteristics of enterotoxin distribution, hemolysis, lecithinase, and starch hydrolysis of Bacillus cereus isolated from infant formulas and ready-to-eat foods.

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Hwang, Ji-Yeon; Park, Jong-Hyun

2015-03-01

Bacillus cereus is a ubiquitous environmental microbe implicated as a main cause of food poisoning with various symptoms, depending on the strain type and the isolation source. In this study, the potential virulence factors and biochemical properties of B. cereus isolated from infant formulas and ready-to-eat (RTE) foods were analyzed and compared. A total of 347 B. cereus strains were isolated and identified from 687 infant food formulas and RTE food samples. All the isolates had one or more enterotoxin genes, and one-half of the strains had all 3 enterotoxin genes (hbl, nhe, and cytK) that are involved in food poisoning in humans. Here, all the 3 genes were detected in 50% of the B. cereus isolates from RTE foods and only 14% of the isolates were identified from infant formulas. The latter harbored low cytK and bceT, and very low hbl genes. Most B. cereus isolates possessed the hemolysis gene, but not the ces gene. The infant formula isolates showed stronger hemolysis activity than the other isolates. In addition, 26% of the total isolates showed low lecithinase activities and 10% showed high lecithinase activities. A greater number of isolates from the infant formula showed high lecithinase activity than those from the RTE foods. Approximately 83% of the isolates were positive and 17% were negative for starch hydrolysis. Over 90% of the RTE food isolates and only 35% of the infant formula isolates were positive for starch hydrolysis. However, all the strains possessed nhe, but their harboring patterns of hbl and cytK were significantly different. Most starch-hydrolyzing strains possessed hbl, but only 23% nonstarch-hydrolyzing isolates possessed this gene. Moreover, very low nonstarch hydrolyzing strains harbored cytK. Most nonstarch-hydrolyzing isolates showed high lecithinase and strong hemolysis activities, and very low hbl and cytK harboring. In summary, most infant formula isolates showed stronger hemolysis and higher lecithinase activities with lower

Prophage-Encoded Staphylococcal Enterotoxin A: Regulation of Production in Staphylococcus aureus Strains Representing Different Sea Regions

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Nikoleta Zeaki

2015-12-01

Full Text Available The present study investigates the nature of the link between the staphylococcal enterotoxin A (SEA gene and the lifecycle of Siphoviridae bacteriophages, including the origin of strain variation regarding SEA production after prophage induction. Five strains representing three different genetic lines of the sea region were studied under optimal and prophage-induced growth conditions and the Siphoviridae lifecycle was followed through the phage replicative form copies and transcripts of the lysogenic repressor, cro. The role of SOS response on prophage induction was addressed through recA transcription in a recA-disruption mutant. Prophage induction was found to increase the abundance of the phage replicative form, the sea gene copies and transcripts and enhance SEA production. Sequence analysis of the sea regions revealed that observed strain variances were related to strain capacity for prophage induction, rather than sequence differences in the sea region. The impact of SOS response activation on the phage lifecycle was demonstrated by the absence of phage replicative form copies in the recA-disruption mutant after prophage induction. From this study it emerges that all aspects of SEA-producing strain, the Siphoviridae phage and the food environment must be considered when evaluating SEA-related hazards.

Suppression of colorectal tumorigenesis by recombinant Bacteroides fragilis enterotoxin-2 in vivo.

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Lv, You; Ye, Tao; Wang, Hui-Peng; Zhao, Jia-Ying; Chen, Wen-Jie; Wang, Xin; Shen, Chen-Xia; Wu, Yi-Bin; Cai, Yuan-Kun

2017-01-28

To evaluate the impact of recombinant Bacteroides fragilis enterotoxin-2 (BFT-2, or Fragilysin) on colorectal tumorigenesis in mice induced by azoxymethane/dextran sulfate sodium (AOM/DSS). Recombinant proBFT-2 was expressed in Escherichia coli strain Rosetta (DE3) and BFT-2 was obtained and tested for its biological activity via colorectal adenocarcinoma cell strains SW-480. Seventy C57BL/6J mice were randomly divided into a blank (BC; n = 10), model (AD; n = 20), model + low-dose toxin (ADLT; n = 20, 10 μg), and a model + high-dose toxin (ADHT; n = 20, 20 μg) group. Mice weight, tumor formation and pathology were analyzed. Immunohistochemistry determined Ki-67 and Caspase-3 expression in normal and tumor tissues of colorectal mucosa. Recombinant BFT-2 was successfully obtained, along with its biological activity. The most obvious weight loss occurred in the AD group compared with the ADLT group (21.82 ± 0.68 vs 23.23 ± 0.91, P ADHT group (21.82 ± 0.68 vs 23.57 ± 1.06, P ADHT groups (19.75 ± 3.30 vs 6.50 ± 1.73, P ADHT group. The incidence of colorectal adenocarcinoma in both the ADHT group and the ADHT group was reduced compared to that in the AD group ( P ADHT group was 50% and 40%, respectively, both of which were lower than that found in the AD group (94.44%, P ADHT group was 45% and 55%, both of which were higher than that found in the BC group (16.67%, P < 0.05, P < 0.05). Oral administration with lower-dose biologically active recombinant BFT-2 inhibited colorectal tumorigenesis in mice.

Rhinosinusitis derived Staphylococcal enterotoxin B possibly associates with pathogenesis of ulcerative colitis

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An Zi-Yuan

2005-09-01

Full Text Available Abstract Background During clinical practice, we noticed that some patients with both ulcerative colitis (UC and chronic rhinosinusitis (CRS showed amelioration of UC after treatment of CRS. This study was designed to identify a possible association between CRS and UC. Methods Thirty-two patients with both CRS and UC received treatment with functional endoscopic sinus surgery (FESS for CRS. Clinical symptom scores for CRS and UC, as well as serum levels of anti-Staphylococcal enterotoxin B (SEB were evaluated at week 0 and week 12. Sinus wash fluid SEB content was measured with enzyme-linked immunosorbent assay (ELISA. The surgically removed tissues were cultured to identify growth of Staphylococcus. aureus (S. aureus. Immunohistochemistry was employed to identify anti-SEB positive cells in the colonic mucosa. Colonic biopsies were obtained and incubated with SEB. Mast cell activation in the colonic mucosa in response to incubation with SEB was observed with electron microscopy and immunoassay. Results The clinical symptom scores of CRS and UC severe scores (UCSS were significantly reduced in the UC-CRS patients after FESS. The number of cultured S. aureus colonies from the surgically removed sinus mucosa significantly correlated with the decrease in UCSS. High levels of SEB were detected in the sinus wash fluids of the patients with UC-CRS. Histamine and tryptase release was significantly higher in the culture supernate in the patients with UC-CRS than the patients with UC-only and normal controls. Anti-SEB positive cells were located in the colonic mucosa. Conclusion The pathogenesis of UC in some patients may be associated with their pre-existing CRS by a mechanism of swallowing sinusitis-derived SEB. We speculate that SEB initiates inappropriate immune reactions and inflammation in the colonic mucosa that further progresses to UC.

Attenuation of massive cytokine response to the staphylococcal enterotoxin B superantigen by the innate immunomodulatory protein lactoferrin

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Hayworth, J L; Kasper, K J; Leon-Ponte, M; Herfst, C A; Yue, D; Brintnell, W C; Mazzuca, D M; Heinrichs, D E; Cairns, E; Madrenas, J; Hoskin, D W; McCormick, J K; Haeryfar, S M M

2009-01-01

Staphylococcal enterotoxin B (SEB) is a pyrogenic exotoxin and a potent superantigen which causes massive T cell activation and cytokine secretion, leading to profound immunosuppression and morbidity. The inhibition of SEB-induced responses is thus considered a goal in the management of certain types of staphylococcal infections. Lactoferrin (LF) is a multi-functional glycoprotein with both bacteriostatic and bactericidal activities. In addition, LF is known to have potent immunomodulatory properties. Given the anti-microbial and anti-inflammatory properties of this protein, we hypothesized that LF can modulate T cell responses to SEB. Here, we report that bovine LF (bLF) was indeed able to attenuate SEB-induced proliferation, interleukin-2 production and CD25 expression by human leucocyte antigen (HLA)-DR4 transgenic mouse T cells. This inhibition was not due to bLF's iron-binding capacity, and could be mimicked by the bLF-derived peptide lactoferricin. Cytokine secretion by an engineered SEB-responsive human Jurkat T cell line and by peripheral blood mononuclear cells from healthy donors was also inhibited by bLF. These findings reveal a previously unrecognized property of LF in modulation of SEB-triggered immune activation and suggest a therapeutic potential for this naturally occurring protein during toxic shock syndrome. PMID:19659771

Evaluation of Handheld Assays for the Detection of Ricin and Staphylococcal Enterotoxin B in Disinfected Waters

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Mary Margaret Wade

2011-01-01

Full Text Available Development of a rapid field test is needed capable of determining if field supplies of water are safe to drink by the warfighter during a military operation. The present study sought to assess the effectiveness of handheld assays (HHAs in detecting ricin and Staphylococcal Enterotoxin B (SEB in water. Performance of HHAs was evaluated in formulated tap water with and without chlorine, reverse osmosis water (RO with chlorine, and RO with bromine. Each matrix was prepared, spiked with ricin or SEB at multiple concentrations, and then loaded onto HHAs. HHAs were allowed to develop and then read visually. Limits of detection (LOD were determined for all HHAs in each water type. Both ricin and SEB were detected by HHAs in formulated tap water at or below the suggested health effect levels of 455 ng/mL and 4.55 ng/mL, respectively. However, in brominated or chlorinated waters, LODs for SEB increased to approximately 2,500 ng/mL. LODs for ricin increased in chlorinated water, but still remained below the suggested health effect level. In brominated water, the LOD for ricin increased to approximately 2,500 ng/mL. In conclusion, the HHAs tested were less effective at detecting ricin and SEB in disinfected water, as currently configured.

Modeling the effects of a Staphylococcal Enterotoxin B (SEB on the apoptosis pathway

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Hammamieh Rasha

2006-05-01

Full Text Available Abstract Background The lack of detailed understanding of the mechanism of action of many biowarfare agents poses an immediate challenge to biodefense efforts. Many potential bioweapons have been shown to affect the cellular pathways controlling apoptosis 1234. For example, pathogen-produced exotoxins such as Staphylococcal Enterotoxin B (SEB and Anthrax Lethal Factor (LF have been shown to disrupt the Fas-mediated apoptotic pathway 24. To evaluate how these agents affect these pathways it is first necessary to understand the dynamics of a normally functioning apoptosis network. This can then serve as a baseline against which a pathogen perturbed system can be compared. Such comparisons can expose both the proteins most susceptible to alteration by the agent as well as the most critical reaction rates to better instill control on a biological network. Results We explore this through the modeling and simulation of the Fas-mediated apoptotic pathway under normal and SEB influenced conditions. We stimulated human Jurkat cells with an anti-Fas antibody in the presence and absence of SEB and determined the relative levels of seven proteins involved in the core pathway at five time points following exposure. These levels were used to impute relative rate constants and build a quantitative model consisting of a series of ordinary differential equations (ODEs that simulate the network under both normal and pathogen-influenced conditions. Experimental results show that cells exposed to SEB exhibit an increase in the rate of executioner caspase expression (and subsequently apoptosis of 1 hour 43 minutes (± 14 minutes, as compared to cells undergoing normal cell death. Conclusion Our model accurately reflects these results and reveals intervention points that can be altered to restore SEB-influenced system dynamics back to levels within the range of normal conditions.

Ultrasensitive electrochemical immunoassay of staphylococcal enterotoxin B in food using enzyme-nanosilica-doped carbon nanotubes for signal amplification.

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Tang, Dianping; Tang, Juan; Su, Biling; Chen, Guonan

2010-10-27

A new sandwich-type electrochemical immunoassay for ultrasensitive detection of staphylococcal enterotoxin B (SEB) in food was developed using horseradish peroxidase-nanosilica-doped multiwalled carbon nanotubes (HRPSiCNTs) for signal amplification. Rabbit polyclonal anti-SEB antibodies immobilized on the screen-printed carbon electrode (SPCE) and covalently bound to the HRPSiCNTs were used as capture antibodies and detection antibodies, respectively. In the presence of SEB analyte, the sandwich-type immunocomplex could be formed between the immobilized anti-SEB on the SPCE and anti-SEB-labeled HRPSiCNTs, and the carried HRP could catalyze the electrochemical reduction of H2O2 with the help of thionine. The high content of HRP in the HRPSiCNTs could greatly amplify the electrochemical signal. Under optimal conditions, the reduction current increased with the increase of SEB in the sample, and exhibited a dynamic range of 0.05-15 ng/mL with a low detection limit (LOD) of 10 pg/mL SEB (at 3σ). Intra- and interassay coefficients of variation were below 10%. In addition, the assay was evaluated with SEB spiked samples including watermelon juice, soymilk, apple juice, and pork food, receiving excellent correlation with results from commercially available enzyme-linked immunosorbent assay (ELISA).

Nitrate salts suppress sporulation and production of enterotoxin in Clostridium perfringens strain NCTC8239.

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Yasugi, Mayo; Otsuka, Keisuke; Miyake, Masami

2016-10-01

Clostridium perfringens type A is a common source of food-borne illness in humans. Ingested vegetative cells sporulate in the small intestinal tract and in the process produce C. perfringens enterotoxin (CPE). Although sporulation plays a critical role in the pathogenesis of food-borne illness, the molecules triggering/inhibiting sporulation are still largely unknown. It has previously been reported by our group that sporulation is induced in C. perfringens strain NCTC8239 co-cultured with Caco-2 cells in Dulbecco's Modified Eagle Medium (DMEM). In contrast, an equivalent amount of spores was not observed when bacteria were co-cultured in Roswell Park Memorial Institute-1640 medium (RPMI). In the present study it was found that, when these two media are mixed, RPMI inhibits sporulation and CPE production induced in DMEM. When a component of RPMI was added to DMEM, it was found that calcium nitrate (Ca[NO 3 ] 2 ) significantly inhibits sporulation and CPE production. The number of spores increased when Ca(NO 3 ) 2 -deficient RPMI was used. The other nitrate salts significantly suppressed sporulation, whereas the calcium salts used did not. qPCR revealed that nitrate salts increased expression of bacterial nitrate/nitrite reductase. Furthermore, it was found that nitrite and nitric oxide suppress sporulation. In the sporulation stages, Ca(NO 3 ) 2 down-regulated the genes controlled by Spo0A, a master regulator of sporulation, but not spo0A itself. Collectively, these results indicate that nitrate salts suppress sporulation and CPE production by down-regulating Spo0A-regulated genes in C. perfringens strain NCTC8239. Nitrate reduction may be associated with inhibition of sporulation. © 2016 The Societies and John Wiley & Sons Australia, Ltd.

Novel Tissue Level Effects of the Staphylococcus aureus Enterotoxin Gene Cluster Are Essential for Infective Endocarditis.

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Stach, Christopher S; Vu, Bao G; Merriman, Joseph A; Herrera, Alfa; Cahill, Michael P; Schlievert, Patrick M; Salgado-Pabón, Wilmara

2016-01-01

Superantigens are indispensable virulence factors for Staphylococcus aureus in disease causation. Superantigens stimulate massive immune cell activation, leading to toxic shock syndrome (TSS) and contributing to other illnesses. However, superantigens differ in their capacities to induce body-wide effects. For many, their production, at least as tested in vitro, is not high enough to reach the circulation, or the proteins are not efficient in crossing epithelial and endothelial barriers, thus remaining within tissues or localized on mucosal surfaces where they exert only local effects. In this study, we address the role of TSS toxin-1 (TSST-1) and most importantly the enterotoxin gene cluster (egc) in infective endocarditis and sepsis, gaining insights into the body-wide versus local effects of superantigens. We examined S. aureus TSST-1 gene (tstH) and egc deletion strains in the rabbit model of infective endocarditis and sepsis. Importantly, we also assessed the ability of commercial human intravenous immunoglobulin (IVIG) plus vancomycin to alter the course of infective endocarditis and sepsis. TSST-1 contributed to infective endocarditis vegetations and lethal sepsis, while superantigens of the egc, a cluster with uncharacterized functions in S. aureus infections, promoted vegetation formation in infective endocarditis. IVIG plus vancomycin prevented lethality and stroke development in infective endocarditis and sepsis. Our studies support the local tissue effects of egc superantigens for establishment and progression of infective endocarditis providing evidence for their role in life-threatening illnesses. In contrast, TSST-1 contributes to both infective endocarditis and lethal sepsis. IVIG may be a useful adjunct therapy for infective endocarditis and sepsis.

Parenteral adjuvant potential of recombinant B subunit of Escherichia coli heat-labile enterotoxin

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Carlos Eduardo Pouey da Cunha

Full Text Available BACKGROUND The B subunit of Escherichia coli heat-labile enterotoxin (LTB is a potent mucosal immune adjuvant. However, there is little information about LTB's potential as a parenteral adjuvant. OBJECTIVES We aimed at evaluating and better understanding rLTB's potential as a parenteral adjuvant using the fused R1 repeat of Mycoplasma hyopneumoniae P97 adhesin as an antigen to characterise the humoral immune response induced by this construct and comparing it to that generated when aluminium hydroxide is used as adjuvant instead. METHODS BALB/c mice were immunised intraperitoneally with either rLTBR1 or recombinant R1 adsorbed onto aluminium hydroxide. The levels of systemic anti-rR1 antibodies (total Ig, IgG1, IgG2a, and IgA were assessed by enzyme-linked immunosorbent assay (ELISA. The ratio of IgG1 and IgG2a was used to characterise a Th1, Th2, or mixed Th1/Th2 immune response. FINDINGS Western blot confirmed rR1, either alone or fused to LTB, remained antigenic; anti-cholera toxin ELISA confirmed that LTB retained its activity when expressed in a heterologous system. Mice immunised with the rLTBR1 fusion protein produced approximately twice as much anti-rR1 immunoglobulins as mice vaccinated with rR1 adsorbed onto aluminium hydroxide. Animals vaccinated with either rLTBR1 or rR1 adsorbed onto aluminium hydroxide presented a mixed Th1/Th2 immune response. We speculate this might be a result of rR1 immune modulation rather than adjuvant modulation. Mice immunised with rLTBR1 produced approximately 1.5-fold more serum IgA than animals immunised with rR1 and aluminium hydroxide. MAIN CONCLUSIONS The results suggest that rLTB is a more powerful parenteral adjuvant than aluminium hydroxide when administered intraperitoneally as it induced higher antibody titres. Therefore, we recommend that rLTB be considered an alternative adjuvant, even if different administration routes are employed.

The influence of headspace and dissolved oxygen level on growth and haemolytic BL enterotoxin production of a psychrotolerant Bacillus weihenstephanensis isolate on potato based ready-to-eat food products.

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Samapundo, S; Everaert, H; Wandutu, J N; Rajkovic, A; Uyttendaele, M; Devlieghere, F

2011-04-01

The major objective of this study was to determine the influence of the initial headspace and dissolved O(2) level and vacuum packaging on growth and diarrhoeal enterotoxin production by Bacillus weihenstephanensis on potato based ready-to-eat food products. In general, the lower the initial headspace or dissolved O(2) level the slower the maximum growth rate (μ(max), log(10) CFU g(-1) d(-1)), the longer the lag phase duration (λ, d) and the smaller the maximum population density (N(max), log(10) CFU g(-1)) became. The slowest μ(max), the longest λ and the smallest N(max) were generally found for growth under vacuum packaging. This implies shorter shelf-lives will occur at higher initial headspace or dissolved O(2) levels as the growth of B. weihenstephanensis to the infective dose of 10(5) CFU g(-1) in such atmospheres takes a shorter time. Significant consumption of dissolved O(2) only occurred when growth shifted from the lag to the exponential phase and growth generally transitioned from the exponential to the stationary phase when the dissolved O(2) levels fell below ca. 75 ppb. Diarrhoeal enterotoxin production (determined via detection of the L2 component of haemolytic BL) was similar for growth under initial headspace O(2) levels of 1-20.9%, and was only reduced when growth took place under vacuum packaging. The reduction in L2 production when growth took place under vacuum was most probably related to the low final cell densities observed under this condition. Both growth and L2 production were inhibited over a 32-day incubation period at 7 °C by 40% CO(2) irrespective of the headspace or dissolved O(2) levels. The results illustrate the importance of residual O(2) and CO(2) on the shelf-stability and safety of modified atmosphere packaged potato based ready-to-eat food products with regards to B. weihenstephanensis. Copyright © 2010 Elsevier Ltd. All rights reserved.

Acetic acid increases the phage-encoded enterotoxin A expression in Staphylococcus aureus

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da Silva Ayla

2010-05-01

Full Text Available Abstract Background The effects of acetic acid, a common food preservative, on the bacteriophage-encoded enterotoxin A (SEA expression and production in Staphylococcus aureus was investigated in pH-controlled batch cultures carried out at pH 7.0, 6.5, 6.0, 5.5, 5.0, and 4.5. Also, genomic analysis of S. aureus strains carrying sea was performed to map differences within the gene and in the temperate phage carrying sea. Results The sea expression profile was similar from pH 7.0 to 5.5, with the relative expression peaking in the transition between exponential and stationary growth phase and falling during stationary phase. The levels of sea mRNA were below the detection limit at pH 5.0 and 4.5, confirmed by very low SEA levels at these pH values. The level of relative sea expression at pH 6.0 and 5.5 were nine and four times higher, respectively, in the transitional phase than in the exponential growth phase, compared to pH 7.0 and pH 6.5, where only a slight increase in relative expression in the transitional phase was observed. Furthermore, the increase in sea expression levels at pH 6.0 and 5.5 were observed to be linked to increased intracellular sea gene copy numbers and extracellular sea-containing phage copy numbers. The extracellular SEA levels increased over time, with highest levels produced at pH 6.0 in the four growth phases investigated. Using mitomycin C, it was verified that SEA was at least partially produced as a consequence of prophage induction of the sea-phage in the three S. aureus strains tested. Finally, genetic analysis of six S. aureus strains carrying the sea gene showed specific sea phage-groups and two versions of the sea gene that may explain the different sea expression and production levels observed in this study. Conclusions Our findings suggest that the increased sea expression in S. aureus caused by acetic acid induced the sea-encoding prophage, linking SEA production to the lifecycle of the phage.

Produção de enterotoxinas e da toxina da síndrome do choque tóxico por cepas de Staphylococcus aureus isoladas na mastite bovina Production of enterotoxins and toxic shock syndrome toxin by Staphylococcus aureus strains isolated from bovine mastitis

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A. Nader Filho

2007-10-01

Full Text Available A total of 72 strains of Staphylococcus aureus were examined for the production of staphylococcal enterotoxins (SE A, B, C, D and toxic shock syndrome toxin (TSST-1. The strains were isolated from milk samples from cows with mastitis in dairy herds of São Paulo State, Brazil. Off 72 isolates, 38 (52.8% produced SEA, 38 (52.8% SEB, 32 (44.4% SED, 28 (38.9% SEC and 27 (37.5% TSST-1. From the 72 strains, 66 (91.7% produced, at least, one or more toxin, including TSST-1.

Perfil de contaminação por Staphylococcus e suas enterotoxinas e monitorização das condições de higiene em uma linha de produção de queijo de coalho Contamination profile for staphylococci and its enterotoxins and monitorization of the conditions of hygiene in a ‘coalho' cheese production line

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Maria de Fatima Borges

2008-08-01

Full Text Available Avaliou-se o perfil de contaminação por Staphylococcus e suas enterotoxinas e monitorou-se as condições de higiene em uma linha de produção de queijo de coalho por meio da técnica de bioluminescência (ATP. A população de Staphylococcus sp. variou de This research aimed to evaluate the contamination by staphylococci and its enterotoxins as well as to monitor the conditions of hygiene from a coalho cheese production line, using ATP bioluminescence assay. Staphylococcus sp. population varied from <1CFU mL-1, in pasteurized milk to 1.5 x 107CFU mL-1, in raw milk, whereas coagulase-positive staphylococci count ranged from <1CFU mL-1, in pasteurized milk to 5.0 x 106CFU mL-1 in raw milk. Coagulase-positive staphylococci were detected in 100% (25/25 of the raw milk samples and in 8% (2/25 of cheese samples. Twelve Staphylococcus species were identified within the selected 68 isolates, being nine negative and three positive for coagulase. Raw milk samples showed a high rate of coagulase-positive, being S. aureus the most common, whereas other product samples and equipment surfaces, pieces of furniture, utensils and manipulator gloves samples presented a high frequency of coagulase-negative and low frequency of coagulase-positive. Staphylococcal enterotoxin was detected in 20% of the raw milk samples and therefore in pasteurized milk, curd and cheese. ATP measurement permitted to assess the effectiveness of the surfaces cleaning, being considered adequate in 62.1% (36/95, "alert state" in 23.2% (22/95 and inadequate in 14.7% (14/95 of surfaces evaluated. Detection of staphylococci species with enterotoxigenic potential as well as enterotoxin presence reveal dissemination of contamination at the "coalho" cheese production line, possibly due inappropriate Good Manufacturing Practices (GMP from the initial milking step until the final cheese production.

Effect of different ionizing radiation dose rates on the Staphylococcal enterotoxin in mechanically deboned chicken meat

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Azevedo, Heliana de; Brito, Poliana de Paula; Fukuma, Henrique Takuji; Roque, Claudio Vitor; Custodio, Wilson; Kodama, Yasko; Miya, Norma Terugo Nago; Pereira, Jose Luiz

2009-01-01

Samples weighing 50g each were prepared from allotments of back with skin MDCM, to the EEB contamination or not (control). Each sample of MDCM contaminated or not with EEB was conditioned in low density polyethylene bag, frozen (-18 ± 1 deg C) for one night in a tunnel and irradiated with gamma rays from 60 Co source in this state with doses of 0.0 kGy (control), 1.5 kGy (5.7 kGy.h -1 - higher dose rate, 1.8 kGy.h -1 - intermediary dose rate and 0.6 kGy.h -1 - lower dose rate) and 3.0 kGy (8.4 kGy.h - '1 - higher dose rate, 2.4 kGy.h -1 - intermediary dose rate and 0.6 kGy.h -1 - lower dose rate). Irradiated or non irradiated MDCM samples were processed to the EEB extraction, according to the VIDAS Staph enterotoxin II kit (bioMerieux) manufacturer protocol. The calculation to determinate the MDCM EEB recovery after the sample (control or irradiated) processing were carried out applying the principle of mass balance, along the whole process. Described experiment was performed in triplicate. Results showed that the irradiation process was effective to remove the MDCM EEB, to both 1.5 kGy and 3.0 kGy. According to the expected, doses of 3.0 kGy showed the highest values of MDCM EEB removal. Regarding the effect of dose rate of radiation on the removal of EEB of the MDCM, it could be observed only for samples irradiated with 1.5 kGy radiation dose; in these processing conditions, the highest value of EEB removal was obtained for samples processed with low radiation dose rate. (author)

Identification of novel Clostridium perfringens type E strains that carry an iota toxin plasmid with a functional enterotoxin gene.

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Kazuaki Miyamoto

Full Text Available Clostridium perfringens enterotoxin (CPE is a major virulence factor for human gastrointestinal diseases, such as food poisoning and antibiotic associated diarrhea. The CPE-encoding gene (cpe can be chromosomal or plasmid-borne. Recent development of conventional PCR cpe-genotyping assays makes it possible to identify cpe location (chromosomal or plasmid in type A isolates. Initial studies for developing cpe genotyping assays indicated that all cpe-positive strains isolated from sickened patients were typable by cpe-genotypes, but surveys of C. perfringens environmental strains or strains from feces of healthy people suggested that this assay might not be useful for some cpe-carrying type A isolates. In the current study, a pulsed-field gel electrophoresis Southern blot assay showed that four cpe-genotype untypable isolates carried their cpe gene on a plasmid of ∼65 kb. Complete sequence analysis of the ∼65 kb variant cpe-carrying plasmid revealed no intact IS elements and a disrupted cytosine methyltransferase (dcm gene. More importantly, this plasmid contains a conjugative transfer region, a variant cpe gene and variant iota toxin genes. The toxin genes encoded by this plasmid are expressed based upon the results of RT-PCR assays. The ∼65 kb plasmid is closely related to the pCPF4969 cpe plasmid of type A isolates. MLST analyses indicated these isolates belong to a unique cluster of C. perfringens. Overall, these isolates carrying a variant functional cpe gene and iota toxin genes represent unique type E strains.

Staphylococcal enterotoxin A regulates bone marrow granulocyte trafficking during pulmonary inflammatory disease in mice

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Takeshita, W.M.; Gushiken, V.O.; Ferreira-Duarte, A.P.; Pinheiro-Torres, A.S.; Roncalho-Buck, I.A. [Department of Biology and Physiology, Faculty of Medicine of Jundiai (FMJ), Jundiai, SP (Brazil); Squebola-Cola, D.M.; Mello, G.C.; Anhê, G.F.; Antunes, E. [Department of Pharmacology, Faculty of Medical Sciences, University of Campinas (UNICAMP), Campinas, SP (Brazil); DeSouza, I.A., E-mail: ivanidesouza@uol.com.br [Department of Biology and Physiology, Faculty of Medicine of Jundiai (FMJ), Jundiai, SP (Brazil)

2015-09-15

Pulmonary neutrophil infiltration produced by Staphylococcal enterotoxin A (SEA) airway exposure is accompanied by marked granulocyte accumulation in bone marrow (BM). Therefore, the aim of this study was to investigate the mechanisms of BM cell accumulation, and trafficking to circulating blood and lung tissue after SEA airway exposure. Male BALB/C mice were intranasally exposed to SEA (1 μg), and at 4, 12 and 24 h thereafter, BM, circulating blood, bronchoalveolar lavage (BAL) fluid and lung tissue were collected. Adhesion of BM granulocytes and flow cytometry for MAC-1, LFA1-α and VLA-4 and cytokine and/or chemokine levels were assayed after SEA-airway exposure. Prior exposure to SEA promoted a marked PMN influx to BAL and lung tissue, which was accompanied by increased counts of immature and/or mature neutrophils and eosinophils in BM, along with blood neutrophilia. Airway exposure to SEA enhanced BM neutrophil MAC-1 expression, and adhesion to VCAM-1 and/or ICAM-1-coated plates. Elevated levels of GM-CSF, G-CSF, INF-γ, TNF-α, KC/CXCL-1 and SDF-1α were detected in BM after SEA exposure. SEA exposure increased production of eosinopoietic cytokines (eotaxin and IL-5) and BM eosinophil VLA-4 expression, but it failed to affect eosinophil adhesion to VCAM-1 and ICAM-1. In conclusion, BM neutrophil accumulation after SEA exposure takes place by integrated action of cytokines and/or chemokines, enhancing the adhesive responses of BM neutrophils and its trafficking to lung tissues, leading to acute lung injury. BM eosinophil accumulation in SEA-induced acute lung injury may occur via increased eosinopoietic cytokines and VLA-4 expression. - Highlights: • Airway exposure to SEA causes acute lung inflammation. • SEA induces accumulation of bone marrow (BM) in immature and mature neutrophils. • SEA increases BM granulocyte or BM PMN adhesion to ICAM-1 and VCAM-1, and MAC-1 expression. • SEA induces BM elevations of CXCL-1, INF-γ, TNF-α, GM-CSF, G-CSF and

Staphylococcal enterotoxin A regulates bone marrow granulocyte trafficking during pulmonary inflammatory disease in mice

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Takeshita, W.M.; Gushiken, V.O.; Ferreira-Duarte, A.P.; Pinheiro-Torres, A.S.; Roncalho-Buck, I.A.; Squebola-Cola, D.M.; Mello, G.C.; Anhê, G.F.; Antunes, E.; DeSouza, I.A.

2015-01-01

Pulmonary neutrophil infiltration produced by Staphylococcal enterotoxin A (SEA) airway exposure is accompanied by marked granulocyte accumulation in bone marrow (BM). Therefore, the aim of this study was to investigate the mechanisms of BM cell accumulation, and trafficking to circulating blood and lung tissue after SEA airway exposure. Male BALB/C mice were intranasally exposed to SEA (1 μg), and at 4, 12 and 24 h thereafter, BM, circulating blood, bronchoalveolar lavage (BAL) fluid and lung tissue were collected. Adhesion of BM granulocytes and flow cytometry for MAC-1, LFA1-α and VLA-4 and cytokine and/or chemokine levels were assayed after SEA-airway exposure. Prior exposure to SEA promoted a marked PMN influx to BAL and lung tissue, which was accompanied by increased counts of immature and/or mature neutrophils and eosinophils in BM, along with blood neutrophilia. Airway exposure to SEA enhanced BM neutrophil MAC-1 expression, and adhesion to VCAM-1 and/or ICAM-1-coated plates. Elevated levels of GM-CSF, G-CSF, INF-γ, TNF-α, KC/CXCL-1 and SDF-1α were detected in BM after SEA exposure. SEA exposure increased production of eosinopoietic cytokines (eotaxin and IL-5) and BM eosinophil VLA-4 expression, but it failed to affect eosinophil adhesion to VCAM-1 and ICAM-1. In conclusion, BM neutrophil accumulation after SEA exposure takes place by integrated action of cytokines and/or chemokines, enhancing the adhesive responses of BM neutrophils and its trafficking to lung tissues, leading to acute lung injury. BM eosinophil accumulation in SEA-induced acute lung injury may occur via increased eosinopoietic cytokines and VLA-4 expression. - Highlights: • Airway exposure to SEA causes acute lung inflammation. • SEA induces accumulation of bone marrow (BM) in immature and mature neutrophils. • SEA increases BM granulocyte or BM PMN adhesion to ICAM-1 and VCAM-1, and MAC-1 expression. • SEA induces BM elevations of CXCL-1, INF-γ, TNF-α, GM-CSF, G-CSF and

Chimeric anti-staphylococcal enterotoxin B antibodies and lovastatin act synergistically to provide in vivo protection against lethal doses of SEB.

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Mulualem E Tilahun

Full Text Available Staphylococcal enterotoxin B (SEB is one of a family of toxins secreted by Staphylococcus aureus that act as superantigens, activating a large fraction of the T-cell population and inducing production of high levels of inflammatory cytokines that can cause toxic shock syndrome (TSS and death. Extracellular engagement of the TCR of T-cells and class II MHC of antigen presenting cells by SEB triggers the activation of many intracellular signaling processes. We engineered chimeric antibodies to block the extracellular engagement of cellular receptors by SEB and used a statin to inhibit intracellular signaling. Chimeric human-mouse antibodies directed against different neutralizing epitopes of SEB synergistically inhibited its activation of human T-cells in vitro. In the in vivo model of lethal toxic shock syndrome (TSS in HLA-DR3 transgenic mice, two of these antibodies conferred significant partial protection when administered individually, but offered complete protection in a synergistic manner when given together. Similarly, in vivo, lovastatin alone conferred only partial protection from TSS similar to single anti-SEB antibodies. However, used in combination with one chimeric neutralizing anti-SEB antibody, lovastatin provided complete protection against lethal TSS in HLA-DR3 transgenic mice. These experiments demonstrate that in vivo protection against lethal doses of SEB can be achieved by a statin of proven clinical safety and chimeric human-mouse antibodies, agents now widely used and known to be of low immunogenicity in human hosts.

Organization and ELISA-Based Results of the First Proficiency Testing to Evaluate the Ability of European Union Laboratories to Detect Staphylococcal Enterotoxin Type B (SEB in Buffer and Milk

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Yacine Nia

2016-09-01

Full Text Available The aim of this work was to organize the first proficiency test (PT dedicated to staphylococcal enterotoxin B (SEB detection in milk and buffer solutions. This paper describes the organization of the PT trial according to EN ISO 17043 requirements. Characterization of the SEB stock solution was performed using SDS-PAGE and SE-specific ELISA, and amino acid analysis was used to assign its protein concentration. The solution was then used to prepare six PT materials (four milk and two buffer batches at a ng/g toxin level, which included one blank and one SEA-containing milk as specificity control. Suitable material homogeneity and stability were assessed using screening and quantitative ELISAs. Among the methods used by the participants, ELISA-based methods demonstrated their efficiency for the detection of SEB in both simple and complex matrices. The results serve as a basis for further improving the detection capabilities in expert laboratories and can therefore be considered as a contribution to biopreparedness.

Food-Borne Outbreak Investigation and Molecular Typing: High Diversity of Staphylococcus aureus Strains and Importance of Toxin Detection

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Denayer, Sarah; Nia, Yacine; Botteldoorn, Nadine

2017-01-01

Staphylococcus aureus is an important aetiological agent of food intoxications in the European Union as it can cause gastro-enteritis through the production of various staphylococcal enterotoxins (SEs) in foods. Reported enterotoxin dose levels causing food-borne illness are scarce and varying. Three food poisoning outbreaks due to enterotoxin-producing S. aureus strains which occurred in 2013 in Belgium are described. The outbreaks occurred in an elderly home, at a barbecue event and in a kindergarten and involved 28, 18, and six cases, respectively. Various food leftovers contained coagulase positive staphylococci (CPS). Low levels of staphylococcal enterotoxins ranging between 0.015 ng/g and 0.019 ng/g for enterotoxin A (SEA), and corresponding to 0.132 ng/g for SEC were quantified in the food leftovers for two of the reported outbreaks. Molecular typing of human and food isolates using pulsed-field gel electrophoresis (PFGE) and enterotoxin gene typing, confirmed the link between patients and the suspected foodstuffs. This also demonstrated the high diversity of CPS isolates both in the cases and in healthy persons carrying enterotoxin genes encoding emetic SEs for which no detection methods currently exist. For one outbreak, the investigation pointed out to the food handler who transmitted the outbreak strain to the food. Tools to improve staphylococcal food poisoning (SFP) investigations are presented. PMID:29261162

Food-Borne Outbreak Investigation and Molecular Typing: High Diversity of Staphylococcus aureus Strains and Importance of Toxin Detection.

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Denayer, Sarah; Delbrassinne, Laurence; Nia, Yacine; Botteldoorn, Nadine

2017-12-20

Staphylococcus aureus is an important aetiological agent of food intoxications in the European Union as it can cause gastro-enteritis through the production of various staphylococcal enterotoxins (SEs) in foods. Reported enterotoxin dose levels causing food-borne illness are scarce and varying. Three food poisoning outbreaks due to enterotoxin-producing S. aureus strains which occurred in 2013 in Belgium are described. The outbreaks occurred in an elderly home, at a barbecue event and in a kindergarten and involved 28, 18, and six cases, respectively. Various food leftovers contained coagulase positive staphylococci (CPS). Low levels of staphylococcal enterotoxins ranging between 0.015 ng/g and 0.019 ng/g for enterotoxin A (SEA), and corresponding to 0.132 ng/g for SEC were quantified in the food leftovers for two of the reported outbreaks. Molecular typing of human and food isolates using pulsed-field gel electrophoresis (PFGE) and enterotoxin gene typing, confirmed the link between patients and the suspected foodstuffs. This also demonstrated the high diversity of CPS isolates both in the cases and in healthy persons carrying enterotoxin genes encoding emetic SEs for which no detection methods currently exist. For one outbreak, the investigation pointed out to the food handler who transmitted the outbreak strain to the food. Tools to improve staphylococcal food poisoning (SFP) investigations are presented.

Safety and efficacy of E coli enterotoxin adjuvant for urease-based rectal immunization against Helicobacter pylori.

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Sougioultzis, Stavros; Lee, Cynthia K; Alsahli, Mazen; Banerjee, Subhas; Cadoz, Michel; Schrader, Robert; Guy, Bruno; Bedford, Philip; Monath, Thomas P; Kelly, Ciaran P; Michetti, Pierre

2002-12-13

Low dose E. coli heat-labile enterotoxin (LT), delivered orally or enterically, has been used as an adjuvant for Helicobacter pylori (H. pylori) urease in healthy adults. In this study we aim to test the safety and adjuvant efficacy of LT delivered rectally together with recombinant H. pylori urease. Eighteen healthy adults without present or past H. pylori infection were enrolled in a double blind, randomized, ascending dose study to receive either urease (60 mg), or urease (60 mg) + LT (5 or 25 microg). The immunization preparation was administered per rectum on days 0, 14 and 28. Serum, stool and saliva anti-urease and anti-LT IgG and IgA antibodies (Abs) were measured and urease-specific and LT-specific antigen secreting cells (ASCs) were counted in peripheral blood at baseline and 7 (ASC counts) or 14 days (antibody levels) after each dosing. Peripheral blood lymphoproliferation assays were also performed at baseline and at the end of the study. Rectally delivered urease and LT were well tolerated. Among the 12 subjects assigned to urease+LT, 2 (16.7%) developed anti-urease IgG Abs, 1 (8.3%) developed anti-urease IgA Abs, and 3 (25%) showed urease-specific IgA(+) ASCs. Immune responses to LT were more vigorous, especially in subjects exposed to 5 microg LT. In the urease+ 5 microg LT group, anti-LT IgG and IgA Abs developed in 60 and 80% of the subjects, respectively, while LT-specific IgG(+) and IgA(+) ASCs were detected in all subjects. The magnitude of the anti-LT response was much higher than the response to urease. No IgA anti-urease or anti-LT Abs were detected in stool or saliva and lymphocyte proliferative responses to urease were unsatisfactory. In conclusion, rectal delivery of 5 microg LT is safe and induces vigorous systemic anti-LT immune responses. Further studies are needed to determine if LT can be an effective adjuvant for rectally delivered antigens.

Distinct cytokine profiles of circulating mononuclear cells stimulated with Staphylococcus aureus enterotoxin A in vitro during early and late episodes of chronic osteomyelitis

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Gabriella Freitas Ferreira

2012-05-01

Full Text Available We investigated the cytokine profile of peripheral mononuclear cells from chronic osteomyelitis (OST patients following in vitro stimulation with staphylococcal enterotoxin A (SEA. We demonstrate that stimulation with SEA induced prominent lymphocyte proliferation and high levels of tumour necrosis factor (TNF-α, interleukin (IL-4 and IL-10 secretion in both OST and non-infected individuals (NI. Even though stimulation with SEA had no impact on IL-6 production in either patient group, the baseline level of IL-6 production by cells from OST patients was always significantly less than that produced by cells from NI. After classifying the osteomyelitic episodes based on the time after the last reactivation event as "early" (1-4 months or "late" osteomyelitis (5-12 months, we found that increased levels of TNF-α and IL-4 in combination with decreased levels of IL-6 were observed in the early episodes. By contrast, increased levels of IL-10, IL-2 and IL-6 were hallmarks of late episodes. Our data demonstrate that early osteomyelitic episodes are accompanied by an increased frequency of "high producers" of TNF-α and IL-4, whereas late events are characterised by increased frequencies of "high producers" of IL-10, IL-6 and IL-2. These findings demonstrate the distinct cytokine profiles in chronic osteomyelitis, with a distinct regulation of IL-6 production during early and late episodes.

Distribusi Gen Enterotoksin Staphylococcus aureus dari Susu Segar dan Pangan Asal Hewan

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Siti Isrina Oktavia Salasia

2009-09-01

Full Text Available Staphylococcus aureus is a potential pathogen causing disease in human and animals due to severalvirulence factors. Staphylococcal enterotoxins which is responsible for foodborne disease is considered tobe one of the important virulence factor for the bacteria. The research was conducted to identify variousenterotoxin genes of S. aureus. Twenty three S. aureus isolates from milk cows (12 isolates and foodanimal products (11 isolates were used to study various enterotoxins genes (sea, seb, sec, sed, see, seg, seh,sei, and sej. The enterotoxins genes of S. aureus were investigated by using polymerase chain reaction(PCR with specific primers. There were 3 isolates (13.04% negative for staphylococcal enterotoxin genes.Twenty isolates (86.96% harboured for one or more staphylococcal enterotoxin genes such as: sec (6 isolates/26.09%, see, seh for 1 isolate (4.35%, combination of 2 genes se (b,i, se (c,g, se (g,i for 1 isolate of each(4.35%, se(c,e for 2 isolates (8.70%, se(b,c for 4 isolates (17.39%. Staphylococcal enterotoxin could bedetected in 3 combination genes of se(b,c,i, se(c,e,i, se(c,g,i for 1 isolate of each (4.35%.

Frequency of enterotoxins, toxic shock syndrome toxin-1, and biofilm formation genes in Staphylococcus aureus isolates from cows with mastitis in the Northeast of Brazil.

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Costa, F N; Belo, N O; Costa, E A; Andrade, G I; Pereira, L S; Carvalho, I A; Santos, R L

2018-06-01

Staphylococcus aureus is among the microorganisms more frequently associated with subclinical bovine mastitis. S. aureus may produce several virulence factors. This study aimed at determining the frequency of virulence factors such as enterotoxins, toxic shock syndrome toxin 1, and ica adhesion genes. In addition, we assessed antimicrobial drug resistance in S. aureus isolated from clinical and subclinical cases of mastitis. A total of 88 cows with clinical or subclinical mastitis were sampled, resulting in 38 S. aureus isolates, from which 25 (65.78%) carried toxin genes, including seb, sec, sed, tst, and icaD adhesion gene. These S. aureus isolates belong to 21 ribotypes and three S. aureus strains belonged to the same ribotype producing ica adhesion gene. Approximately 90% of S. aureus strains obtained in our study demonstrated multiple resistance to different antimicrobial agents. The most efficacious antimicrobial agents against the isolates were gentamicin, amoxicillin, and norfloxacin. Gentamicin was the most efficacious agent inhibiting 78.95% of the S. aureus isolates. The least efficacious were penicillin, streptomycin, and ampicillin. Our results can help in understanding the relationship between virulence factors and subclinical mastitis caused by S. aureus. Further research about diversity of S. aureus isolates and genes responsible for the pathogenicity of subclinical mastitis is essential.

Staphylococcal enterotoxin-like X (SElX is a unique superantigen with functional features of two major families of staphylococcal virulence factors.

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Ries J Langley

2017-09-01

Full Text Available Staphylococcus aureus is an opportunistic pathogen that produces many virulence factors. Two major families of which are the staphylococcal superantigens (SAgs and the Staphylococcal Superantigen-Like (SSL exoproteins. The former are immunomodulatory toxins that induce a Vβ-specific activation of T cells, while the latter are immune evasion molecules that interfere with a wide range of innate immune defences. The superantigenic properties of Staphylococcal enterotoxin-like X (SElX have recently been established. We now reveal that SElX also possesses functional characteristics of the SSLs. A region of SElX displays high homology to the sialyl-lactosamine (sLacNac-specific binding site present in a sub-family of SSLs. By analysing the interaction of SElX with sLacNac-containing glycans we show that SElX has an equivalent specificity and host cell binding range to the SSLs. Mutation of key amino acids in this conserved region affects the ability of SElX to bind to cells of myeloid origin and significantly reduces its ability to protect S. aureus from destruction in a whole blood killing (WBK assay. Like the SSLs, SElX is up-regulated early during infection and is under the control of the S. aureus exotoxin expression (Sae two component gene regulatory system. Additionally, the structure of SElX in complex with the sLacNac-containing tetrasaccharide sialyl Lewis X (sLeX reveals that SElX is a unique single-domain SAg. In summary, SElX is an 'SSL-like' SAg.

Detection of target staphylococcal enterotoxin B antigen in orange juice and popular carbonated beverages using antibody-dependent antigen-capture assays.

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Principato, MaryAnn; Njoroge, Joyce M; Perlloni, Andrei; O' Donnell, Michael; Boyle, Thomas; Jones, Robert L

2010-10-01

There is a critical need for qualitative and quantitative methodologies that provide the rapid and accurate detection of food contaminants in complex food matrices. However, the sensitivity of the assay can be affected when antigen-capture is applied to certain foods or beverages that are extremely acidic. This study was undertaken to assess the effects of orange juice and popular carbonated soft drink upon the fidelity of antibody-based antigen-capture assays and to develop simple approaches that could rescue assay performance without the introduction of additional or extensive extraction procedures. We examined the effects of orange juice and a variety of popular carbonated soft drink beverages upon a quantitative Interleukin-2 (IL-2) enzyme-linked immunosorbent assay (ELISA) assay system and a lateral flow device (LFD) adapted for the detection of staphylococcal enterotoxin B (SEB) in foods. Alterations in the performance and sensitivity of the assay were directly attributable to the food matrix, and alterations in pH were especially critical. The results demonstrate that approaches such as an alteration of pH and the use of milk as a blocking agent, either singly or in combination, will partially rescue ELISA performance. The same approaches permit lateral flow to efficiently detect antigen. Practical Application: The authors present ways to rescue an ELISA assay compromised by acidity in beverages and show that either the alteration of pH, or the use of milk as a blocking agent are not always capable of restoring the assay to its intended efficiency. However, the same methods, when employed with lateral flow technology, are rapid and extremely successful.

Isolation of a new ssDNA aptamer against staphylococcal enterotoxin B based on CNBr-activated sepharose-4B affinity chromatography.

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Hedayati Ch, Mojtaba; Amani, Jafar; Sedighian, Hamid; Amin, Mohsen; Salimian, Jafar; Halabian, Raheleh; Imani Fooladi, Abbas Ali

2016-09-01

Staphylococcus aureus are potent human pathogens possessing arsenal of virulence factors. Staphylococcal food poisoning (SFP) and respiratory infections mediated by staphylococcal enterotoxin B (SEB) are common clinical manifestations. Many diagnostic techniques are based on serological detection and quantification of SEB in different food and clinical samples. Aptamers are known as new therapeutic and detection tools which are available in different ssDNA, dsDNA and protein structures. In this study, we used a new set of ssDNA aptamers against SEB. The methods used included preparation of a dsDNA library using standard SEB protein as the target analyte, affinity chromatography matrix in microfuge tubes, SELEX procedures to isolate specific ssDNA-aptamer as an affinity ligand, aptamer purification using ethanol precipitation method, affinity binding assay using ELISA, aptamer cloning and specificity test. Among 12 readable sequences, three of them were selected as the most appropriate aptamer because of their affinity and specificity to SEB. This study presents a new set of ssDNA aptamer with favorable selectivity to SEB through 12 rounds of SELEX. Selected aptamers were used to detect SEB in infected serum samples. Results showed that SEB c1 aptamer (2 µg SEB/100 nM aptamer) had favorable specificity to SEB (kd  = 2.3 × 10(-11) ). In conclusion, aptamers can be considered as useful tools for detecting and evaluating SEB. The results showed that affinity chromatography was an affordable assay with acceptable accuracy to isolate sensitive and selective novel aptamers. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

Review Over a 3-Year Period of European Union Proficiency Tests for Detection of Staphylococcal Enterotoxins in Food Matrices.

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Nia, Yacine; Mutel, Isabelle; Assere, Adrien; Lombard, Bertrand; Auvray, Frederic; Hennekinne, Jacques-Antoine

2016-04-13

Staphylococcal food poisoning outbreaks are a major cause of foodborne illnesses in Europe and their notifications have been mandatory since 2005. Even though the European regulation on microbiological criteria for food defines a criterion on staphylococcal enterotoxin (SE) only in cheese and dairy products, European Food Safety Authority (EFSA) data reported that various types of food matrices are involved in staphylococcal food poisoning outbreaks. The European Screening Method (ESM) of European Union Reference Laboratory for Coagulase Positive Staphylococci (EURL CPS) was validated in 2011 for SE detection in food matrices and is currently the official method used for screening purposes in Europe. In this context, EURLCPS is annually organizing Inter-Laboratory Proficiency Testing Trials (ILPT) to evaluate the competency of the European countries' National Reference Laboratories (NRLs) to analyse SE content in food matrices. A total of 31 NRLs representing 93% of European countries participated in these ILPTs. Eight food matrices were used for ILPT over the period 2013-2015, including cheese, freeze-dried cheese, tuna, mackerel, roasted chicken, ready-to-eat food, milk, and pastry. Food samples were spiked with four SE types (i.e., SEA, SEC, SED, and SEE) at various concentrations. Homogeneity and stability studies showed that ILPT samples were both homogeneous and stable. The analysis of results obtained by participants for a total of 155 blank and 620 contaminated samples allowed for evaluation of trueness (>98%) and specificity (100%) of ESM. Further to the validation study of ESM carried out in 2011, these three ILPTs allowed for the assessment of the proficiency of the NRL network and the performance of ESM on a large variety of food matrices and samples. The ILPT design presented here will be helpful for the organization of ILPT on SE detection by NRLs or other expert laboratories.

Review Over a 3-Year Period of European Union Proficiency Tests for Detection of Staphylococcal Enterotoxins in Food Matrices

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Yacine Nia

2016-04-01

Full Text Available Staphylococcal food poisoning outbreaks are a major cause of foodborne illnesses in Europe and their notifications have been mandatory since 2005. Even though the European regulation on microbiological criteria for food defines a criterion on staphylococcal enterotoxin (SE only in cheese and dairy products, European Food Safety Authority (EFSA data reported that various types of food matrices are involved in staphylococcal food poisoning outbreaks. The European Screening Method (ESM of European Union Reference Laboratory for Coagulase Positive Staphylococci (EURL CPS was validated in 2011 for SE detection in food matrices and is currently the official method used for screening purposes in Europe. In this context, EURLCPS is annually organizing Inter-Laboratory Proficiency Testing Trials (ILPT to evaluate the competency of the European countries’ National Reference Laboratories (NRLs to analyse SE content in food matrices. A total of 31 NRLs representing 93% of European countries participated in these ILPTs. Eight food matrices were used for ILPT over the period 2013–2015, including cheese, freeze-dried cheese, tuna, mackerel, roasted chicken, ready-to-eat food, milk, and pastry. Food samples were spiked with four SE types (i.e., SEA, SEC, SED, and SEE at various concentrations. Homogeneity and stability studies showed that ILPT samples were both homogeneous and stable. The analysis of results obtained by participants for a total of 155 blank and 620 contaminated samples allowed for evaluation of trueness (>98% and specificity (100% of ESM. Further to the validation study of ESM carried out in 2011, these three ILPTs allowed for the assessment of the proficiency of the NRL network and the performance of ESM on a large variety of food matrices and samples. The ILPT design presented here will be helpful for the organization of ILPT on SE detection by NRLs or other expert laboratories.

Involvement of protein kinase C in the mechanism of action of Escherichia coli heat-stable enterotoxin (STa) in a human colonic carcinoma cell line, COLO-205

International Nuclear Information System (INIS)

Gupta, Dyuti Datta; Saha, Subhrajit; Chakrabarti, Manoj K.

2005-01-01

The present study was undertaken to determine the involvement of calcium-protein kinase C pathway in the mechanism of action of Escherichia coli heat stable enterotoxin (STa) apart from STa-induced activation of guanylate cyclase in human colonic carcinoma cell line COLO-205, which was used as a model cultured cell line to study the mechanism of action of E. coli STa. In response to E. coli STa, protein kinase C (PKC) activity was increased in a time-dependent manner with its physical translocation from cytosol to membrane. Inhibition of the PKC activity in membrane fraction and inhibition of its physical translocation in response to IP 3 -mediated calcium release inhibitor dantrolene suggested the involvement of intracellular store depletion in the regulation of PKC activity. Among different PKC isoforms, predominant involvement of calcium-dependent protein kinase C (PKCα) was specified using isotype-specific pseudosubstrate, which showed pronounce enzyme activity. Inhibition of enzyme activity by PKCα-specific inhibitor Goe6976 and immunoblott study employing isotype-specific antibody further demonstrated the involvement of calcium-dependent isoform of PKC in the mechanism of action of E. coli STa. Moreover, inhibition of guanylate cyclase activity by PKCα-specific inhibitor Goe6976 suggested the involvement of PKCα in the regulation of guanylate cyclase activity

Experimental investigation into the effects of irradiation with mixed neutron-gamma-rays on the immune system as demonstrated by the model of immunity to Staphylococcal Enterotoxin B in rabbits

International Nuclear Information System (INIS)

Kraus, W.

1984-01-01

Using the model of immunisation of rabbits with Staphylococcal Enterotoxin B the influence of neutron radiation on the antibody response was investigated. Four groups were formed, which differed by the moment of irradiation with regard to the moment of immunisation and re-immunisation, and were irradiated with neutron-rays of 200 rad and 300 rad doses. Both doses were each given before and after vaccination, respectively before and after re-vaccination. The antibody response has been evaluated by the Radio-Immuno-Assay (RIA): Neutron-radiation given 24 hours before vaccination effects a stronger immune suppression than given 24 hours after vaccination. By pre-antigenic radiation a secondary immune response is more suppressed than by radiation following the antigen; the recovery of lymphoid tissue, however, takes place in a shorter time than in case of radiation before or after a primary vaccination. The secondary antibody response to a re-vaccination is not influenced by a subsequent neutron-radiation. The formation of secondary antibodies is not as radiosensitive as the formation of primary ones. By a neutron-radiation preformed antibodies are hardly damaged. (orig./MG) [de

[Escherichia coli heat-labile enterotoxin B subunit enhances the immune response against canine parvovirus VP2 in mice immunized by VP2 DNA vaccine].

Science.gov (United States)

Han, Dongmei; Zhong, Fei; Li, Xiujin; Wang, Wei; Wang, Xingxing; Pan, Sumin

2011-01-01

To investigate the effect of Escherichia coli heat-labile enterotoxin (LT) B subunit (LTB) gene on canine parvovirus (CPV) VP2 gene vaccine. The LTB gene was amplified by PCR from genomic DNA of E. coli 44815 strain. The VP2-70 fragment (210 bp) encoding major epitope of VP2 (70 amino acids) was amplified by PCR from a plasmid encoding VP2 gene. VP2-70 and LTB genes were inserted into the eukaryotic vector to construct VP2-70 gene,LTB gene and VP2-70-LTB fused gene vectors. The mice were immunized with VP2-70 vector, VP2-70-LTB fused vector, or VP2-70 vector plus LTB vector, respectively. The antibody titers at the different time were measured by using ELISA method. The spleen lymphocyte proliferation activity was analyzed by 3-(4, 5-Dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay. The sequence of VP2-70 and LTB genes was identified. The recombinant VP2-70 and LTB proteins could be expressed in HEK293T cells in a secretory manner. The mice immunized with VP2-70 vector, VP2-70-LTB vector or VP2-70 vector plus LTB vector could generate the specific antibody against VP2 protein. The antibody titer immunized with VP2-70-LTB vector reached 1:5120 at 35 d post immunization, significantly higher than that of other two groups (P vaccine in mice.

The immunogenic SigA enterotoxin of Shigella flexneri 2a binds to HEp-2 cells and induces fodrin redistribution in intoxicated epithelial cells.

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Keith Al-Hasani

Full Text Available BACKGROUND: We have previously shown that the enterotoxin SigA which resides on the she pathogenicity island (PAI of S. flexneri 2a is an autonomously secreted serine protease capable of degrading casein. We have also demonstrated that SigA is cytopathic for HEp-2 cells and plays a role in the intestinal fluid accumulation associated with S. flexneri infections. METHODS/PRINCIPAL FINDINGS: In this work we show that SigA binds specifically to HEp-2 cells and degrades recombinant human alphaII spectrin (alpha-fodrin in vitro, suggesting that the cytotoxic and enterotoxic effects mediated by SigA are likely associated with the degradation of epithelial fodrin. Consistent with our data, this study also demonstrates that SigA cleaves intracellular fodrin in situ, causing its redistribution within cells. These results strongly implicate SigA in altering the cytoskeleton during the pathogenesis of shigellosis. On the basis of these findings, cleavage of fodrin is a novel mechanism of cellular intoxication for a Shigella toxin. Furthermore, information regarding immunogenicity to SigA in infected patients is lacking. We studied the immune response of SigA from day 28 post-challenge serum of one volunteer from S. flexneri 2a challenge studies. Our results demonstrate that SigA is immunogenic following infection with S. flexneri 2a. CONCLUSIONS: This work shows that SigA binds to epithelial HEp-2 cells as well as being able to induce fodrin degradation in vitro and in situ, further extending its documented role in the pathogenesis of Shigella infections.

Effects of tetrodotoxin and ion replacements on the short-circuit current induced by Escherichiacoli heat stable enterotoxin across small intestine of the gerbil (Gerbillus cheesmani

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Fawzia Yaqoub Al-Balool

2004-03-01

Full Text Available The effects of mucosally added Escherichia coli heat stable enterotoxin (STa 30 ng ml-1 on the basal short-circuit current (Isc in µA cm-2 across stripped and unstripped sheets of jejuna and ilea taken from fed, starved (4 days, water ad lib and undernourished (50% control food intake for 21 days gerbil (Gerbillus cheesmani were investigated. The effect of neurotoxin tetrodotoxin (TTX 10 µM and the effects of replacing chloride by gluconate or the effects of removing bicarbonate from bathing buffers on the maximum increase in Isc induced by STa were also investigated. The maximum increase in Isc which resulted from the addition of STa were significantly higher in jejuna and ilea taken from starved and undernourished gerbils when compared with the fed control both using stripped and unstripped sheets. In the two regions of the small intestine taken from fed and starved animals TTX reduced the maximum increase in Isc induced by STa across unstripped sheets only. Moreover in jejuna and ilea taken from undernourished gerbils TTX reduced significantly the maximum increase in Isc induced by STa across stripped and unstripped sheets. Replacing chloride by gluconate decreased the maximum increase in Isc induced by STa across jejuna and ilea taken from undernourished gerbils only. Removing bicarbonates from bathing buffer decreased the maximum increase in Isc across the jejuna and ilea taken from starved and undernourished gerbils.

Chromosomal and plasmid-encoded factors of Shigella flexneri induce secretogenic activity ex vivo.

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Christina S Faherty

Full Text Available Shigella flexneri is a Gram-negative, facultative intracellular pathogen that causes millions of cases of watery or bloody diarrhea annually, resulting in significant global mortality. Watery diarrhea is thought to arise in the jejunum, and subsequent bloody diarrhea occurs as a result of invasion of the colonic epithelium. Previous literature has demonstrated that Shigella encodes enterotoxins, both chromosomally and on the 220 kilobase virulence plasmid. The ShigellaEnterotoxins 1 and 2 (ShET1 and ShET2 have been shown to increase water accumulation in the rabbit ileal loop model. In addition, these toxins increase the short circuit current in rabbit tissue mounted in Ussing chambers, which is a model for the ion exchange that occurs during watery diarrhea. In this study, we sought to validate the use of mouse jejunum in Ussing chamber as an alternative, more versatile model to study bacterial pathogenesis. In the process, we also identified enterotoxins in addition to ShET1 and ShET2 encoded by S. flexneri. Through analysis of proteins secreted from wildtype bacteria and various deletion mutants, we have identified four factors responsible for enterotoxin activity: ShET1 and Pic, which are encoded on the chromosome; ShET2 (encoded by sen or ospD3, which requires the type-III secretion system for secretion; and SepA, an additional factor encoded on the virulence plasmid. The use of mouse jejunum serves as a reliable and reproducible model to identify the enterotoxins elaborated by enteric bacteria. Moreover, the identification of all Shigella proteins responsible for enterotoxin activity is vital to our understanding of Shigella pathogenicity and to our success in developing safe and effective vaccine candidates.

Identification of Staphylococcus spp. isolated during the ripening process of a traditional minas cheese

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B.M. Borelli

2011-04-01

Full Text Available The population dynamics of Staphylococcus spp. was studied during the ripening of Canastra Minas cheese at three farms located in the State of Minas Gerais, Brazil. The presence of coagulase (coa, thermonuclease (nuc, and enterotoxin (sea, seb, sec, and sed genes was investigated in Staphylococcus strains isolated during the 60-day cheese-ripening period. The presence of the staphylococcal enterotoxins A, C, and D was also investigated in the cheese samples. Cheese samples that were matured for 0, 7, 15, 30, and 45 days presented staphylococci counts from 10³ to 10(8cfu/g. All isolates considered coagulase-positive by physiological tests had the coa gene. However, no association was observed between the results obtained with biochemical tests and those obtained by PCR using gene-specific primers for coagulase-negative strains. Coagulase and thermonuclease genes occurred simultaneously in 41.3% of Staphylococcus spp. tested. None of the investigated Staphylococcus strains expressed enterotoxins SEA, SEB, SEC, and SED. Enterotoxins A, C, and D were not detected in any of the cheese samples.

ANIMAL ENTEROTOXIGENIC ESCHERICHIA COLI

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Dubreuil, J. Daniel; Isaacson, Richard E.; Schifferli, Dieter M.

2016-01-01

Enterotoxigenic Escherichia coli (ETEC) is the most common cause of E. coli diarrhea in farm animals. ETEC are characterized by the ability to produce two types of virulence factors; adhesins that promote binding to specific enterocyte receptors for intestinal colonization and enterotoxins responsible for fluid secretion. The best-characterized adhesins are expressed in the context of fimbriae, such as the F4 (also designated K88), F5 (K99), F6 (987P), F17 and F18 fimbriae. Once established in the animal small intestine, ETEC produces enterotoxin(s) that lead to diarrhea. The enterotoxins belong to two major classes; heat-labile toxin that consist of one active and five binding subunits (LT), and heat-stable toxins that are small polypeptides (STa, STb, and EAST1). This chapter describes the disease and pathogenesis of animal ETEC, the corresponding virulence genes and protein products of these bacteria, their regulation and targets in animal hosts, as well as mechanisms of action. Furthermore, vaccines, inhibitors, probiotics and the identification of potential new targets identified by genomics are presented in the context of animal ETEC. PMID:27735786

Genome sequencing and molecular characterisation of Staphylococcus aureus ST772-MRSA-V, "Bengal Bay Clone".

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Monecke, Stefan; Baier, Vico; Coombs, Geoffrey W; Slickers, Peter; Ziegler, Albrecht; Ehricht, Ralf

2013-12-20

The PVL-positive ST772-MRSA-V is an emerging community-associated (CA-) MRSA clone that has been named Bengal Bay Clone since most patients have epidemiological connections to the Indian subcontinent. It is found increasingly common in other areas of the world. One isolate of ST772-MRSA-V was sequenced using the Illumina Genome Analyzer System. After initial assembling the multiple sequence contigs were analysed using different in-house annotation scripts. Results were compared to microarray hybridisation results of clinical isolates of ST772-MRSA-V, of related strains and to another ST772-MRSA-V genome sequence. According to MLST e-burst analysis, ST772-MRSA-V belongs to Clonal Complex (CC)1, differing from ST1 only in one MLST allele (pta-22). However, there are several additional differences including agr alleles (group II rather than III), capsule type (5 rather than 8), the presence of the egc enterotoxin gene cluster and of the enterotoxin homologue ORF CM14 as well as the absence of the enterotoxin H gene seh. Enterotoxin genes sec and sel are present. ST772-MRSA-V harbours the genes encoding enterotoxin A (sea) and PVL (lukS/F-PV). Both are located on the same prophage. ST772-MRSA-V may have emerged from the same lineage as globally spread CC1 and CC5 strains. It has acquired a variety of virulence factors, and for a CA-MRSA strain it has an unusually high number of genes associated with antibiotic resistance.

Up-regulation of granzyme B and perforin by staphylococcal enterotoxin C2 mutant induces enhanced cytotoxicity in Hepa1–6 cells

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Zhang, Guojun [Institute of Applied Ecology, Chinese Academy of Sciences, No.72 Wenhua Road Shenhe Dis., Shenyang, Liaoning (China); University of Chinese Academy of Sciences, Beijing (China); Xu, Mingkai, E-mail: mkxu@iae.ac.cn [Institute of Applied Ecology, Chinese Academy of Sciences, No.72 Wenhua Road Shenhe Dis., Shenyang, Liaoning (China); Zhang, Huiwen [Institute of Applied Ecology, Chinese Academy of Sciences, No.72 Wenhua Road Shenhe Dis., Shenyang, Liaoning (China); Song, Yubo [Institute of Applied Ecology, Chinese Academy of Sciences, No.72 Wenhua Road Shenhe Dis., Shenyang, Liaoning (China); University of Chinese Academy of Sciences, Beijing (China); Wang, Jian; Zhang, Chenggang [Institute of Applied Ecology, Chinese Academy of Sciences, No.72 Wenhua Road Shenhe Dis., Shenyang, Liaoning (China)

2016-12-15

Staphylococcal enterotoxin C2 (SEC2), a member of bacterial superantigen, is one of the most potent known activators of T lymphocytes. With this property, SEC2 has already been used in clinic as a tumor immunotherapy agent in China. To increase the antitumor activity, a SEC2 mutant named ST-4 (GKVTG102-106WWH) with amino acid substitutions in T cell receptor (TCR)-binding domain was generated by site-directed mutagenesis, and the molecular mechanism of the enhanced antitumor activity was investigated. Results showed that ST-4 could activate much more Vβ 8.2 and 8.3 T cells and NK cells compared with SEC2, and exhibited significantly enhanced immunocyte stimulation and antitumor activity in vitro. The synthetic peptide sequencing the residues of mutant TCR-binding domain could competitively inhibit the immunocyte stimulation activity of ST-4. Most importantly, ST-4 up-regulated granzyme B and perforin at both mRNA and protein levels. We also found that expression of proapoptotic proteins cytochrome c, BAX and activation of caspase-3, 9 was up-regulated, and antiapoptotic protein Bcl-xL was down-regulated in the treatment with either ST-4 or SEC2. When granzyme B inhibitor or perforin inhibitor is presented, tumor cell viability was significantly rescued. Taken together, we demonstrate that increased ST-4-TCR recognition contributed to massive T cells and NK cells activation. These activated cells released up-regulated granzyme B and perforin, which induced the enhanced tumor cells apoptosis by mitochondrial apoptotic pathway, and ultimately led to enhanced tumor cell growth inhibition. ST-4 may be a promising candidate for antitumor clinic usage in future. - Highlights: • We obtained a SEC2 mutant ST-4 with enhanced superantigen and antitumor activity. • Increased ST-4-TCR recognition contributed to massive T cells and NK cells activation. • Up-regulated GzmB and PRF1 in T cell by ST-4 induced enhanced tumor cells apoptosis. • Enhanced tumor cell apoptosis

Claudin-4 Overexpression in Epithelial Ovarian Cancer Is Associated with Hypomethylation and Is a Potential Target for Modulation of Tight Junction Barrier Function Using a C-Terminal Fragment of Clostridium perfringens Enterotoxin

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Babak Litkouhi

2007-04-01

Full Text Available BACKGROUND: Claudin-4, a tight junction (TJ protein and receptor for the C-terminal fragment of Clostridium perfringens enterotoxin (C-CPE, is overexpressed in epithelial ovarian cancer (EOC. Previous research suggests DNA methylation is a mechanism for claudin-4 overexpression in cancer and that C-CPE acts as an absorption-enhancing agent in claudin-4expressing cells. We sought to correlate claudin-4 overexpression in EOC with clinical outcomes and TJ barrier function, investigate DNA methylation as a mechanism for overexpression, and evaluate the effect of C-CPE on the TJ. METHODS: Claudin-4 expression in EOC was quantified and correlated with clinical outcomes. Claudin-4 methylation status was determined, and claudin-4-negative cell lines were treated with a demethylating agent. Electric cell-substrate impedance sensing was used to calculate junctional (paracellular resistance (Rb in EOC cells after claudin-4 silencing and after C-CPE treatment. RESULTS: Claudin4 overexpression in EOC does not correlate with survival or other clinical endpoints and is associated with hypomethylation. Claudin-4 overexpression correlates with Rb and C-CPE treatment of EOC cells significantly decreased Rb in a dose- and claudin-4-dependent noncytotoxic manner. CONCLUSIONS: C-CPE treatment of EOC cells leads to altered TJ function. Further research is needed to determine the potential clinical applications of C-CPE in EOC drug delivery strategies.

Molecular Typing of Staphylococcus Aureus Isolate Responsible for Staphylococcal Poisoning Incident in Homemade Food.

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Macori, Guerrino; Bellio, Alberto; Bianchi, Daniela Manila; Gallina, Silvia; Adriano, Daniela; Zuccon, Fabio; Chiesa, Francesco; Acutis, Pier Luigi; Casalinuovo, Francesco; Decastelli, Lucia

2016-04-19

In October 2012, two persons fell ill with symptoms consistent with staphylococcal food poisoning after eating home-canned tuna fish and tomatoes. Laboratory investigation detected the enterotoxins in the home-canned tuna and molecular analysis of the isolated Staphylococcus aureus confirmed it carried toxin genes. Qualitative enzyme-linked immunosorbent assay and enzime linked fluorescent assay methods and quantitative assay identified the enterotoxins in the food leftovers, specifically staphylococcal enterotoxins type A (SEA) and D (SED), respectively 0.49 and 2.04 ng/g. The laboratory results are discussed considering the relation to the fish in oil, survival and heat resistance of S. aureus , and presumptive microbial contamination due to improper handling during home-canning procedures. This is the first reported cluster of foodborne illnesses due to staphylococcal enterotoxins in tuna in Italy. In this study, we reported cases described and analysed for their spa -type. Showing a high heterogeneity of isolates, spa -type t 13252 is correlated in a node of the minimum spanning tree and it has never been reported as responsible for foodborne outbreak. This case underlines the importance of risk communication and dissemination of home-canning guidelines to reduce the incidence of foodborne outbreaks caused by homemade conserves.

Molecular typing of Staphylococcus aureus isolate responsible for staphylococcal poisoning incident in homemade food

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Guerrino Macori

2016-06-01

Full Text Available In October 2012, two persons fell ill with symptoms consistent with staphylococcal food poisoning after eating home-canned tuna fish and tomatoes. Laboratory investigation detected the enterotoxins in the home-canned tuna and molecular analysis of the isolated Staphylococcus aureus confirmed it carried toxin genes. Qualitative enzyme-linked immunosorbent assay and enzime linked fluorescent assay methods and quantitative assay identified the enterotoxins in the food leftovers, specifically staphylococcal enterotoxins type A (SEA and D (SED, respectively 0.49 and 2.04 ng/g. The laboratory results are discussed considering the relation to the fish in oil, survival and heat resistance of S. aureus, and presumptive microbial contamination due to improper handling during home-canning procedures. This is the first reported cluster of foodborne illnesses due to staphylococcal enterotoxins in tuna in Italy. In this study, we reported cases described and analysed for their spa-type. Showing a high heterogeneity of isolates, spa-type t13252 is correlated in a node of the minimum spanning tree and it has never been reported as responsible for foodborne outbreak. This case underlines the importance of risk communication and dissemination of home-canning guidelines to reduce the incidence of foodborne outbreaks caused by homemade conserves.

Possible Application of Biotechnology to the Development of Biological Agents by Potential Enemies

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1987-06-01

7 beta chains Shigella dysenteriae type 1 plus other Inhibits 60S ribosome in eukaryotes -22- E. coli enterotoxin (heat labile) 2 subunits...toxin and those from E. colt because they do not act directly on intestinal cells. The same clinical response is obtained in monkeys following...there arc few good models in which to stud> the toxicity of the enterotoxins. Monkeys show many of the symptoms of toxic shock syndrome, however

A Study To Assess the Numbers and Prevalence of Bacillus cereus and Its Toxins in Pasteurized Fluid Milk.

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Saleh-Lakha, Saleema; Leon-Velarde, Carlos G; Chen, Shu; Lee, Susan; Shannon, Kelly; Fabri, Martha; Downing, Gavin; Keown, Bruce

2017-07-01

Bacillus cereus is a pathogenic adulterant of raw milk and can persist as spores and grow in pasteurized milk. The objective of this study was to determine the prevalence of B. cereus and its enterotoxins in pasteurized milk at its best-before date when stored at 4, 7, and 10°C. More than 5.5% of moderately temperature-abused products (stored at 7°C) were found to contain >10 5 CFU/mL B. cereus , and about 4% of them contained enterotoxins at a level that may result in foodborne illness; in addition, more than 31% of the products contained >10 5 CFU/mL B. cereus and associated enterotoxins when stored at 10°C. Results from a growth kinetic study demonstrated that enterotoxin production by B. cereus in pasteurized milk can occur in as short as 7 to 8 days of storage at 7°C. The higher B. cereus counts were associated with products containing higher butterfat content or with those produced using the conventional high-temperature, short-time pasteurization process. Traditional indicators, aerobic colony counts and psychrotrophic counts, were found to have no correlation with level of B. cereus in milk. The characterization of 17 representative B. cereus isolates from pasteurized milk revealed five toxigenic gene patterns, with all the strains carrying genes encoding for diarrheal toxins but not for an emetic toxin, and with one strain containing all four diarrheal enterotoxin genes (nheA, entFM, hblC, and cytK). The results of this study demonstrate the risks associated even with moderately temperature-abused pasteurized milk and the necessity of a controlled cold chain throughout the shelf life of fluid milk to enhance product safety and minimize foodborne illness.

Prevalence of toxin genes among the clinical isolates of Staphylococcus aureus and its clinical impact

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Divya Deodhar

2015-01-01

Full Text Available Introduction: Staphylococcus aureus (S. aureus causes a variety of infections, ranging from a mild skin infection to blood stream infections and deep seated infections. As Stapylococcus aureus bacteremia (SAB has the tendency to cause endovascular and metastatic infections, complications can occur at almost all sites of the body. Hence, SAB is associated with increased morbidity and mortality in spite of appropriate antimicrobial treatment. The virulence in S. aureus is determined by the presence of adhesins and toxins, which behave like superantigens (SAgs and leads to a massive release of proinflammatory cytokines causing overwhelming inflammatory response leading to endothelial leakage, hemodynamic shock, multiorgan failure, and possibly death. Materials and Methods: One year prospective study conducted in a tertiary care hospital in southern part of India included all patients with SAB. Clinical details were filled according to. All isolates were subjected to polymerase chain reaction (PCR for enterotoxin profiling. Results: A total of 101 patients of SAB were identified which comprises of 61 (60.4% patients with methicillin-susceptible S. aureus (MSSA and 40 (39.6% patients with methicillin-resistant S. aureus (MRSA. Most common predictors of mortality were prior hospitalization and antibiotic intake, severe organ dysfunction, shock, tachycardia, and leukocytosis. Two-third of the isolates had at least one enterotoxin, most prevalent was sea; 28% and 27% (P - value = 0.001 MSSA isolates had seg and sei; whereas, 38.6% (P - value < 0.001 of MRSA isolates were found to have sea. The most common enterotoxin associated with mortality was sei, which comprised of 38% of all mortality. Conclusion: In SAB, the significant predictors of mortality were prior hospitalization and antibiotic intake, presence of multiorgan dysfunction, and shock. Although overall significance between the enterotoxin and shock could not be demonstrated, it successfully

Prevalence of Toxin Genes among the Clinical Isolates of Staphylococcus aureus and its Clinical Impact.

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Deodhar, Divya; Varghese, George; Balaji, Veeraraghavan; John, James; Rebekah, Grace; Janardhanan, Jeshina; Jeyaraman, Ranjith; Jasmine, Sudha; Mathews, Prasad

2015-01-01

Staphylococcus aureus (S. aureus) causes a variety of infections, ranging from a mild skin infection to blood stream infections and deep seated infections. As Stapylococcus aureus bacteremia (SAB) has the tendency to cause endovascular and metastatic infections, complications can occur at almost all sites of the body. Hence, SAB is associated with increased morbidity and mortality in spite of appropriate antimicrobial treatment. The virulence in S. aureus is determined by the presence of adhesins and toxins, which behave like superantigens (SAgs) and leads to a massive release of proinflammatory cytokines causing overwhelming inflammatory response leading to endothelial leakage, hemodynamic shock, multiorgan failure, and possibly death. One year prospective study conducted in a tertiary care hospital in southern part of India included all patients with SAB. Clinical details were filled according to. All isolates were subjected to polymerase chain reaction (PCR) for enterotoxin profiling. A total of 101 patients of SAB were identified which comprises of 61 (60.4%) patients with methicillin-susceptible S. aureus (MSSA) and 40 (39.6%) patients with methicillin-resistant S. aureus (MRSA). Most common predictors of mortality were prior hospitalization and antibiotic intake, severe organ dysfunction, shock, tachycardia, and leukocytosis. Two-third of the isolates had at least one enterotoxin, most prevalent was sea; 28% and 27% (P - value = 0.001) MSSA isolates had seg and sei; whereas, 38.6% (P - value < 0.001) of MRSA isolates were found to have sea. The most common enterotoxin associated with mortality was sei, which comprised of 38% of all mortality. In SAB, the significant predictors of mortality were prior hospitalization and antibiotic intake, presence of multiorgan dysfunction, and shock. Although overall significance between the enterotoxin and shock could not be demonstrated, it successfully demonstrated the difference of enterotoxin between MSSA and MRSA.

Natural indoles, indole-3-carbinol and 3,3′-diindolymethane, inhibit T cell activation by staphylococcal enterotoxin B through epigenetic regulation involving HDAC expression

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Busbee, Philip B.; Nagarkatti, Mitzi; Nagarkatti, Prakash S.

2014-01-01

Staphylococcal enterotoxin B (SEB) is a potent exotoxin produced by the Staphylococcus aureus. This toxin is classified as a superantigen because of its ability to directly bind with MHC-II class molecules followed by activation of a large proportion of T cells bearing specific Vβ-T cell receptors. Commonly associated with classic food poisoning, SEB has also been shown to induce toxic shock syndrome, and is also considered to be a potential biological warfare agent because it is easily aerosolized. In the present study, we assessed the ability of indole-3-carbinol (I3C) and one of its byproducts, 3,3′-diindolylmethane (DIM), found in cruciferous vegetables, to counteract the effects of SEB-induced activation of T cells in mice. Both I3C and DIM were found to decrease the activation, proliferation, and cytokine production by SEB-activated Vβ8 + T cells in vitro and in vivo. Interestingly, inhibitors of histone deacetylase class I (HDAC-I), but not class II (HDAC-II), showed significant decrease in SEB-induced T cell activation and cytokine production, thereby suggesting that epigenetic modulation plays a critical role in the regulation of SEB-induced inflammation. In addition, I3C and DIM caused a decrease in HDAC-I but not HDAC-II in SEB-activated T cells, thereby suggesting that I3C and DIM may inhibit SEB-mediated T cell activation by acting as HDAC-I inhibitors. These studies not only suggest for the first time that plant-derived indoles are potent suppressors of SEB-induced T cell activation and cytokine storm but also that they may mediate these effects by acting as HDAC inhibitors. - Highlights: • I3C and DIM reduce SEB-induced T cell activation and inflammatory cytokines. • Inhibiting class I HDACs reduces T cell activation and inflammatory cytokines. • Inhibiting class II HDACs increases T cell activation and inflammatory cytokines. • I3C and DIM selectively reduce mRNA expression of class I HDACs. • Novel use and mechanism to counteract SEB

Occurrence of Natural Bacillus thuringiensis Contaminants and Residues of Bacillus thuringiensis-Based Insecticides on Fresh Fruits and Vegetables

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Frederiksen, Kristine; Rosenquist, Hanne; Jørgensen, Kirsten; Wilcks, Andrea

2006-01-01

A total of 128 Bacillus cereus-like strains isolated from fresh fruits and vegetables for sale in retail shops in Denmark were characterized. Of these strains, 39% (50/128) were classified as Bacillus thuringiensis on the basis of their content of cry genes determined by PCR or crystal proteins visualized by microscopy. Random amplified polymorphic DNA analysis and plasmid profiling indicated that 23 of the 50 B. thuringiensis strains were of the same subtype as B. thuringiensis strains used as commercial bioinsecticides. Fourteen isolates were indistinguishable from B. thuringiensis subsp. kurstaki HD1 present in the products Dipel, Biobit, and Foray, and nine isolates grouped with B. thuringiensis subsp. aizawai present in Turex. The commercial strains were primarily isolated from samples of tomatoes, cucumbers, and peppers. A multiplex PCR method was developed to simultaneously detect all three genes in the enterotoxin hemolysin BL (HBL) and the nonhemolytic enterotoxin (NHE), respectively. This revealed that the frequency of these enterotoxin genes was higher among the strains indistinguishable from the commercial strains than among the other B. thuringiensis and B. cereus-like strains isolated from fruits and vegetables. The same was seen for a third enterotoxin, CytK. In conclusion, the present study strongly indicates that residues of B. thuringiensis-based insecticides can be found on fresh fruits and vegetables and that these are potentially enterotoxigenic. PMID:16672488

Incorporation of membrane-anchored flagellin or Escherichia coli heat-labile enterotoxin B subunit enhances the immunogenicity of rabies virus-like particles in mice and dogs

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Yinglin eQi

2015-03-01

Full Text Available Rabies remains an important worldwide public health threat, so safe, effective and affordable vaccines are still being sought. Virus-like particle (VLP-based vaccines targeting various viral pathogens have been successfully produced, licensed and commercialized. Here, we designed and constructed two chimeric rabies virus-like particles (cRVLPs containing rabies virus (RABV glycoprotein (G, matrix (M protein, and membrane-anchored flagellin (EVLP-F or Escherichia coli heat-labile enterotoxin B subunit (EVLP-L as molecular adjuvants to enhance the immune response against rabies. The immunogenicity and potential of cRVLPs as novel rabies vaccine were evaluated by intramuscular vaccination in mouse and dog models. Mouse studies demonstrated that both EVLP-F and EVLP-L induced faster and larger virus-neutralizing antibodies (VNA responses and elicited greater numbers of CD4+ and CD8+ T cells secreting IFN-γ or IL-4 compared with a standard rabies VLP (sRVLP containing only G and M. Moreover, cRVLPs recruited and/or activated more B cells and dendritic cells in inguinal lymph nodes. EVLP-F induced a strong, specific IgG2a response but not an IgG1 response, suggesting the activation of Th1 class immunity; in contrast, Th2 class immunity was observed with EVLP-L. The significantly enhanced humoral and cellular immune responses induced by cRVLPs provided complete protection against lethal challenge with RABV. Most importantly, dogs vaccinated with EVLP-F or EVLP-L exhibited increased VNA titers in sera and enhanced IFN-γ and IL-4 secretion from peripheral blood mononuclear cells. Taken together, these results illustrate that when incorporated into sRVLP, membrane-anchored flagellin and LTB possess strong adjuvant activity. EVLP-F and EVLP-L induce significantly enhanced RABV-specific humoral and cellular immune responses in both mouse and dog. Therefore, these cRVLPs may be developed as safe and more efficacious rabies vaccine candidate for animals.

Isolation of Bacillus cereus Group from the Fecal Material of Endangered Wood Turtles.

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Nfor, Nancy Ngvumbo; Lapin, Carly N; McLaughlin, Richard William

2015-10-01

Members of the Bacillus cereus group are opportunistic human pathogens. They can be found in a broad range of foods. Diarrheal food poisoning and/or emetic type syndromes can result from eating contaminated food. In this study, seven B. cereus group members were isolated from the fecal material of Wood Turtles (Glyptemys insculpta). The isolates were then assessed for the presence of enterotoxin genes (nheA, entFM, hblC, and cytK) using PCR. The most prevalent is the nonhemolytic enterotoxin gene which was found in all seven isolates.

Staphylococcal Entertotoxins of the Enterotoxin Gene Cluster (egcSEs Induce Nitrous Oxide- and Cytokine Dependent Tumor Cell Apoptosis in a Broad Panel of Human Tumor Cells

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David eTerman

2013-08-01

Full Text Available The egcSEs comprise five genetically linked staphylococcal enterotoxins, SEG, SEI, SElM, SElN and SElO and two pseudotoxins which constitute an operon present in up to 80% of Staphylococcus aureus isolates. A preparation containing theses proteins was recently used to treat advanced lung cancer with pleural effusion. We investigated the hypothesis that egcSEs induce nitrous oxide (NO and associated cytokine production and that these agents may be involved in tumoricidal effects against a broad panel of clinically relevant human tumor cells. Preliminary studies showed that egcSEs and SEA activated T cells (range: 11-25% in a concentration dependent manner. Peripheral blood mononuclear cells (PBMCs stimulated with equimolar quantities of egcSEs expressed NO synthase and generated robust levels of nitrite (range: 200-250 µM, a breakdown product of NO; this reaction was inhibited by NG-monomethyl-L-arginine (L-NMMA (0.3 mM, an NO synthase antagonist. Cell free supernatants (CSFs of all egcSE-stimulated PBMCs were also equally effective in inducing concentration dependent tumor cell apoptosis in a broad panel of human tumor cells. The latter effect was due in part to the generation of NO and TNF-α since it was significantly abolished by L-NMMA, anti-TNF-α antibodies respectively and a combination thereof. A hierarchy of tumor cell sensitivity to these CFSs was as follows: lung carcinoma>osteogenic sarcoma>melanoma>breast carcinoma>neuroblastoma. Notably, SEG induced robust activation of NO/TNFα-dependent tumor cell apoptosis comparable to the other egcSEs and SEA despite TNF-α and IFN-γ levels that were 2 and 8 fold lower respectively than the other egcSEs and SEA. Thus, egcSEs produced by S. aureus induce NO synthase and the increased NO formation together with TNF-α appear to contribute to egcSE-mediated apoptosis against a broad panel of human tumor cells.

Methicillin-resistant Staphylococcus aureus: a controversial food-borne pathogen.

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Sergelidis, D; Angelidis, A S

2017-06-01

Methicillin-resistant Staphylococcus aureus (MRSA) is a major cause of severe healthcare-associated (HA) infections. Although during the last decade the incidence of HA invasive infections has dropped, the incidence of community-associated MRSA (CA-MRSA) infections has risen among the general population. Moreover, CA-MRSA, livestock-associated MRSA (LA-MRSA) and HA-MRSA (HA-MRSA) can be found in foods intended for human consumption. Several studies from different geographical areas have reported the presence of enterotoxin genes in several MRSA food isolates. Molecular typing studies have revealed genetic relatedness of these enterotoxigenic isolates with isolates incriminated in human infections. The contamination sources for foods, especially animal-origin foods, may be livestock as well as humans involved in animal husbandry and food-processing. Under favourable environmental conditions for growth and enterotoxin production, enterotoxigenic S. aureus isolates present in foods can cause staphylococcal food poisoning (SFP), irrespective of the contamination origin. Owing to the typically moderate clinical manifestations of SFP, the S. aureus strains responsible for SFP (cases or outbreaks) are frequently either not identified or not further characterized. Antimicrobial susceptibility testing is rarely performed, because administration of antimicrobial therapy is not required in the vast majority of cases. Staphylococcal food poisoning is the result of consumption of foods with preformed enterotoxins. Hence, similar to methicillin-sensitive enterotoxigenic S. aureus, enterotoxigenic MRSA can also act as food-borne pathogens upon favourable conditions for growth and enterotoxin production. The severity of the intoxication is not related to the antimicrobial resistance profile of the causative S. aureus strain and therefore MRSA food-borne outbreaks are not expected to be more severe. This review evaluates the potential of methicillin-resistant Staphylococcus

Natural indoles, indole-3-carbinol and 3,3′-diindolymethane, inhibit T cell activation by staphylococcal enterotoxin B through epigenetic regulation involving HDAC expression

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Busbee, Philip B.; Nagarkatti, Mitzi; Nagarkatti, Prakash S., E-mail: prakash@mailbox.sc.edu

2014-01-01

Staphylococcal enterotoxin B (SEB) is a potent exotoxin produced by the Staphylococcus aureus. This toxin is classified as a superantigen because of its ability to directly bind with MHC-II class molecules followed by activation of a large proportion of T cells bearing specific Vβ-T cell receptors. Commonly associated with classic food poisoning, SEB has also been shown to induce toxic shock syndrome, and is also considered to be a potential biological warfare agent because it is easily aerosolized. In the present study, we assessed the ability of indole-3-carbinol (I3C) and one of its byproducts, 3,3′-diindolylmethane (DIM), found in cruciferous vegetables, to counteract the effects of SEB-induced activation of T cells in mice. Both I3C and DIM were found to decrease the activation, proliferation, and cytokine production by SEB-activated Vβ8{sup +} T cells in vitro and in vivo. Interestingly, inhibitors of histone deacetylase class I (HDAC-I), but not class II (HDAC-II), showed significant decrease in SEB-induced T cell activation and cytokine production, thereby suggesting that epigenetic modulation plays a critical role in the regulation of SEB-induced inflammation. In addition, I3C and DIM caused a decrease in HDAC-I but not HDAC-II in SEB-activated T cells, thereby suggesting that I3C and DIM may inhibit SEB-mediated T cell activation by acting as HDAC-I inhibitors. These studies not only suggest for the first time that plant-derived indoles are potent suppressors of SEB-induced T cell activation and cytokine storm but also that they may mediate these effects by acting as HDAC inhibitors. - Highlights: • I3C and DIM reduce SEB-induced T cell activation and inflammatory cytokines. • Inhibiting class I HDACs reduces T cell activation and inflammatory cytokines. • Inhibiting class II HDACs increases T cell activation and inflammatory cytokines. • I3C and DIM selectively reduce mRNA expression of class I HDACs. • Novel use and mechanism to counteract

Influence of Host Interleukin-10 Polymorphisms on Development of Traveler's Diarrhea Due to Heat-Labile Enterotoxin-Producing Escherichia coli in Travelers from the United States Who Are Visiting Mexico▿

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Flores, Jose; DuPont, Herbert L.; Lee, Stephanie A.; Belkind-Gerson, Jaime; Paredes, Mercedes; Mohamed, Jamal A.; Armitige, Lisa Y.; Guo, Dong-Chuan; Okhuysen, Pablo C.

2008-01-01

Up to 60% of U.S. visitors to Mexico develop traveler's diarrhea (TD), mostly due to enterotoxigenic Escherichia coli (ETEC) strains that produce heat-labile (LT) and/or heat-stable (ST) enterotoxins. Distinct single-nucleotide polymorphisms (SNPs) within the interleukin-10 (IL-10) promoter have been associated with high, intermediate, or low production of IL-10. We conducted a prospective study to investigate the association of SNPs in the IL-10 promoter and the occurrence of TD in ETEC LT-exposed travelers. Sera from U.S. travelers to Mexico collected on arrival and departure were studied for ETEC LT seroconversion by using cholera toxin as the antigen. Pyrosequencing was performed to genotype IL-10 SNPs. Stools from subjects who developed diarrhea were also studied for other enteropathogens. One hundred twenty-one of 569 (21.3%) travelers seroconverted to ETEC LT, and among them 75 (62%) developed diarrhea. Symptomatic seroconversion was more commonly seen in subjects who carried a genotype producing high levels of IL-10; it was seen in 83% of subjects with the GG genotype versus 54% of subjects with the AA genotype at IL-10 gene position −1082 (P, 0.02), in 71% of those with the CC genotype versus 33% of those with the TT genotype at position −819 (P, 0.005), and in 71% of those with the CC genotype versus 38% of those with the AA genotype at position −592 (P, 0.02). Travelers with the GCC haplotype were more likely to have symptomatic seroconversion than those with the ATA haplotype (71% versus 38%; P, 0.002). Travelers genetically predisposed to produce high levels of IL-10 were more likely to experience symptomatic ETEC TD. PMID:18579697

Influence of host interleukin-10 polymorphisms on development of traveler's diarrhea due to heat-labile enterotoxin-producing Escherichia coli in travelers from the United States who are visiting Mexico.

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Flores, Jose; DuPont, Herbert L; Lee, Stephanie A; Belkind-Gerson, Jaime; Paredes, Mercedes; Mohamed, Jamal A; Armitige, Lisa Y; Guo, Dong-Chuan; Okhuysen, Pablo C

2008-08-01

Up to 60% of U.S. visitors to Mexico develop traveler's diarrhea (TD), mostly due to enterotoxigenic Escherichia coli (ETEC) strains that produce heat-labile (LT) and/or heat-stable (ST) enterotoxins. Distinct single-nucleotide polymorphisms (SNPs) within the interleukin-10 (IL-10) promoter have been associated with high, intermediate, or low production of IL-10. We conducted a prospective study to investigate the association of SNPs in the IL-10 promoter and the occurrence of TD in ETEC LT-exposed travelers. Sera from U.S. travelers to Mexico collected on arrival and departure were studied for ETEC LT seroconversion by using cholera toxin as the antigen. Pyrosequencing was performed to genotype IL-10 SNPs. Stools from subjects who developed diarrhea were also studied for other enteropathogens. One hundred twenty-one of 569 (21.3%) travelers seroconverted to ETEC LT, and among them 75 (62%) developed diarrhea. Symptomatic seroconversion was more commonly seen in subjects who carried a genotype producing high levels of IL-10; it was seen in 83% of subjects with the GG genotype versus 54% of subjects with the AA genotype at IL-10 gene position -1082 (P, 0.02), in 71% of those with the CC genotype versus 33% of those with the TT genotype at position -819 (P, 0.005), and in 71% of those with the CC genotype versus 38% of those with the AA genotype at position -592 (P, 0.02). Travelers with the GCC haplotype were more likely to have symptomatic seroconversion than those with the ATA haplotype (71% versus 38%; P, 0.002). Travelers genetically predisposed to produce high levels of IL-10 were more likely to experience symptomatic ETEC TD.

Bacteroides fragilis Enterotoxin Induces Formation of Autophagosomes in Endothelial Cells but Interferes with Fusion with Lysosomes for Complete Autophagic Flux through a Mitogen-Activated Protein Kinase-, AP-1-, and C/EBP Homologous Protein-Dependent Pathway.

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Ko, Su Hyuk; Jeon, Jong Ik; Myung, Hyun Soo; Kim, Young-Jeon; Kim, Jung Mogg

2017-10-01

Bacteroides fragilis enterotoxin (BFT), a virulence factor of enterotoxigenic B. fragilis (ETBF), plays an essential role in mucosal inflammation. Although autophagy contributes to the pathogenesis of diverse infectious diseases, little is known about autophagy in ETBF infection. This study was conducted to investigate the role of BFT in the autophagic process in endothelial cells (ECs). Stimulation of human umbilical vein ECs (HUVECs) with BFT increased light chain 3 protein II (LC3-II) conversion from LC3-I and protein expression of p62, Atg5, and Atg12. In addition, BFT-exposed ECs showed increased indices of autophagosomal fusion with lysosomes such as LC3-lysosome-associated protein 2 (LAMP2) colocalization and the percentage of red vesicles monitored by the expression of dual-tagged LC3B. BFT also upregulated expression of C/EBP homologous protein (CHOP), and inhibition of CHOP significantly increased indices of autophagosomal fusion with lysosomes. BFT activated an AP-1 transcription factor, in which suppression of AP-1 activity significantly downregulated CHOP and augmented autophagosomal fusion with lysosomes. Furthermore, suppression of Jun N-terminal protein kinase (JNK) mitogen-activated protein kinase (MAPK) significantly inhibited the AP-1 and CHOP signals, leading to an increase in autophagosomal fusion with lysosomes in BFT-stimulated ECs. These results suggest that BFT induced accumulation of autophagosomes in ECs, but activation of a signaling pathway involving JNK, AP-1, and CHOP may interfere with complete autophagy. Copyright © 2017 American Society for Microbiology.

Housefly (Musca domestica as Carrier of Enterotoxigenic Staphylococcus aureus in Broiler Farms in Iran: Is it Important for Public Health?

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Mehdi Rashki Ghalehnoo

2015-08-01

Full Text Available Background: During the last decade, the prevalence of insect borne diseases due to contaminated food as well as the outbreaks of diseases due to enterotoxin-producing Staphylococcus aureus (ETSA strains has increased. Objectives: This study was conducted to determine the prevalence rate, enterotoxigenecity, and antimicrobial resistance of S. aureus isolated from M. domestica collected from the residential areas of nine districts broiler farms in Zabol, Iran. Materials and Methods: The flies were captured with a sterile nylon net and washed twice with distilled water. The contents were streaked onto selective media and S. aureus was identified using the conventional biochemical tests. Antimicrobial susceptibility testing was performed using CLSI guideline. Staphylococcal enterotoxins (SEs, SEA, SEB, SEC and SED, was detected by the reverse passive latex agglutination (PRLA method. Coagulase typing was performed with coagulase typing kit. Results: A total of 87 (17% S. aureus was isolated from 450 samples. The ability to synthesize staphylococcal enterotoxins (SEs was determined in 62 of 87 (71% isolates. SE type B was the most common enterotoxin found in the isolated S. aureus (45%, followed by SE type A (26%, SE type C (5%, SE type D (8%, SEA + SEC (7% and SEA + SEB (7%. Among the antibiotic tested, Penicillin was the most resistant antibiotic tested. Sixty seven percent of isolates belonged to coagulase type III, VI, VII, VIII, IV, and V. Conclusions: Our results suggest that S. aureus, including ETSA, is being carried by house flies such as M. domestica and may contribute to the spread of pathogenic isolates, with an impact on public health.

Enterotoxigenic coagulase positive Staphylococcus in milk and milk products, lben and jben, in northern Morocco.

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Bendahou, Abdrezzak; Abid, Mohammed; Bouteldoun, Nadine; Catelejine, Dierick; Lebbadi, Mariam

2009-04-30

The aim of this research was to determine the prevalence of enterotoxin genes (sea-seo) in Coagulase Positive Staphylococcus (CPS) isolated from unpasteurized milk and milk products. These results were compared with the results obtained by using the detection kit SET-RPLA for the specific detection of staphylococcal enterotoxins (SEA-SED). Eighty-one samples of milk and milk products were analyzed for the presence of Staphylococcus strains. Forty-six coagulase positive Staphylococcus isolates were tested for the production of staphylococcal enterotoxins (SEA-SED) by using the reversed passive latex agglutination method. The strains were also tested for the presence of se genes (sea-seo) by polymerase chain reaction. One or more classical enterotoxin products (SEA-SED) were observed in 39% of the strains tested, while se genes were detected in 56.5%. SEA and sea were most commonly detected. For newly discovered se genes among CPS isolates tested in this study, except the seh gene which was revealed in four isolates (8.7 %), none of the strains harbored any of the other se genes (see, seg, sei, sej, sek, sel, sem, seo and sen). The finding of a pathogen such as staphylococci-producing SEs and containing se genes in milk and milk products in northern Morocco may indicate a problem for public health in this region. The presence of enterotoxigenic strains in food does not always necessarily mean that the toxin will be produced. For that reason, the combination of both methods (RPLA and PCR) is a guarantee for success in diagnostic analysis tests.

The three-dimensional crystal structure of cholera toxin

Energy Technology Data Exchange (ETDEWEB)

Zhang, Rong-Guang; Westbrook, M.L.; Nance, S.; Spangler, B.D. [Argonne National Lab., IL (United States); Scott, D.L. [Yale Univ., New Haven, CT (United States). Dept. of Molecular Biophysics and Biochemistry; Westbrook, E.M. [Northwestern Univ., Evanston, IL (United States)

1996-02-01

The clinical manifestations of cholera are largely attributable to the actions of a secreted hexameric AB{sub 5} enterotoxin (choleragen). We have solved the three-dimensional structure of choleragen at 2.5 {Angstrom} resolution and compared the refined coordinates with those of choleragenoid (isolated B pentamer) and the heat-labile enterotoxin from Escherichia coli (LT). The crystalline coordinates provide a detailed view of the stereochemistry implicated in binding to GM1 gangliosides and in carrying out ADP-ribosylation. The A2 chain of choleragen, in contrast to that of LT, is a nearly continuous {alpha}-helix with an interpretable carboxyl tail.

[The staphylococcal enterotoxin burden determines the ultrastructure of ciliated epithelia and inflammatory changes in maxillary sinus mucosa of rabbits].

Science.gov (United States)

Wei, Hongqi; Zhu, Zhengwen; Cao, Zhongsheng; Liu, Zhiyong; Wu, Xiaofan; Yuan, Hui

2014-12-01

To investigate the ultrastructure of ciliated epithelia and inflammatory changes upon repeated exposure to staphylococcal enterotoxin A (SEA) of different concentrations in the maxillary sinus mucosa of rabbits. The rabbits were randomly divided into 2 groups (24 rabbits per group): low-dose SEA group and high-dose SEA group. The low-dose SEA group and high-dose SEA group received daily injections of 0.6 ng of SEA (2 ml) and 60 ng of SEA (2 ml) into the left maxillary sinus of rabbits for 28 days, respectively. Concurrent treatment of the right maxillary sinus with normal saline was used as control. Six rabbits chosen randomly in two groups were examined by computed tomography (CT) scans and then sacrificed to obtain the sinus mucosa from the two-side of maxillary sinuses for histological assessment on days 3, 7, 14 and 28. To characterize the inflammatory changes of the sinus mucosa examined using light microscope, hematoxylin and eosin (HE) and toluidine blue staining was performed. Scanning and transmission electron microscopy were performed to observe ultrastructure of ciliated epithelia in the maxillary sinus mucosa. SPSS 13.0 software was used to analyze the data. On days 14 and 28, CT images showed opacification of the left maxillary sinus in the high-dose SEA group. The percentage of epithelial disruption was (22.73 ± 5.72) % and (30.79 ± 4.30)% in the high-dose SEA group respectively, and were significantly greater than those in the low-dose SEA group (5.12% ± 1.98% and 5.38% ± 1.64%, q value was 10.079 and 19.132) and control group (4.08% ± 1.29% and 4.81% ± 1.62%, q value was 11.016 and 19.592, respectively, all P microscope, loss of cilia was observed, a few compound cilia and cytoplasmic protrusion were found, an obvious stretching of the endoplasmic reticulum and an obvious turgescence of the mitochondria was also observed. However, in the low-dose SEA group on days 14 and 28, CT scan of the left maxillary sinus showed transparency; light

Natural indoles, indole-3-carbinol (I3C and 3,3'-diindolylmethane (DIM, attenuate staphylococcal enterotoxin B-mediated liver injury by downregulating miR-31 expression and promoting caspase-2-mediated apoptosis.

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Philip B Busbee

Full Text Available Staphylococcal enterotoxin B (SEB is a potent superantigen capable of inducing inflammation characterized by robust immune cell activation and proinflammatory cytokine release. Exposure to SEB can result in food poisoning as well as fatal conditions such as toxic shock syndrome. In the current study, we investigated the effect of natural indoles including indole-3-carbinol (I3C and 3,3'-diindolylmethane (DIM on SEB-mediated liver injury. Injection of SEB into D-galactosamine-sensitized female C57BL/6 mice resulted in liver injury as indicated by an increase in enzyme aspartate transaminase (AST levels, induction of inflammatory cytokines, and massive infiltration of immune cells into the liver. Administration of I3C and DIM (40 mg/kg, by intraperitonal injection, attenuated SEB-induced acute liver injury, as evidenced by decrease in AST levels, inflammatory cytokines and cellular infiltration in the liver. I3C and DIM triggered apoptosis in SEB-activated T cells primarily through activation of the intrinsic mitochondrial pathway. In addition, inhibitor studies involving caspases revealed that I3C and DIM-mediated apoptosis in these activated cells was dependent on caspase-2 but independent of caspase-8, 9 and 3. In addition, I3C and DIM caused a decrease in Bcl-2 expression. Both compounds also down-regulated miR-31, which directly targets caspase-2 and influences apoptosis in SEB-activated cells. Our data demonstrate for the first time that indoles can effectively suppress acute hepatic inflammation caused by SEB and that this may be mediated by decreased expression of miR-31 and consequent caspase-2-dependent apoptosis in T cells.

Inhibition of Clostridium difficile toxin A and B by 1,2-cyclohexanedione modification of an arginine residue.

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Balfanz, J; Rautenberg, P

1989-12-29

Toxin A (enterotoxin) and toxin B (cytotoxin) of Clostridium difficile were both inactivated by the arginine specific reagent 1,2-cyclohexanedione. Molecular stability during the inactivation process was demonstrated by SDS-PAGE analysis showing the same migration rates for modified and unmodified forms of the 230 kDa toxin A and of the 250 kDa toxin B. Cytotoxicity of both toxins as well as mouse lethality of the enterotoxin were drastically decreased as a result of the arginine modification. The reaction followed pseudo-first-order kinetics. Analysis of the data suggested that modification of a single arginine residue was sufficient to abolish the activity of both toxins.

40 CFR 725.421 - Introduced genetic material.

Science.gov (United States)

2010-07-01

... fever toxins, pyrogenic exotoxins) Yersinia enterocolitica Heat-stable enterotoxins (ST) ... Neurotoxin Staphylococcus aureus Alpha toxin (alpha lysin) Yersinia pestis Murine toxin Snake toxins Bungarus...

Occurrence of Salmonella, Listeria monocytogenes, and enterotoxigenic Staphylococcus in goat milk from small and medium-sized farms located in Minas Gerais State, Brazil.

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Cavicchioli, V Q; Scatamburlo, T M; Yamazi, A K; Pieri, F A; Nero, L A

2015-12-01

Consumption of goat milk has been increasing due to its nutritional characteristics and health benefits. Therefore, assessment of the presence of foodborne pathogens in this product is critical to ensure its safety to consumers. The present study aimed to identify common foodborne pathogens in raw goat milk. Fifty-three samples of raw goat milk from 11 farms were collected and cultured for the presence of Salmonella spp. and Listeria monocytogenes, as well as for enumeration and isolation of coagulase-positive and coagulase-negative Staphylococcus (CPS and CNS, respectively). All samples tested negative for Salmonella spp. and L. monocytogenes. The CPS counts in raw goat milk samples were predominantly less than 2 log cfu/mL (n=39), and CNS counts were predominantly higher than 3 log cfu/mL (n=42). Based on Staphylococcus counts, 51 isolates were selected (CPS=26; CNS=25) and tested by PCR for the presence of classic enterotoxin-encoding genes (sea, seb, sec, sed, and see). Only 3 isolates (CPS=2, CNS=1) were negative for all enterotoxin-encoding genes tested, and the genotype sec and see was the most frequent (n=16), followed by sea, sec, and see (n=13) and sec (n=13); sed was not detected in any isolate. The frequencies of enterotoxin-encoding genes for CPS and CNS were similar, demonstrating the equivalence of both groups in harboring these virulent markers. These results suggest that Salmonella and L. monocytogenes are not frequent contaminants of raw goat milk, but that Staphylococcus spp. that are capable of producing enterotoxins are prevalent; therefore, consumers of raw goat milk and products made from raw milk are at risk. Copyright © 2015 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Bacillus cereus Fnr binds a [4Fe-4S] cluster and forms a ternary complex with ResD and PlcR

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Esbelin Julia

2012-06-01

Full Text Available Abstract Background Bacillus cereus is a facultative anaerobe that causes diarrheal disease in humans. Diarrheal syndrome may result from the secretion of various virulence factors including hemolysin BL and nonhemolytic enterotoxin Nhe. Expression of genes encoding Hbl and Nhe is regulated by the two redox systems, ResDE and Fnr, and the virulence regulator PlcR. B. cereus Fnr is a member of the Crp/Fnr family of iron-sulfur (Fe-S proteins. Only its apo-form has so far been studied. A major goal in deciphering the Fnr-dependent regulation of enterotoxin genes is thus to obtain and characterize holoFnr. Results Fnr has been subjected to in vitro Fe-S cluster reconstitution under anoxic conditions. UV-visible and EPR spectroscopic analyses together with the chemical estimation of the iron content indicated that Fnr binds one [4Fe-4S]2+ cluster per monomer. Atmospheric O2 causes disassembly of the Fe-S cluster, which exhibited a half-life of 15 min in air. Holo- and apoFnr have similar affinities for the nhe and hbl promoter regions, while holoFnr has a higher affinity for fnr promoter region than apoFnr. Both the apo- and holo-form of Fnr interact with ResD and PlcR to form a ternary complex. Conclusions Overall, this work shows that incorporation of the [4Fe-4S]2+ cluster is not required for DNA binding of Fnr to promoter regions of hbl and nhe enterotoxin genes or for the formation of a ternary complex with ResD and PlcR. This points to some new unusual properties of Fnr that may have physiological relevance in the redox regulation of enterotoxin gene regulation.

Prevalence of foodborne pathogens in grilled chicken from street vendors and retail outlets in Reynosa, Tamaulipas, Mexico.

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Díaz-López, A; Cantú-Ramírez, R C; Garza-González, E; Ruiz-Tolentino, L; Tellez-Luis, S J; Rivera, G; Bocanegra-García, V

2011-08-01

We analyzed a total of 70 grilled chicken samples bought randomly from street vendors and retail outlets in the city of Reynosa, Mexico, to determine the prevalence of Escherichia coli (Shiga toxin producing and enterotoxin producing), Salmonella spp., Staphylococcus aureus, Listeria spp., and Campylobacter spp. using microbiological methods and PCR detection of bacterial sequences. Of the 70 samples, 27 (38.5%) were from retail outlets and 43 (61.4%) from street vendors. All specimens were negative by both microbiological and molecular methods for Listeria monocytogenes, Shiga toxin 2 of Shiga toxin-producing E. coli, lt of enterotoxin-producing E. coli, and st enterotoxin, and all were negative for Salmonella spp. and Campylobacter jejuni by PCR. Of the samples studied, 49 (70%) had undetectable levels of the foodborne pathogens studied with the methods used. In the remaining 21 (30%) specimens, at least one pathogen was isolated or detected, with E. coli being the pathogen most frequently isolated and with two samples bearing the hlyA gene. We found no statistical difference in bacterial prevalence between retail and street vendor samples. The presence of pathogens in grilled chicken is an important public health risk because of the great demand for and daily consumption of this product in this region.

Prevalence and molecular characteristics of Staphylococcus aureus, including methicillin resistant strains, isolated from bulk can milk and raw milk products in pastoral communities of South-West Uganda.

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Asiimwe, Benon B; Baldan, Rossella; Trovato, Alberto; Cirillo, Daniela M

2017-06-13

Staphylococcus aureus strains are now regarded as zoonotic agents. In pastoral settings where human-animal interaction is intimate, multi-drug resistant microorganisms have become an emerging zoonotic issue of public health concern. The study of S. aureus prevalence, antimicrobial resistance and clonal lineages in humans, animals and food in African settings has great relevance, taking into consideration the high diversity of ethnicities, cultures and food habits that determine the lifestyle of the people. Little is known about milk carriage of methicillin resistant S. aureus strains (MRSA) and their virulence factors in Uganda. Here, we present the prevalence of MRSA in bulk can milk and raw milk products in pastoral communities of south-west Uganda. We also present PFGE profiles, spa-types, as well as frequency of enterotoxins genes. S. aureus was identified by the coagulase test, susceptibility testing by the Kirby-Bauer disc diffusion and E-test methods and MRSA by detection of the mecA gene and SCCmec types. The presence of Panton - Valentine Leucocidin (PVL) genes and staphylococcal enterotoxins was determined by PCR, while genotyping was by PFGE and spa typing. S. aureus were isolated from 30/148 (20.3%) milk and 11/91(12%) sour milk samples. mecA gene carriage, hence MRSA, was detected in 23/41 (56.1%) of the isolates, with 21 of the 23 (91.3%) being SCCmec type V; while up to 30/41 (73.2%) of the isolates were resistant to tetracycline. Only five isolates carried the PVL virulence gene, while PFGE typing revealed ten clusters (ranging from two seven isolates each) that comprised 83% of the sample, and only eight isolates with unique pulsotypes. The largest PFGE profile (E) consisted of seven isolates while t7753, t1398, and t2112 were the most common spa-types. Thirty seven of the 41 strains (90.2%) showed at least one of the eight enterotoxin genes tested, with sem 29 (70.7%), sei 25 (61%) and seg 21 (51.2%) being the most frequently observed genes. This

21 CFR 866.3700 - Staphylococcus aureus serological reagents.

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2010-04-01

... affecting the intestine) producing staphylococci from cultured isolates. The identification aids in the... while growing in meat, dairy, or bakery products. After ingestion, this enterotoxin is absorbed in the...

Marine toxins and their toxicological significance: An overview

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Sarkar, A.

, Hemolysins-1 and hemolysin-2, saxitoxin, neosaxitoxin, gonyautoxin, tetrodotoxin, ptychodiscus brevis toxin and theonellamide F. According to their mode of action, these toxins are classified into different categories such as cytotoxin, enterotoxin...

Detection of the enterotoxigenic genes (sei,sej) in Staphylococcus aureus isolates from bovine mastitis milk in the West Azerbaijan of Iran.

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Ahmady, Malahat; Kazemi, Sahar

2013-07-01

Staphylococcus aureus is a major causative pathogen of clinical and subclinical mastitis of dairy domestic ruminants. This organism produces a variety of extracellular toxins and virulence factors such as enterotoxin SEI and SEJ that contribute to its pathogenic potential. In this study 25 S. aureus isolates obtained from four dairy herds of Urmia region which is located in West Azerbaijan province in Iran. The tested isolates were identified on the basis of the cultural and biochemical properties, as well as amplification of the aroA gene which is specific for S. aureus . All isolates were also analyzed for the presence of the SEI ( sei ) and SEJ ( sej ) encoding genes using polymerase chain reaction (PCR). Seven positive isolates were detected for sei , but sej gene was not detected in any of the total number of 25 isolates. The present study revealed that the PCR amplification of the aroA gene could be used as a powerful tool for identification of S. aureus from the cases of bovine mastitis. Results of the present study also showed that the strains of S. aureus which cause mastitis can potentially produce enterotoxin SEI. Overall, our results suggest that it is of special importance to follow the presence of enterotoxin-producing S. aureus in other dairy products, especially for protecting the consumers from staphylococcal food poisoning.

Manufacturing of a novel double-function ssDNA aptamer for sensitive diagnosis and efficient neutralization of SEA.

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Sedighian, Hamid; Halabian, Raheleh; Amani, Jafar; Heiat, Mohammad; Taheri, Ramezan Ali; Imani Fooladi, Abbas Ali

2018-05-01

Staphylococcal enterotoxin A (SEA) is an enterotoxin produced mainly by Staphylococcus aureus. In recent years, it has become the most prevalent compound for staphylococcal food poisoning (SFP) around the world. In this study, we isolate new dual-function single-stranded DNA (ssDNA) aptamers by using some new methods, such as the Taguchi method, by focusing on the detection and neutralization of SEA enterotoxin in food and clinical samples. For the asymmetric polymerase chain reaction (PCR) optimization of each round of systematic evolution of ligands by exponential enrichment (SELEX), we use Taguchi L9 orthogonal arrays, and the aptamer mobility shift assay (AMSA) is used for initial evaluation of the protein-DNA interactions on the last SELEX round. In our investigation the dissociation constant (K D ) value and the limit of detection (LOD) of the candidate aptamer were found to be 8.5 ± 0.91 of nM and 5 ng/ml using surface plasmon resonance (SPR). In the current study, the Taguchi and mobility shift assay methods were innovatively harnessed to improve the selection process and evaluate the protein-aptamer interactions. To the best of our knowledge, this is the first report on employing these two methods in aptamer technology especially against bacterial toxin. Copyright © 2018 Elsevier Inc. All rights reserved.

Portación y caracterización de Staphylococcus aureus en manipuladores de alimentos Carriage and characterization of Staphylococcus aureus in food handlers

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Graciela B Jordá

2012-06-01

Full Text Available Staphylococcus aureus es una causa de intoxicaciones alimentarias por su capacidad de producir enterotoxinas. Los manipuladores de alimentos que portan S. aureus productores de enterotoxinas pueden provocar intoxicaciones alimentarias. Se estudiaron muestras tomadas de fosas nasales de 88 manipuladores de alimentos en la provincia de Misiones. El 37,5 % de los individuos analizados eran portadores de S. aureus. Mediante técnicas de amplificación (PCR, se detectaron genes que codifican la producción de enterotoxinas en 13 de los 33 aislamientos obtenidos (39,4 % y en el 14,7 % de los manipuladores. De estos aislamientos, 10 portaban el gen sea y 3 el gen sec. El estudio de sensibilidad a los antibióticos mostró un 100 % de sensibilidad a teicoplanina, gentamiclna y rifampicina; 2 aislamientos fueron resistentes a clindamicina y a eritromicina y 4 resultaron resistentes a la meticilina. Estos resultados son un alerta e indicarían la necesidad de desarrollar medidas racionales para reducir el riesgo potencial de intoxicaciones alimentarias.Staphylococcus aureus causes food poisoning due to its ability to produce enterotoxins. Food handlers carrying enterotoxin-producing S. aureus can contaminate food, thus leading to food poisoning. Samples were obtained from 88 food handlers in the Province of Misiones, Argentina. S. aureus was isolated from nasal swaps and PCR amplification was performed for genes encoding staphylococcal enterotoxins. A total of 37.5 % food handlers were positive for S. aureus. Expression of enterotoxin genes was found in 13 of the 33 (39.4 % S. aureus isolates studied, accounting for 14.7 % of food handlers. Gene sea was detected in 10 isolates followed by gene sec in 3 isolates. All isolates were susceptible to teicoplanin, gentamicin and rifampicin. Four isolates were resistant to methicillin whereas 2 isolates were resistant to clindamycin and erythromycin. These results constitute a critical alert and indicate the need

Polyclonal antibodies production against Staphylococcus aureus ...

African Journals Online (AJOL)

STORAGESEVER

2010-02-01

Feb 1, 2010 ... ... and the enterotoxins staphylococcies dried dosage remains the unique means to define the food innocuousness. The standard methods of cultivation used to detect .... A second sample of pasteurized and packaged milk.

Adaptation in Bacillus cereus: from stress to disease

Directory of Open Access Journals (Sweden)

Catherine Duport

2016-10-01

Full Text Available Bacillus cereus is a foodborne pathogen that causes diarrheal disease in humans. After ingestion B. cereus experiences in the human gastro-intestinal tract abiotic physical variables encountered in food, such as acidic pH in the stomach and changing oxygen conditions in the human intestine. B. cereus responds to environmental changing conditions (stress by reversibly adjusting its physiology to maximize resource utilization while maintaining structural and genetic integrity by repairing and minimizing damage to cellular infrastructure. As reviewed in this article, B. cereus adapts to acidic pH and changing oxygen conditions through diverse regulatory mechanisms and then exploits its metabolic flexibility to grow and produce enterotoxins. We then focus on the intricate link between metabolism, redox homeostasis and enterotoxins, which are recognized as important contributors of food-borne disease.

Enhancement of Intranasal Vaccination in Mice with Deglycosylated Chain A Ricin by LTR72, a Novel Mucosal Adjuvant

National Research Council Canada - National Science Library

Kende, Meir; Del Giudice, Giuseppe; Rivera, Noelia; Hewetson, John

2006-01-01

.... However, in the presence of 4, 2, or 1 microg of the mucosal adjuvant LTR72, a mutant of the heat-labile enterotoxin of Escherichia coli, the low antibody response and protection were substantially enhanced...

Real-Time Fluorogenic Polymerase Chain Reaction (PCR) Assays to Nucleic Acid-Based Detection of Simulants and Biothreat Agents

National Research Council Canada - National Science Library

Khan, Akbar S; O'Connell, Kevin P; Bucher, Jennifer R; Anderson, Patricia E; Cao, Cheng J; Gostomski, Mark V; Valdes, James J

2003-01-01

.... We describe here assays for the detection of the gene encoding staphylococcal enterotoxin A (SEA), a toxin produced by the bacterium Staphylococcus aureus and an agent responsible for a significant fraction of food poisoning incidents worldwide...

Antidiarrhoeal activity of different plants used in traditional medicine

African Journals Online (AJOL)

SERVER

2007-12-28

Dec 28, 2007 ... components such as artificial sweeteners and lactose and the result is the .... decrease in severe diarrhoea of rats treated with castor oil. This extract also ..... heat-labile enterotoxin and ganglioside interaction. J. Ethnopharma-.

Enhancement of Intranasal Vaccination in Mice with Deglycosylated Chain A Ricin by LTR72, a Novel Mucosal Adjuvant

National Research Council Canada - National Science Library

Kende, Meir; Del Giudice, Giuseppe; Rivera, Noelia; Hewetson, John

2006-01-01

.... However, in the presence of 4, 2, or 1 micro-gram of the mucosal adjuvant LTR72, a mutant of the heat-labile enterotoxin of Escherichia coli, the low antibody response and protection were substantially enhanced...

Aeromonas caviae strain induces Th1 cytokine response in mouse intestinal tract

Science.gov (United States)

Aeromonas caviae has been associated with human gastrointestinal disease. Strains of this species typically lack virulence factors (VFs) such as enterotoxins and hemolysins that are produced by other human pathogens of the Aeromonas genus,. Microarray profiling of...

PCR detection and serotyping of enterotoxigenic and shigatoxigenic Escherichia coli isolates obtained from chicken meat in Mumbai, India

Directory of Open Access Journals (Sweden)

R. J. Zende,

2013-08-01

Full Text Available Aim: Present study was undertaken to find out the frequency of few virulent genes and prevalence of related strains of Escherichia coli isolated from chicken meat obtained from chicken retail shops by Polymerase Chain Reaction (PCR.Materials and Methods: 66 samples of freshly slaughtered chicken meat were collected from 22 identified retail shops located at Mumbai city, randomly. Processed meat samples were cultured in EMB agar and presumptive colonies were confirmed by various biochemical tests. PCR method was accustomed for identification of the genes coding for heat-stable enterotoxin a (STa, heat labile enterotoxin (LT, shiga-like toxins 1 and 2 (SLT1 and SLT2. E. coli isolates were sent to National Salmonella and Escherichia Centre, CRI, Kasauli, HP, India for serotyping.Results: 11 (16.67% E. coli strains were isolated from 66 chicken meat samples. 3 (27.27% out of 11 harbored the gene for SLT2, and 2 (18.18% for STa. None of the strain contains SLT1 and LT genes. Serotypes detected were rough, O2, O20, O22, O102 each for one isolate and 6 isolates were untypable (UT.Conclusion: The results concluded that chicken meat samples analysed harbored genes for shiga like toxins and enterotoxins and different serotypes of E. coli. These findings indicating that regular monitoring of chicken meat is essential for this pathogen to prevent potential public health problems.

Virulence Factors of Staphylococcus aureus Isolates in an Iranian Referral Children's Hospital.

Science.gov (United States)

Sabouni, Farah; Mahmoudi, Shima; Bahador, Abbas; Pourakbari, Babak; Sadeghi, Reihaneh Hosseinpour; Ashtiani, Mohammad Taghi Haghi; Nikmanesh, Bahram; Mamishi, Setareh

2014-04-01

The clinical importance of Staphylococcus aureus (S. aureus) is attributed to notable virulence factors, surface proteins, toxins, and enzymes as well as the rapid development of drug resistance. The aim of this study was to compare the occurrence of virulence factors produced by S. aureus strains isolated from children in an Iranian referral children's hospital. The presence of genes encoding for the enterotoxins A (sea), B (seb), C (sec), D (sed), TSST-1 (tsst), exfoliative toxin A (eta), and exfoliative toxin B (etb) were detected by Multiplex polymerase chain reaction (PCR) using specific primers. In addition, the standardized Kirby-Bauer disc-diffusion method was performed on Mueller-Hinton agar. In total, 133 S. aureus isolates were obtained from different patients. Of these S. aureus isolates, 64 (48%) were methicillin-resistant S. aureus (MRSA), and all of these tested positive for the mecA gene. Regarding the classical enterotoxin genes, sea gene (40.6%) was the most prevalent followed by seb (19.6%), tsst (12.8%), eta (11.3%), etb (9%), sed (4.5%), and sec (3%). Among methicillin-susceptible S. aureus (MSSA) isolates, seb and tsst were the more prevalent toxins in comparison with MRSA isolates (p  0.05). In our study enterotoxin A was produced by 40.6% of the isolates (48% from MRSA and 33% from MSSA isolates) which was higher than in previous reports. According to our results, strict hygiene and preventative measures during food processing are highly recommended.

Superantigen and HLA-DR ligation induce phospholipase-C gamma 1 activation in class II+ T cells

DEFF Research Database (Denmark)

Kanner, S B; Odum, Niels; Grosmaire, L

1992-01-01

Bacterial enterotoxin superantigens bind directly to HLA class II molecules (HLA-DR) expressed on both APC and activated human T cells, and simultaneously bind to certain V beta chains of the TCR. In this report, we compared early T cell signaling events in human alloantigen-stimulated T cells when...... activated by HLA-DR ligation through antibody cross-linking or by direct enterotoxin superantigen binding. Both types of stimuli induced tyrosine phosphorylation of phosphatidylinositol-specific phospholipase C gamma 1 (PLC gamma 1) and an increase in intracellular calcium concentration; however......, superantigen-induced signaling was stronger than class II ligation alone. Antibody-mediated ligation of HLA-DR with CD3 resulted in augmented PLC gamma 1 activation and increased calcium mobilization, consistent with a mechanism of superantigen activity through a combination of class II and CD3/Ti signals...

Incidence of enterotoxigenic staphylococci and their toxins in foods.

Science.gov (United States)

Soriano, J M; Font, G; Rico, H; Moltó, J C; Mañes, J

2002-05-01

Of 504 food samples collected from cafeterias, 19 (3.8%) yielded strains of enterotoxigenic staphylococci, and 10 (52.6%), 4 (21.1%), 3 (15.8%), and 2 (10.5%) of these strains produced enterotoxins C (SEC), D (SED), B (SEB), and A (SEA), respectively. Moreover, SEA, SEB, and SEC were isolated from three hamburger samples. Of 181 food samples collected from four restaurants before the implementation of the hazard analysis and critical control point (HACCP) system, 7 (3.9%) were found to contain enterotoxigenic strains, and SED, SEC, and SEA were produced by 4 (57.1%), 2 (28.6%), and 1 (14.3%) of these strains, respectively. One meatball sample with SEC was detected in a restaurant. After the implementation of the HACCP system in four restaurants, neither enterotoxigenic staphylococci nor enterotoxins were detected in 196 studied samples.

Geographical variation in the presence of genes encoding superantigenic exotoxins and beta-hemolysin among Staphylococcus aureus isolated from bovine mastitis in Europe and USA

DEFF Research Database (Denmark)

Larsen, H. D.; Aarestrup, Frank Møller; Jensen, N. E.

2002-01-01

The object was to examine the geographical variation in the presence of superantigenic exotoxins and beta-hemolysin among epidemiologically independent Staphyirrcoccus aureus isolates from bovine mastitis. A total of 462 S. aureus isolates from nine European countries and USA were examined...... for the individual exotoxins. The genes encoding enterotoxin C, TSST-1, and enterotoxin D were the most common superantigens. The present and earlier studies demonstrate that the superantigenic exotoxins that were investigated in this study, do not play a role in the pathogenesis of bovine S. aureus mastitis...... regions in the beta-hemolysin encoding gene of the Norwegian isolates is suggested, and should be investigated further. The consistent presence of beta-hemolysin suggests that this factor, or a co-existing gene correlated to beta-hemolysin, may be an active virulence factor in the pathogenesis of bovine S...

Selection of bacteriocin producer strains of lactic acid bacteria from a dairy environment.

Science.gov (United States)

Lasagno, M; Beoleito, V; Sesma, F; Raya, R; Font de Valdez, G; Eraso, A

2002-01-01

Two strains showing bacteriocin production were selected from a total of 206 lactic acid bacteria isolated from samples of milk, milk serum, whey and homemade cheeses in Southern Cordoba, Argentina. This property was detected by means of well diffusion assays. The strains were identified as Enterococcus hirae and Enterococcus durans. The protein nature of those substances was proved by showing their sensitivity to type IV and XXV proteases, papaine, trypsin, pepsin and K proteinase. The bacteriocins inhibited the growth of Listeria monocytogenes, Bacillus cereus, Clostridium perfringes and two strains of Staphylococcus aureus, an A-enterotoxin and a B-enterotoxin producers. All of these bacteria are common pathogens usually associated with food borne diseases (ETA). These lactic acid bacteria or their bacteriocins could be suitable candidates for food preservation and specially useful in the our regional dairy industry.

Histopathological Changes Induced By Staphylococcal Enterotoxin ...

African Journals Online (AJOL)

In this study, six Staphylococcus aureus strains isolated from contaminated yoghurt were evaluated for enterotoxigenicity. Two o the strains were enterotoxigenic and caused fluid accumulation in rabbit ileal loops. Fluid aspirated from the loops was bloody and histopathological changes in sections collected from rabbit ...

Nanodisc-based Co-immunoprecipitation for Mass Spectrometric Identification of Membrane-interacting Proteins

DEFF Research Database (Denmark)

Borch-Jensen, Jonas; Roepstorff, Peter; Møller-Jensen, Jakob

2011-01-01

enterotoxigenic Escherischia coli, GM1-nanodiscs were employed for co-immunoprecipitation. The B subunit of heat labile enterotoxin was identified as a specific interaction partner by mass spectrometry, thus demonstrating that nanodisc technology is useful for highly specific detection and identification...

Characterisation of MRSA from Malta and the description of a Maltese epidemic MRSA strain.

LENUS (Irish Health Repository)

Scicluna, E A

2010-02-01

Malta has one of the highest prevalence rates of methicillin-resistant Staphylococcus aureus (MRSA) in Europe. However, only limited typing data are currently available. In order to address this situation, 45 MRSA isolates from the Mater Dei Hospital in Msida, Malta, were characterised using DNA microarrays. The most common strain was ST22-MRSA-IV (UK-EMRSA-15, 30 isolates). Sporadic strains included ST36-MRSA-II (UK-EMRSA-16, two isolates), PVL-positive ST80-MRSA-IV (European Clone, one isolate), ST228-MRSA-I (Italian Clone\\/South German Epidemic Strain, one isolate) and ST239-MRSA-III (Vienna\\/Hungarian\\/Brazilian Epidemic Strain, one isolate). Ten MRSA isolates belonged to a clonal complex (CC) 5\\/ST149, spa type t002 strain. This strain harboured an SCCmec IV element (mecA, delta mecR, ugpQ, dcs, ccrA2 and ccrB2), as well as novel alleles of ccrA\\/B and the fusidic acid resistance element Q6GD50 (previously described in the sequenced strain MSSA476, BX571857.1:SAS0043). It also carried the gene for enterotoxin A (sea) and the egc enterotoxin locus, as well as (in nine out of ten isolates) genes encoding the toxic shock syndrome toxin (tst1) and enterotoxins C and L (sec, sel). While the presence of the other MRSA strains suggests foreign importation due to travel between Malta and other European countries, the CC5\\/t002 strain appears, so far, to be restricted to Malta.

Identification of virulence factors in 16S-23S rRNA intergenic spacer genotyped Staphylococcus aureus isolated from water buffaloes and small ruminants.

Science.gov (United States)

Cremonesi, P; Zottola, T; Locatelli, C; Pollera, C; Castiglioni, B; Scaccabarozzi, L; Moroni, P

2013-01-01

Staphylococcus aureus is an important human and animal pathogen, and is regarded as an important cause of intramammary infection (IMI) in ruminants. Staphylococcus aureus genetic variability and virulence factors have been well studied in veterinary medicine, especially in cows as support for control and management of IMI. The aim of the present study was to genotype 71 Staph. aureus isolates from the bulk tank and foremilk of water buffaloes (n=40) and from udder tissue (n=7) and foremilk (n=24) from small ruminants. The method used was previously applied to bovine Staph. aureus and is based on the amplification of the 16S-23S rRNA intergenic spacer region. The technique applied was able to identify different Staph. aureus genotypes isolated from dairy species other than the bovine species, and cluster the genotypes according to species and herds. Virulence gene distribution was consistent with genotype differentiation. The isolates were also characterized through determination of the presence of 19 virulence-associated genes by specific PCR. Enterotoxins A, C, D, G, I, J, and L were associated with Staph. aureus isolates from buffaloes, whereas enterotoxins C and L were linked to small ruminants. Genes coding for methicillin resistance, Panton-Valentine leukocidin, exfoliative toxins A and B, and enterotoxins B, E, and H were undetected. These findings indicate that RNA template-specific PCR is a valid technique for typing Staph. aureus from buffaloes and small ruminants and is a useful tool for understanding udder infection epidemiology. Copyright © 2013 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Fate and effect of ingested Bacillus cereus spores and vegetative cells in the intestinal tract of human-flora-associated rats

DEFF Research Database (Denmark)

Wilcks, Andrea; Hansen, Bjarne Munk; Hendriksen, Niels Bohse

2006-01-01

The fate and effect of Bacillus cereus F4433/73R in the intestine of human-flora-associated rats was studied using bacteriological culturing techniques and PCR-denaturing gradient gel electrophoresis in combination with cell assays and immunoassays for detection of enterotoxins. In faecal samples...

TCR-Vß8 as alternative to animal testing for quantifying active SEE

Science.gov (United States)

Staphylococcal food poisoning is a result of ingestion of Staphylococcal enterotoxins (SEs) produced by the bacterium Staphylococcus aureus. SEs cause gastroenteritis and also cause activation of T cells and massive cytokine release. A current method for the detection of active SEs relies on its eme...

The N-Terminal Flanking Region of the Invariant Chain Peptide Augments the Immunogenicity of a Cryptic “Self” Epitope from a Tumor-Associated Antigen

NARCIS (Netherlands)

Hess, A.D.; Thoburn, C.; Chen, W.; Miura, Y.; Wall, E. van der

2001-01-01

The N-terminal flanking region of the invariant chain peptide termed CLIP appears to have superagonistic properties interacting with the T cell receptor and the MHC class II molecule at or near the binding site for the bacterial superantigen Staphylococcal enterotoxin B (SEB). The present studies

Studies on the irradiation of toxins of Clostridium botulinum and Staphylococcus aureus

International Nuclear Information System (INIS)

Rose, S.A.; Bailey, N.E.; Stringer, M.F.; Modi, N.K.; Tranter, H.S.; Hambleton, P.

1989-01-01

The effects of irradiation of Clostridium botulinum neutotoxin type A (BNTA) and staphylococcal enterotoxin A (SEA) in gelatin phosphate buffer and cooked mince beef slurries were investigated. Estimation of toxins by immunoassays showed that in buffer, toxins were destroyed by irradiation at 8.0 kGy; in mince slurries however, 45% of BTNA and 27-34% of SEA remained after this level of irradiation. At 23.7 kGy, over twice the dose of irradiation proposed for legal acceptance in the UK, 15% of BNTA and 16-26% of SEA still remained. Increasing concentrations of mince conferred increased protection against the effect of irradiation on both toxins. The biological activity of BNTA was more sensitive to irradiation than the immunological activity. Staphylococcal enterotoxin was more resistant to irradiation than BNTA. Irradiation should therefore only be used in conjunction with good manufacturing practices to prevent microbial proliferation and toxin production prior to irradiation. (author)

Studies on the irradiation of toxins of Clostridium botulinum and Staphylococcus aureus

Energy Technology Data Exchange (ETDEWEB)

Rose, S A; Bailey, N E; Stringer, M F [Campden Food and Drink Research Association, Chipping Campden (UK); Modi, N K; Tranter, H S [Porton International plc., London (UK); Hambleton, P [Centre for Applied Microbiological Research, Porton Down (UK)

1988-10-01

The effects of irradiation of Clostridium botulinum neurotoxin type A (BNTA) and staphylococcal enterotoxin A (SEA) in gelatin phosphate buffer and cooked mince beef slurries were investigated. Estimation of toxins by immunoassays showed that in buffer, toxins were destroyed by irradiation at 8.0 kGy; in mince slurries however, 45% of BTNA and 27-34% of SEA remained after this level of irradiation. At 23.7 kGy, over twice the dose of irradiation proposed for legal acceptance in the UK, 15% of BNTA and 16-26% of SEA still remained. Increasing concentrations of mince conferred increased protection against the effect of irradiation on both toxins. The biological activity of BNTA was more sensitive to irradiation than the immunological activity. Staphylococcal enterotoxin was more resistant to irradiation than BNTA. Irradiation should therefore only be used in conjunction with good manufacturing practices to prevent microbial proliferation and toxin production prior to irradiation. (author).

Enterotoxigenic Genes in strains of Staphylococcus spp., isolated from cheese made in Pamplona-Colombia

Directory of Open Access Journals (Sweden)

Fanny Herrera A.

2015-01-01

Full Text Available Objective. To determine the incidence of coagulase-positive strains of enterotoxigenic Staphylococcus in doble crema (double cream cheese samples produced in Pamplona. Materials and methods. Bacterial isolation was performed following the routine method for coagulase positive Staphylococcus provided by the Colombian Technical Standard 4779, by using Baird Parker medium with confirmation of typical colonies by performing the coagulase test. Detection of genes for principal enterotoxins was done by PCR. Results. The prevalence of coagulase positive Staphylococcus in cheese samples was 31%, with 27% of the samples failing to meet the requirements of the NTC 750. In 24.6% of the studied isolates, genes for enterotoxin production were detected. The presence, in the isolated strains, of genes for SEB, SEA and SED was 18.5%, 4.6% and 3.0%, respectively. Conclusions. The significant presence of enterotoxigenic genes found in the isolates obtained from samples of double cream cheese made in Pamplona, suggests an important hazard to the health of consumers.

The 2.3 {angstrom} crystal structure of cholera toxin B subunit pentamer: Choleragenoid

Energy Technology Data Exchange (ETDEWEB)

Zhang, Rong-Guang; Westbrook, M.L. [Argonne National Lab., IL (United States); Maulik, P.R.; Reed, R.A.; Shipley, G. [Boston Univ., MA (United States). School of Medicine; Westbrook, E.M. [Argonne National Lab., IL (United States)]|[Northwestern Univ., Evanston, IL (United States); Scott, D.L.; Otwinowski, Z. [Yale Univ., New Haven, CT (United States)

1996-02-01

Cholera toxin, a heterohexameric AB{sub 5} enterotoxin released by Vibrio cholera, induces a profuse secretory diarrhea in susceptible hosts. Choleragenoid, the B subunit pentamer of cholera toxin, directs the enzymatic A subunit to its target by binding to GM{sub 1} gangliosides exposed on the luminal surface of intestinal epithelial cells. We have solved the crystal structure of choleragenoid at 2.3 {Angstrom} resolution by combining single isomorphous replacement with non-crystallographic symmetry averaging. The structure of the B subunits, and their pentameric arrangement, closely resembles that reported for the intact holotoxin (choleragen), the heat-labile enterotoxin from E. coli, and for a choleragenoid-GM{sub 1} pentasaccharide complex. In the absence of the A subunit the central cavity of the B pentamer is a highly solvated channel. The binding of the A subunit or the receptor pentasaccharide to choleragenoid has only a modest effect on the local stereochemistry and does not perceptibly alter the subunit interface.

Prevalence and Toxin Characteristics of Bacillus thuringiensis Isolated from Organic Vegetables.

Science.gov (United States)

Kim, Jung-Beom; Choi, Ok-Kyung; Kwon, Sun-Mok; Cho, Seung-Hak; Park, Byung-Jae; Jin, Na Young; Yu, Yong Man; Oh, Deog-Hwan

2017-08-28

The prevalence and toxin characteristics of Bacillus thuringiensis isolated from 39 organic vegetables were investigated. B. thuringiensis was detected in 30 out of the 39 organic vegetables (76.9%) with a mean value of 2.60 log CFU/g. Twenty-five out of the 30 B. thuringiensis isolates (83.3%) showed insecticidal toxicity against Spodoptera exigua . The hblCDA, nheABC , and entFM genes were found to be the major toxin genes, but the ces gene was not detected in any of the tested B. thuringiensis isolates. The hemolysin BL enterotoxin was detected in all 30 B. thuringiensis isolates (100%). The non-hemolytic enterotoxin complex was found in 27 out of 30 B. thuringiensis isolates (90.0%). The B. thuringiensis tested in this study had similar toxin gene characteristics to B. cereus , which possessed more than one toxin gene. B. thuringiensis could have the potential risk of foodborne illness based on the toxin genes and toxin-producing ability.

Studies on the irradiation of toxins of Clostridium botulinum and Staphylococcus aureus

International Nuclear Information System (INIS)

Rose, S.A.; Bailey, N.E.; Stringer, M.F.; Modi, N.K.; Tranter, H.S.; Hambleton, P.

1988-01-01

The effects of irradiation of Clostridium botulinum neurotoxin type A (BNTA) and staphylococcal enterotoxin A (SEA) in gelatin phosphate buffer and cooked mince beef slurries were investigated. Estimation of toxins by immunoassays showed that in buffer, toxins were destroyed by irradiation at 8.0 kGy; in mince slurries however, 45% of BTNA and 27-34% of SEA remained after this level of irradiation. At 23.7 kGy, over twice the dose of irradiation proposed for legal acceptance in the UK, 15% of BNTA and 16-26% of SEA still remained. Increasing concentrations of mince conferred increased protection against the effect of irradiation on both toxins. The biological activity of BNTA was more sensitive to irradiation than the immunological activity. Staphylococcal enterotoxin was more resistant to irradiation than BNTA. Irradiation should therefore only be used in conjunction with good manufacturing practices to prevent microbial proliferation and toxin production prior to irradiation. (author)

Studies on the irradiation of toxins of Clostridium botulinum and Staphylococcus aureus

Energy Technology Data Exchange (ETDEWEB)

Rose, S.A.; Bailey, N.E.; Stringer, M.F. (Campden Food and Drink Research Association, Chipping Campden (UK)); Modi, N.K.; Tranter, H.S. (Centre for Applied Micobiological Research, Porton Down, (UK)); Hambleton, P. (Porton International plc, London (UK))

1989-09-01

The effects of irradiation of Clostridium botulinum neutotoxin type A (BNTA) and staphylococcal enterotoxin A (SEA) in gelatin phosphate buffer and cooked mince beef slurries were investigated. Estimation of toxins by immunoassays showed that in buffer, toxins were destroyed by irradiation at 8.0 kGy; in mince slurries however, 45% of BTNA and 27-34% of SEA remained after this level of irradiation. At 23.7 kGy, over twice the dose of irradiation proposed for legal acceptance in the UK, 15% of BNTA and 16-26% of SEA still remained. Increasing concentrations of mince conferred increased protection against the effect of irradiation on both toxins. The biological activity of BNTA was more sensitive to irradiation than the immunological activity. Staphylococcal enterotoxin was more resistant to irradiation than BNTA. Irradiation should therefore only be used in conjunction with good manufacturing practices to prevent microbial proliferation and toxin production prior to irradiation. (author).

THE STUDY OF ENTEROTOXIGENICITY OF THE BIOTYPE 1A YERSINIA ENTEROCOLITICA

Directory of Open Access Journals (Sweden)

E. A. Bogumilchik

2011-01-01

Full Text Available Abstract. The representatives of Y. enterocolitica biotype 1А which are considered as nonpathogenic microorganisms were tested for production of the thermostable enterotoxin YST B (Yersinia Stable Toxin. This toxin is characterized by strong toxic action and it can bring on diarrhea in human and animals. The chromosome gene of thermostable enterotoxin ystB was detected by PCR in 87.1% out of 116 studied strains of different origin and territorial isolation. To determine toxin production in vitro the studied strains cultivated in various conditions: in 26°C and 37°С in usual culture medium and in 37°С in the medium corresponded to the content of intestine. In part of the studied strains the toxin production was revealed on the model of newborn mice in both temperature regimes of cultivation 26°С and 37°С. The study of toxin production in representatives of Y. enterocolitica biotype 1А showed their possible role as etiological agents of diarrhea.

Staphylococcal poisoning foodborne outbreak: epidemiological investigation and strain genotyping.

Science.gov (United States)

Gallina, S; Bianchi, D M; Bellio, A; Nogarol, C; Macori, G; Zaccaria, T; Biorci, F; Carraro, E; Decastelli, L

2013-12-01

In June 2011, an outbreak of Staphylococcus aureus enterotoxin food poisoning gastroenteritis occurred in Turin, Italy, following a catered dinner party at a private home. Within a few hours, 26 of the 47 guests experienced gastrointestinal illness, and 9 were hospitalized. A retrospective cohort study using a standardized questionnaire was carried out, and the risk ratios for each food item were calculated. The analysis indicated consumption of seafood salad as the most probable cause of the outbreak (risk ratio = 11.72; 95 % confidence interval, 1.75 to 78.54). Biological samples were collected from four of the hospitalized guests (stool and vomit), nasal mucosa swabs from three food handlers employed with the caterer, and available food residuals. All stool and vomit samples tested positive for enterotoxigenic S. aureus. As residues of the seafood salad were no longer available for sampling, suspected contamination could not be verified. However, no other food was found contaminated by S. aureus or its enterotoxins. All isolates from the biological samples were characterized at the genomic level by means of two multiplex PCR protocols to determine the presence of genes encoding staphylococcal enterotoxins, pulsed-field gel electrophoresis and staphylococcal protein A gene (spa) typing to describe their genetic profiles. All the isolates presented genes encoding SEA and SEI; the pulsed-field gel electrophoresis genetic profiles revealed the same pulsotype in the microorganism isolated from the hospitalized guests as in one of the isolates from a food handler's nasal mucosa, and the spa typing analysis reported two closely related spa types (t701 and t267), implicating the food handler as the most likely outbreak source.

Modulation of Interleukin-8 and staphylococcal flora by Avène hydrotherapy in patients suffering from chronic inflammatory dermatoses.

Science.gov (United States)

Casas, C; Ribet, V; Alvarez-Georges, S; Sibaud, V; Guerrero, D; Schmitt, A-M; Redoulès, D

2011-02-01

A number of studies argue in favour of an important role of microbial colonization, in particular of Staphylococcus aureus, in triggering atopic dermatitis (AD) flare-up and psoriasis, in particular through the superantigenic properties of toxins generated by S. aureus. The aim of this study was to assess the efficacy of a 3-week Avène hydrotherapy on the skin surface of patients suffering from psoriasis or atopic dermatitis. Skin samples were taken from healthy subjects or atopic (n = 18) or psoriatic patients (n = 39) undergoing hydrotherapy at Avène at the beginning (D0) and the end of treatment (D18). The severity of the dermatosis was evaluated according to SCORing Atopic Dermatitis (SCORAD) or Psoriasis Area Severity Index (PASI) scores at D0 and D18. Marker of inflammation interleukin 8 (IL-8), S. aureus colonization (protein A) and enterotoxins were assessed in skin samples using RT-PCR. At D0, significant differences were observed between healthy subjects and atopic or psoriatic patients in all the parameters evaluated (IL-8, protein A). At the end of the hydrotherapy, a significant decrease in SCORAD was associated with a significant reduction of IL-8, S. aureus colonization and enterotoxin D in patients with atopic dermatitis. Similarly, a significant decrease in PASI was associated with a significant reduction of IL-8, S. aureus colonization and enterotoxin N in patients with psoriasis. This study demonstrates the positive effects of Avène hydrotherapy on the skin of patients suffering from chronic dermatosis, with decreased inflammation and reduced colonization by S. aureus. © 2010 The Authors. JEADV © 2010 European Academy of Dermatology and Venereology.

Multilocus Sequence Typing And Antibiotic Resistance Of Staphylococcus Aureus Isolated From The Brazilian Dairy Industry

DEFF Research Database (Denmark)

Dittmann, Karen Kiesbye; Chaul, Luiza; Lee, Sarah

2015-01-01

Staphylococcus aureus is a common cause of food poisoning due to enterotoxin production. This is particularly an issue in the dairy industry, where S. aureus can contaminate the product e.g. from raw milk or the handlers. In Brazil, soft cheese is mainly produced in small dairy plants where good ...

Staphylococcal enterotoxins stimulate lymphoma-associated immune dysregulation

DEFF Research Database (Denmark)

Krejsgaard, Thorbjørn Frej; Willerslev-Olsen, Andreas; Lindahl, Lise Maria

2014-01-01

a cascade of events involving cell-cell and asymmetric cytokine interactions between malignant and benign T cells, which stimulated the malignant T cells to express high levels of IL-10. Much evidence supports that malignant activation of the Stat3/IL-10 axis plays a key role in driving the immune...... dysregulation and severe immunodeficiency that characteristically develops in CTCL patients. The present findings thereby establish a novel link between SEs and immune dysregulation in CTCL strengthening the rationale for antibiotic treatment of colonized patients with severe or progressive disease....

Prevalence, antibiotic-resistance properties and enterotoxin gene ...

African Journals Online (AJOL)

Methods: Three-hundred milk-based infant foods were collected and immediately transferred to the laboratory. ... and entFM (55.5 %) were the most commonly detected enterotoxigenic genes. Bacteria showed the ... conditions like pasteurization and sterilization. [2,3]. ... spaghetti, noodle, milk powder and infant formula.

Prevalence, antibiotic-resistance properties and enterotoxin gene ...

African Journals Online (AJOL)

milk-based infant foods in Iran, represent an important public health issue which should be considered ... Keywords: Prevalence, Bacillus cereus, Antibiotic resistance, Enterotoxigenic genes, Milk-based infant food. Tropical Journal of Pharmaceutical Research is indexed by Science ..... and cereals collected in Korea.

DNA microarray genotyping and virulence and antimicrobial resistance gene profiling of methicillin-resistant Staphylococcus aureus bloodstream isolates from renal patients.

LENUS (Irish Health Repository)

McNicholas, Sinead

2012-02-01

Thirty-six methicillin-resistant Staphylococcus aureus (MRSA) bloodstream isolates from renal patients were genetically characterized by DNA microarray analysis and spa typing. The isolates were highly clonal, belonging mainly to ST22-MRSA-IV. The immune evasion and enterotoxin gene clusters were found in 29\\/36 (80%) and 33\\/36 (92%) isolates, respectively.

DNA microarray genotyping and virulence and antimicrobial resistance gene profiling of methicillin-resistant Staphylococcus aureus bloodstream isolates from renal patients.

LENUS (Irish Health Repository)

McNicholas, Sinead

2011-12-01

Thirty-six methicillin-resistant Staphylococcus aureus (MRSA) bloodstream isolates from renal patients were genetically characterized by DNA microarray analysis and spa typing. The isolates were highly clonal, belonging mainly to ST22-MRSA-IV. The immune evasion and enterotoxin gene clusters were found in 29\\/36 (80%) and 33\\/36 (92%) isolates, respectively.

Subinhibitory concentrations of perilla oil affect the expression of secreted virulence factor genes in Staphylococcus aureus.

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Jiazhang Qiu

Full Text Available BACKGROUND: The pathogenicity of staphylococcus aureus is dependent largely upon its ability to secrete a number of virulence factors, therefore, anti-virulence strategy to combat S. aureus-mediated infections is now gaining great interest. It is widely recognized that some plant essential oils could affect the production of staphylococcal exotoxins when used at subinhibitory concentrations. Perilla [Perilla frutescens (L. Britton], a natural medicine found in eastern Asia, is primarily used as both a medicinal and culinary herb. Its essential oil (perilla oil has been previously demonstrated to be active against S. aureus. However, there are no data on the influence of perilla oil on the production of S. aureus exotoxins. METHODOLOGY/PRINCIPAL FINDINGS: A broth microdilution method was used to determine the minimum inhibitory concentrations (MICs of perilla oil against S. aureus strains. Hemolysis, tumour necrosis factor (TNF release, Western blot, and real-time RT-PCR assays were performed to evaluate the effects of subinhibitory concentrations of perilla oil on exotoxins production in S. aureus. The data presented here show that perilla oil dose-dependently decreased the production of α-toxin, enterotoxins A and B (the major staphylococcal enterotoxins, and toxic shock syndrome toxin 1 (TSST-1 in both methicillin-sensitive S. aureus (MSSA and methicillin-resistant S. aureus (MRSA. CONCLUSIONS/SIGNIFICANCE: The production of α-toxin, SEA, SEB, and TSST-1 in S. aureus was decreased by perilla oil. These data suggest that perilla oil may be useful for the treatment of S. aureus infections when used in combination with β-lactam antibiotics, which can increase exotoxins production by S. aureus at subinhibitory concentrations. Furthermore, perilla oil could be rationally applied in food systems as a novel food preservative both to inhibit the growth of S. aureus and to repress the production of exotoxins, particularly staphylococcal enterotoxins.

Outbreak of Staphylococcal food poisoning due to SEA-producing Staphylococcus aureus.

Science.gov (United States)

Johler, Sophia; Tichaczek-Dischinger, Petra S; Rau, Jörg; Sihto, Henna-Maria; Lehner, Angelika; Adam, Maja; Stephan, Roger

2013-09-01

In 2008, 150 people gathered for a wedding celebration in Baden-Württemberg, Germany. Three hours after ingestion of a variety of foods including pancakes filled with minced chicken, several guests exhibited symptoms of acute gastroenteritis such as vomiting, diarrhea, fever, and ague. Twelve guests were reported to have fallen ill, with nine of these seeking medical care in hospitals. At least four patients were admitted to the hospital and received inpatient treatment, among them a 2-year-old child and a woman in the 4th month of pregnancy. Within 24 h of the event, an investigative team collected a variety of samples including refrigerated leftovers, food in the storage unit of the caterer, nasal swabs of the caterer, as well as 21 environmental swabs. Five stool samples from patients were provided by the hospitals. Staphylococcus aureus isolates were gathered from eight samples, among them nasal swabs of the caterer, food samples, and one stool sample. Fourier transform-infrared spectroscopy was used for species identification and for primary clustering of the isolates in a similarity tree. The isolates were further characterized by spa typing and pulsed-field gel electrophoresis, and a DNA microarray was used to determine the presence/absence of genes involved in virulence and antimicrobial resistance. We were able to match an enterotoxigenic strain from the stool sample of a patient to isolates of the same strain obtained from food and the nasal cavity of a food handler. The strain produced the enterotoxin SEA and the toxic shock syndrome toxin-1, and was also found to exhibit the genes encoding enterotoxins SEG and SEI, as well as the enterotoxin gene cluster egc. This is one of only a few studies that were able to link a staphylococcal food poisoning outbreak to its source.

Toxicity and immunogenicity of Enterotoxigenic Escherichia coli heat-labile and heat-stable toxoid fusion 3xSTa(A14Q-LT(S63K/R192G/L211A in a murine model.

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Chengxian Zhang

Full Text Available Diarrhea is the second leading cause of death to young children. Enterotoxigenic Escherichia coli (ETEC are the most common bacteria causing diarrhea. Adhesins and enterotoxins are the virulence determinants in ETEC diarrhea. Adhesins mediate bacterial attachment and colonization, and enterotoxins including heat-labile (LT and heat-stable type Ib toxin (STa disrupt fluid homeostasis in host cells that leads to fluid hyper-secretion and diarrhea. Thus, adhesins and enterotoxins have been primarily targeted in ETEC vaccine development. A recent study reported toxoid fusions with STa toxoid (STa(P13F fused at the N- or C-terminus, or inside the A subunit of LT(R192G elicited neutralizing antitoxin antibodies, and suggested application of toxoid fusions in ETEC vaccine development (Liu et al., Infect. Immun. 79:4002-4009, 2011. In this study, we generated a different STa toxoid (STa(A14Q and a triple-mutant LT toxoid (LT(S63K/R192G/L211A, tmLT, constructed a toxoid fusion (3xSTa(A14Q-tmLT that carried 3 copies of STa(A14Q for further facilitation of anti-STa immunogenicity, and assessed antigen safety and immunogenicity in a murine model to explore its potential for ETEC vaccine development. Mice immunized with this fusion antigen showed no adverse effects, and developed antitoxin antibodies particularly through the IP route. Anti-LT antibodies were detected and were shown neutralizing against CT in vitro. Anti-STa antibodies were also detected in the immunized mice, and serum from the IP immunized mice neutralized STa toxin in vitro. Data from this study indicated that toxoid fusion 3xSTa(A14Q-tmLT is safe and can induce neutralizing antitoxin antibodies, and provided helpful information for vaccine development against ETEC diarrhea.

Enterotoxigenicity and Antimicrobial Resistance of Staphylococcus aureus Isolated from Retail Food in China

Science.gov (United States)

Wang, Wei; Baloch, Zulqarnain; Jiang, Tao; Zhang, Cunshan; Peng, Zixin; Li, Fengqin; Fanning, Séamus; Ma, Aiguo; Xu, Jin

2017-01-01

Staphylococcus aureus is one of the most common causes of zoonotic agent in the world, which are attributable to the contamination of food with enterotoxins. In this study, a total of 1,150 S. aureus isolates were cultured from 27,000 retail foods items from 203 cities of 24 provinces in China in 2015 and were test for antimicrobial susceptibility. Additionally, the role of the genes responsible for the staphylococcal enterotoxins (SEA to SEE), methicillin resistance (mecA) and the toxigenic capabilities were also assessed. The results showed that 4.3% retail foods were contaminated with S. aureus, and 7.9% retail foods isolates were mecA positive. Some 97.6% of S. aureus isolates were resistant to at least one antimicrobial compound, and 57.5% of these were multi drug resistant (MDR). Resistance to penicillin (83.7%, 963/1,150), was common, followed by linezolid (67.7%, 778/1,150) and erythromycin (52.1%, 599/1,150). The isolates cultured from raw meats showed high levels of resistant to tetracycline (42.8%), ciprofloxacin (17.4%), and chloramphenicol (12.0%) and expressed a MDR phenotype (62.4%). A total of 29.7% S. aureus isolates harbored the classical SEs genes (sea, seb, sec, and sed). The sea and seb genes were the most frequent SEs genes detected. Of note, 22% of the SEs genes positive S. aureus harbored two or three SEs genes, and 16 isolates were confirmed with the capacity to simultaneously produce two or three enterotoxin types. Moreover, nearly 50% of the MRSA isolates were positive for at least one SE gene in this study. Therefore, it is important to monitor the antimicrobial susceptibility and enterotoxigenicity of MDR S. aureus and MRSA in the food chain and to use these data to develop food safety measures, designed to reduce the contamination and transmission of this bacterium. PMID:29209290

Optimizing Two-Color Semiconductor Nanocrystal Immunoassays in Single Well Microtiter Plate Formats

Directory of Open Access Journals (Sweden)

W. Russ Algar

2011-08-01

Full Text Available The simultaneous detection of two analytes, chicken IgY (IgG and Staphylococcal enterotoxin B (SEB, in the single well of a 96-well plate is demonstrated using luminescent semiconductor quantum dot nanocrystal (NC tracers. The NC-labeled antibodies were prepared via sulfhydryl-reactive chemistry using a facile protocol that took

Laser Induced Breakdown Spectroscopy (LIBS)

Science.gov (United States)

2010-03-31

Radium 226 Page 34 Nitrogen mustard (HN) Lassa Polonium 210 Phosgene oxime (CX) Staphylococcal Enterotoxin B (SEB) ( Food Poisoning...sample preparation. Diverse solid or liquid samples including environmental, biological, geological, industrial, forensic, food , thin coatings...Thallium 201 Hydrogen cyanide (AC) FUNGI Thorium 232 Cyanogen chloride (CK) Aflatoxin Lead 210 Arsine (SA) Bacillus Sodium 22

Clostridium perfringens in London, July 2009: two weddings and an outbreak.

Science.gov (United States)

Eriksen, J; Zenner, D; Anderson, S R; Grant, K; Kumar, D

2010-06-24

Food poisoning outbreaks caused by Clostridium perfringens enterotoxin occur occasionally in Europe but have become less common in recent years. This paper presents the microbiological and epidemiological results of a large C. perfringens outbreak occurring simultaneously at two weddings that used the same caterer. The outbreak involved several London locations and required coordination across multiple agencies. A case-control study (n=134) was carried out to analyse possible associations between the food consumed and becoming ill. Food, environmental and stool samples were tested for common causative agents, including enterotoxigenic C. perfringens. The clinical presentation and the epidemiological findings were compatible with C. perfringens food poisoning and C. perfringens enterotoxin was detected in stool samples from two cases. The case-control study found statistically significant associations between becoming ill and eating either a specific chicken or lamb dish prepared by the same food handler of the implicated catering company. A rapid outbreak investigation with preliminary real-time results and the successful collaboration between the agencies and the caterer led to timely identification and rectification of the failures in the food handling practices.

Evaluation of PCR and DNA hybridization protocols for detection of viable enterotoxigenic Clostridium perfringens in irradiated beef

International Nuclear Information System (INIS)

Baez, L.A.; Juneja, V.K.; Thayer, D.W.; Sackitey, S.

1997-01-01

The sensitivity of DNA hybridization and polymerase chain reaction (PCR), was evaluated in irradiated cooked and raw beef samples. A membrane-based colony hybridization assay and a PCR protocol, both with specificity for the enterotoxin A gene of Clostridium perfringens, were compared with viable plate counts. The results of the colony hybridization procedure were in agreement with viable plate counts for detection and enumeration of enterotoxigenic C. perfringens. The PCR procedure combined a 4 h enrichment followed by a nucleic acid extraction step and assessed the amplification of 183 and 750 base pair enterotoxin gene targets. Detection of C. perfringens by PCR did not show a reliable correlation with viable plate counts or the colony hybridization assay. C. perfringens killed by irradiation were not detected by the plate count or colony hybridization methods; however, killed cells were detected with the PCR technique. By relying on the growth of viable cells for detection and/or enumeration, the colony hybridization and plate count methods provided a direct correlation with the presence of viable bacteria

Food irradiation and bacterial toxins

International Nuclear Information System (INIS)

Tranter, H.S.; Modi, N.K.; Hambleton, P.; Melling, J.; Rose, S.; Stringer, M.F.

1987-01-01

The authors' findings indicate that irradiation confers no advantage over heat processing in respect of bacterial toxins (clostridium botulinum, neurotoxin A and staphylococcal enterotoxin A). It follows that irradiation at doses less than the ACINF recommended upper limit of 10 kGy could not be used to improve the ambient temperature shelf life on non-acid foods. (author)

Biotype characterization of Staphylococcus aureus isolated from milk and dairy products of private production in the western regions of Ukraine

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M. D. Kukhtyn

2017-08-01

Full Text Available Prevention of foodborne diseases is a priority for the world health system. In the process of manufacturing milk and dairy products, the most important factor compromising their safety is seeding with a conditionally pathogenic and pathogenic microflora. Salmonella, Escherichia coli, Listeria and other microorganisms that reproduce in dairy products without changing their organoleptic properties are a particular danger. Staphylococcus aureus is an opportunistic, conditionally pathogenic microorganism that often contaminates raw milk and dairy products. The aim of the research presented in this article was to determine the dissemination of S. aureus in milk and milk products of household production in the western regions of Ukraine, to identify the biotypes of S. aureus, production of enterotoxins and the presence of methicillin-resistant strains. S. aureus was isolated on BD Baird-Parker Agar. The biotypes of S. aureus were determined according to Meer. The determination of MRSA was carried out on the chromogenic Agar chromID MRSA ("Biomerioux", Russia. The mecA gene was determined using the LightCycler MRSA Advanced Test with LightCycler 2.0 primer (Roche Molecular Biochemicals, Germany. To determine staphylococcal enterotoxins, the test system RIDASCREENSET A, B, C, D, E (R-Biopharm AG, Darmstadt, Germany was used. We isolated saprophyte staphylococci from milk of raw and dairy products in western regions of Ukraine in 82.7–97.4% of samples. S. aureus is much more rarely isolated from these dairy products, so it was isolated from sour cream at 62.8 ± 0.9%, from milk at 35.5 ± 1.3% and cottage cheese at 23.0 ± 1.6%. Of the most well known biotypes of S. aureus present in milk of raw and dairy products of domestic production, two ecological types were distinguished: human and cattle. In this case S. aureus var. hominis was isolated more often than in S. aureus var. bovis. This gives grounds to believe that the main source of

Enterotoxigenic gene profiles of Bacillus cereus and Bacillus megaterium isolates recovered from honey Búsqueda de factores de virulencia en cepas de Bacillus cereus y de Bacillus megaterium aisladas de miel

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A. C. López

2010-09-01

Full Text Available One hundred and thirty two Bacillus cereus and 52 Bacillus megaterium isolates from honeys were evaluated for the presence of genes encoding enterotoxin HBL, enterotoxin-T, cytotoxin K and the NHE complex, respectively. The relationship between hemolytic and coagulase activity and its correlation with the presence of the four mentioned enterotoxins was determined by principal component analysis (PCA. PCA in B. cereus revealed a positive correlation among free coagulase, hemolysis and the presence of genes hblA, hblB, hblC, hblD (HBL complex and bceT (enterotoxin-T, but no correlation with the clumping factor (bound coagulase and the presence of sequences of the NHE complex. On the other hand, PCA in B. megaterium showed a high positive correlation between coagulase (bound and free and the haemolytic activity but no correlation in relation to the presence of genes of the HBL complex, cytotoxin K, enterotoxin T and the NHE complex. To our knowledge, this is the first report of the detection of cytotoxin K and of the NHE complex genes in B. megaterium. The relationship between the coagulase activity and the presence of virulence factors has not been described before in the genus Bacillus, being this work the first report of this correlation. Interestingly, the presence of the cytK gene was almost independent of the presence of the rest of virulence factors herein analyzed both in B. cereus and B. megaterium populations. Our results suggest that honey could be a possible vehicle for foodborne illness due to the presence of toxigenic B. cereus and B. megaterium strains containing different virulence factors.Se evaluaron 132 aislamientos de Bacillus cereus y 52 de Bacillus megaterium provenientes de mieles de distintos orígenes geográficos para investigar la presencia de secuencias de ADN relacionadas con genes de virulencia y su posible correlación con la actividad hemolítica y coagulasa. Con respecto a los genes de virulencia, se analizaron por

Volunteer Challenge With Enterotoxigenic Escherichia coli That Express Intestinal Colonization Factor Fimbriae CS17 and CS19

Science.gov (United States)

2011-07-01

Serotype was determined by classic serological methods at the Universidad Nacional Aut6noma de Mexico [UNAMl. H- indrcates non-motility. b CF...Levine MM, Merson MM. Serologic differentiation between antitoxin responses to infection with Vibrio cholerae and enterotoxin-producing Escherichia coli...prototype cholera B subunit-colonization factor antigen cnterotoxigenic Escherichia coli vaccine. Vaccine 1993; 1[:929-34. 15. Levine MM, Nalin DR

Simulation of carbohydrate-protein interactions : Computer-aided design of a second generation GM1 mimic

NARCIS (Netherlands)

Bernardi, A; Galgano, M; Belvisi, L; Colombo, G

The oligosaccharide of ganglioside GM1 [Gal beta1-3GalNAc beta1-4(NeuAc alpha2-3)Gal beta1-4Glc beta1-1Cer] is the cellular target of two bacterial enterotoxins: the cholera toxin (CT) and the heat-labile toxin of E.coli (LT). We recently reported that the pseudosaccharide 2[Gal beta1-3GalNAc

Potential enterotoxicity and antimicrobial resistance pattern of Aeromonas species isolated from pet turtles and their environment.

Science.gov (United States)

Wimalasena, S H M P; Shin, Gee-Wook; Hossain, Sabrina; Heo, Gang-Joon

2017-05-23

To investigate the potential enterotoxicity and antimicrobial resistance of aeromonads from pet turtles as a risk for human infection, one hundred and two Aeromonas spp. were isolated from the feces, skin and rearing environments of pet turtles and identified by biochemical and gyrB sequence analyses. Aeromonas enteropelogenes was the predominant species among the isolates (52.9%) followed by A. hydrophila (32.4%), A. dharkensis (5.9%), A. veronii (4.9%) and A. caviae (3.9%). Their potential enterotoxicities were evaluated by PCR assays for detecting genes encoding cytotoxic enterotoxin (act) and two cytotonic enterotoxins (alt and ast). 75.8% of A. hydrophila isolates exhibited the act + /alt + /ast + genotype, whereas 94.4% of A. enteropelogenes isolates were determined to be act - /alt - /ast - . In an antimicrobial susceptibility test, most isolates were susceptible to all tested antibiotics except amoxicillin, ampicillin, cephalothin, chloramphenicol and tetracycline. Non-susceptible isolates to penicillins (ampicillin and amoxicillin) and fluoroquinolones (ciprofloxacin and norfloxacin) were frequently observed among the A. enteropelogenes isolates. Few isolates were resistant to imipenem, amikacin, ceftriaxone and cefotaxime. Collectively, these results suggest that pet turtles may pose a public health risk of infection by enterotoxigenic and antimicrobial resistant Aeromonas strains.

Clostridium perfringens Sporulation and Sporulation-Associated Toxin Production

Science.gov (United States)

Li, Jihong; Paredes-Sabja, Daniel; Sarker, Mahfuzur R.; McClane, Bruce A.

2015-01-01

The ability of Clostridium perfringens to form spores plays a key role during the transmission of this Gram-positive bacterium to cause disease. Of particular note, the spores produced by food poisoning strains are often exceptionally resistant to food environment stresses such as heat, cold and preservatives, which likely facilitates their survival in temperature-abused foods. The exceptional resistance properties of spores made by most type A food poisoning strains and some type C foodborne disease strains involves their production of a variant small acid soluble protein-4 that binds more tightly to spore DNA compared to the small acid soluble protein-4 made by most other C. perfringens strains. Sporulation and germination by C. perfringens and Bacillus spp. share both similarities and differences. Finally, sporulation is essential for production of C. perfringens enterotoxin, which is responsible for the symptoms of C. perfringens type A food poisoning, the second most common bacterial foodborne disease in the USA. During this foodborne disease, C. perfringens is ingested with food and then, using sporulation-specific alternate sigma factors, this bacterium sporulates and produces the enterotoxin in the intestines. PMID:27337447

Determination of haemolytic and non haemolytic genes profiles of Bacillus cereus strains isolated from food samples by polymerase chain reaction (pcr) technique

Science.gov (United States)

Jawad, Nisreen; Ahemd, Asmat; Abdullah, Aminah

2018-04-01

The aim of this study was to investigate the presence of Bacillus cereus and detection of enterotoxigenic genes in food samples by utilizing a Polymerase Chain Reaction technique (PCR). In this study the providence of B. cereus was carried out to food samples. The B. cereus isolates were investigated for enterotoxigenic gene. The cooked seafood, and raw milk samples were purchased from several restaurants and market in the area of (Bangi, Kajang, Serdang and UKM) Selangor, Malaysia. A total of 60 samples have been analyzed. B. cereus contamination has been formed between 1.4×105 - 3×105 cfu/mL of cooked seafood and raw milk samples. Five colonies have been detected as B. cereus using biochemical test. All B. cereus isolates named BC1 to BC27, were characterized for haemolytic enterotoxin (HBL) complex encoding genes (hblA), non-haemolytic enterotoxin encoding gene (NheA). 10 isolates have been reported to be positive towards hblA and 12 isolates were positive towards NheA. The presence of B. cereus and their enterotoxigenic genes in cooked seafood and raw milk from to food samples obtained may pose a potential risk for public health.

Microbiological quality and safety of Skipjack tuna loins (Katsuwonus Pelamis intented for canning

Directory of Open Access Journals (Sweden)

Francesco Casalinuovo

2018-03-01

Full Text Available Tuna is a food widely consumed fresh and canned as well. After catching and filleting, a pre-cooking step is normally followed by freezing and shipping to canning industry as loins. The aim of this paper was to assess the microbiological quality and safety of tuna loins (50 samples imported by an Italian canned tuna producer from two different FAO fishing areas. Total bacterial count (TBC, Coliforms, Enterobacteriaceae, Escherichia coli, Listeria monocytogenes, Salmonella, Staphylococcus aureus, Vibrio parahaemolyticus, Vibrio cholera, pH measurement, S. aureus enterotoxin and histamine levels were evaluated. Significant differences were evidenced for TBC, Enterobacteriaceae, E. coli, Coliforms and pH between fishing areas. Staphylococcus aureus was detected in all the samples tested at 1 to 3 Log cfu/g; Staphylococcus warneri, S. saprophyticus, S. epidermidis, S. hominis, S. intermedius, S. vitulinus and S. sciuri were also identified. Listeria monocytogenes, Salmonella, Vibrio parahaemolyticus, Vibrio cholera and Staphylococcus aureus enterotoxin were not detected. Histamine ranged between 0 to 5 mg/kg. The tuna loins analysed in this paper have to be considered safe; staphylococci presence represents a possible concern and has to been taken in account when risk analysis is carried out.

Prevalence and antimicrobial resistance of Bacillus cereus isolated from beef products in Egypt

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Reyad Shawish

2017-12-01

Full Text Available Foodborne pathogens have the main concern in public health and food safety. Bacillus cereus food poisoning is one of the most important foodborne pathogens worldwide. In the present study, a total of 200 random beef product samples were collected from different supermarkets located at Menofia and Cairo governorates were examined for the presence of B. cereus. In addition, the presence of some virulence encoding genes was evaluated using Multiplex PCR. Finally, the antibiogram testing was conveyed to illustrate the resistance pattern of the confirmed B. cereus. The data showed that B. cereus was recovered from 22.5%, 30%, 25%, 37.5% and 15% of the minced meat, burger, sausage, kofta, and luncheon respectively. Among the 20 examined isolates 18/20 (90% were harbor hblC enterotoxin encoding gene compared with 20/20 (100 were have cytK enterotoxin encoding gene. The isolated strains of B. cereus were resistant to penicillin G and sensitive to oxacillin, clindamycin, vancomycin, erythromycin, gentamicin, ciprofloxacin, and ceftriaxone. In all, the obtained data showed the importance of emerging B. cereus in disease control and prevention programs, and in regular clinical and food quality control laboratories in Egypt.

Effect of food-related stress conditions and loss of agr and sigB on seb promoter activity in S. aureus.

Science.gov (United States)

Sihto, Henna-Maria; Stephan, Roger; Engl, Christoph; Chen, John; Johler, Sophia

2017-08-01

Staphylococcal enterotoxin B (SEB) causes staphylococcal food poisoning and is produced in up to ten times higher quantities than other major enterotoxins. While Staphylococcus aureus growth is often repressed by competing flora, the organism exhibits a decisive growth advantage under some stress conditions. So far, data on the influence of food-related stressors and regulatory mutations on seb expression is limited and largely based on laboratory strains, which were later reported to harbor mutations. Therefore, the aim of this study was to investigate the influence of stress and regulatory mutations on seb promoter activity. To this end, transcriptional fusions were created in two strains, USA300 and HG003, carrying different seb upstream sequences fused to a blaZ reporter. NaCl, nitrite, and glucose stress led to significantly decreased seb promoter activity, while lactic acid stress resulted in significantly increased seb promoter activity. Loss of agr decreased seb promoter activity and loss of sigB increased promoter activity, with the magnitude of change depending on the strain. These results demonstrate that mild stress conditions encountered during food production and preservation can induce significant changes in seb promoter activity. Copyright © 2017 Elsevier Ltd. All rights reserved.

Single Chain Variable Fragments Produced in Escherichia coli against Heat-Labile and Heat-Stable Toxins from Enterotoxigenic E. coli.

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Christiane Y Ozaki

Full Text Available Diarrhea is a prevalent pathological condition frequently associated to the colonization of the small intestine by enterotoxigenic Escherichia coli (ETEC strains, known to be endemic in developing countries. These strains can produce two enterotoxins associated with the manifestation of clinical symptoms that can be used to detect these pathogens. Although several detection tests have been developed, minimally equipped laboratories are still in need of simple and cost-effective methods. With the aim to contribute to the development of such diagnostic approaches, we describe here two mouse hybridoma-derived single chain fragment variable (scFv that were produced in E. coli against enterotoxins of ETEC strains.Recombinant scFv were developed against ETEC heat-labile toxin (LT and heat-stable toxin (ST, from previously isolated hybridoma clones. This work reports their design, construction, molecular and functional characterization against LT and ST toxins. Both antibody fragments were able to recognize the cell-interacting toxins by immunofluorescence, the purified toxins by ELISA and also LT-, ST- and LT/ST-producing ETEC strains.The developed recombinant scFvs against LT and ST constitute promising starting point for simple and cost-effective ETEC diagnosis.

Toxin production ability of Bacillus cereus strains from food product of Ukraine

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I. Pylypenko

2017-10-01

Full Text Available Potential pathogens of foodborne toxic infections – bacterial contaminants Bacillus cereus isolated from plant raw materials and food products from the Ukrainian region were investigated. When determining of the proportion of isolated bacilli from the plant samples, it was established that the epidemiologically significant microorganisms of Bacillus cereus as agents of food poisoning are the second largest. The average value of contaminated samples of Ukrainian plant raw materials and processed products with Bacillus cereus is 36,2 %. The ability of Bacillus cereus strains identified by a complex of morphological, tinctorial, cultural and biochemical properties, to produce specific emetic and enterotoxins was studied. Molecular genetic diagnosis and detection of the toxin-producing ability of isolated 42 Bacillus cereus strains showed both the possibility of their rapid identification and the presence of specific toxicity genes. Multiplex polymerase chain reaction (PCR was carried out with specific primers to detect toxicity determined of various bacilli genes: nheA, hblD, cytK, cesВ. The distribution of toxigenic genes is significantly different among the Bacillus cereus isolates from various sources. The nheA, hblD and cytK enterotoxin genes were detected in 100, 83,3 and 61,9 % of the investigated strains of Bacillus cereus, respectively. The cesB gene encoding emetic toxin was detected in 4,8 % of  strains. Molecular-genetic PCR-method confirmed that all the isolated strains belong to the Bacillus cereus group, and the ability to produce toxins can be attributed to five groups. The main toxins that produce the investigated Bacillus cereus strains were nhe and hbl enterotoxins encoded by the corresponding genes of nheA and hblD. The enterotoxic type of Bacillus cereus was predominant in Ukrainian region.  Studies of domestic plant food raw materials and products have confirmed the need to improve microbiological control of product safety

Effect of High Pressure and Heat on Bacterial Toxins

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Dirk Margosch

2005-01-01

Full Text Available Even though the inactivation of microorganisms by high pressure treatment is a subject of intense investigations, the effect of high pressure on bacterial toxins has not been studied so far. In this study, the influence of combined pressure/temperature treatment (0.1 to 800 MPa and 5 to 121 °C on bacterial enterotoxins was determined. Therefore, heat-stable enterotoxin (STa of cholera toxin (CT from Vibrio cholerae, staphylococcal enterotoxins A-E, haemolysin BL (HBL from Bacillus cereus, and Escherichia coli (STa were subjected to different treatment schemes. Structural alterations were monitored in enzyme immunoassays (EIAs. Cytotoxicity of the pressure treated supernatant of toxigenic B. cereus DSM 4384 was investigated with Vero cells. High pressure of 200 to 800 MPa at 5 °C leads to a slight increase of the reactivity of the STa of E. coli. However, reactivity decreased at 800 MPa and 80 °C to (66±21 % after 30 min and to (44±0.3 % after 128 min. At ambient pressure no decrease in EIA reactivity could be observed after 128 min. Pressurization (0.1 to 800 MPa of heat stable monomeric staphylococcal toxins at 5 and 20 °C showed no effect. A combined heat (80 °C and pressure (0.1 to 800 MPa treatment lead to a decrease in the immuno-reactivity to 20 % of its maximum. For cholera toxin a significant loss in latex agglutination was observable only at 80 °C and 800 MPa for holding times higher than 20 min. Interestingly, the immuno-reactivity of B. cereus HBL toxin increased with the increase of pressure (182 % at 800 MPa, 30 °C, and high pressure showed only minor effects on cytotoxicity to Vero cells. Our results indicate that pressurization can increase inactivation observed by heat treatment, and combined treatments may be effective at lower temperatures and/or shorter incubation time.

Genotypic and Phenotypic Characterization of Enterotoxigenic Escherichia coli Strains Isolated from Peruvian Children

Science.gov (United States)

2010-09-01

adhesion, ETEC elabo- rates one or both of two enterotoxins: heat-labile toxin (LT), a protein multimer which shares many features with cholera toxin...pathogens (Shigella, Salmonella. Vibrio , Campylo/Jacter, Gi- ardia Iamblia, Cryptosporidium, and rotavirus) by conventional methods. Five lactose...studies performed in developing coun- tries: 38% in children ɚ years of age in Nicaragua (19), 33% in children ə year of age in Mexico (6), and 18% in

Clostridium perfringens in London, July 2009: two weddings and an outbreak.

OpenAIRE

Eriksen, J.; Zenner, D.; Anderson, S. R.; Grant, K.; Kumar, D.

2010-01-01

: Food poisoning outbreaks caused by Clostridium perfringens enterotoxin occur occasionally in Europe but have become less common in recent years. This paper presents the microbiological and epidemiological results of a large C. perfringens outbreak occurring simultaneously at two weddings that used the same caterer. The outbreak involved several London locations and required coordination across multiple agencies. A case-control study (n=134) was carried out to analyse possible associations b...

Study of immunoglobulin G thin layers obtained by the Langmuir-Blodgett method: application to immunosensors.

Science.gov (United States)

Barraud, A; Perrot, H; Billard, V; Martelet, C; Therasse, J

1993-01-01

Nowadays, immunosensors play a leading part in the field of bioanalytical chemistry research. As with any biosensor, they need appropriate transducers and a suitable technique to immobilize the active biocomponents. In this study, two transduction modes were chosen: mass effects (quartz microbalance measurements) and geometric and dielectric effects (capacitance measurements). The Langmuir-Blodgett (LB) method appears to be quite suitable for generating biospecific surfaces. This work has focused on the detection of staphylococcal enterotoxin B, the corresponding antibody being immobilized at the surface of fatty acids by a variant of the LB method. The composition of the film and the nature of antibody-fatty acid interactions were studied by means of the two transducers mentioned above. FTIR (Fourier transform infra-red) spectroscopy and protein diagnostic assay. Influence of several parameters (pH, ionic strength, transfer pressure, antibody concentration in the subphase) was investigated. The immobilization rate reached its maximum when experimental conditions allowed optimal electrostatic interactions. In this case, the quartz crystal microbalance response, in air, reached 55 Hz per monolayer of immobilized immunoglobulin G and the equivalent capacitance variation, measured in liquid media, was around 300 pF cm-2. Activity of the biospecific LB films, when binding enterotoxin, was checked by the classical ELISA (enzyme immuno-linked assay) technique.

Safety Evaluation of the Coagulase-Negative Staphylococci Microbiota of Salami: Superantigenic Toxin Production and Antimicrobial Resistance

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Raquel Soares Casaes Nunes

2015-01-01

Full Text Available The risks of contracting staphylococci food poisoning by the consumption of improperly manufactured salami and the possibility of this food being reservoirs for antibiotic resistance were evaluated. Nineteen coagulase-negative staphylococci (CNS strains were found in commercial and artisanal salami. The species in commercial salami were S. saprophyticus, S. sciuri, S. xylosus, and S. carnosus. Artisanal salami showed S. succinus, S. epidermidis, and S. hominis but no S. carnosus. Phylogenetic analyses grouped the strains into three major staphylococcal species groups, comprised of 4 refined clusters with similarities superior to 90%. Fifteen strains harbored multiple enterotoxin genes, with high incidence of seb/sec and sea, 57% and 50%, respectively, intermediate incidence of sed/seh/selm and sei/seln/tst-H, 33% and 27%, correspondingly, and low incidence of see/selj/selo and seg, of respectively 13% and 1%. Real time RT-PCR and enzyme-linked-immunosorbent assays confirmed the enterotoxigenicity of the strains, which expressed and produced enterotoxins in vitro. The CNS strains showed multiresistance to several antimicrobials of therapeutic importance in both human and veterinarian medicine, such as β-lactams, vancomycin, and linezolid. The effective control of undue staphylococci in fermented meat products should be adopted to prevent or limit the risk of food poisoning and the spread of antimicrobial-resistant strains.

Prevalence of Staphylococcus aureus Isolated From Various Foods of Animal Origin in Kırşehir, Turkey and Their Enterotoxigenicity

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M. Dilek Avşaroğlu

2016-12-01

Full Text Available The aim of this study was to detect Staphylococcus aureus contamination to different types of animal origin foods collected in the Kırşehir province of Turkey and to examine their enterotoxin production ability. Out of 120 food samples 38 suspected colonies were obtained and 23 of them were identified as S. aureus by biochemical and molecular analyses. Other species detected were S. chromogenes, S. cohnii ssp. cohnii, S. hominis, S. lentus, S. warneri, and S. xylosus. The isolates were also analysed with regard to carry mecA gene. None of them was found to have mecA gene indicating susceptibility to methicillin. To determine the enterotoxigenic ability of the isolates phenotypically, reversed-passive-latex-agglutination test against SEA-SED was used. Six out of 23 S. aureus isolates were determined to produce SEA, SEC and SED. Three of them had only one enterotoxin production, whereas others had SEA and SED production together. The results of phenotypic analyses were confirmed by PCR based examination. None of the coagulase-negative staphylococci were found to be enterotoxigenic by both phenotypical and PCR-based analyses. In conclusion, enterotoxigenic S. aureus is a risk in foods of animal origin in Kırşehir and its counties.

Putative new heat-stable cytotoxic and enterotoxic factors in culture supernatant of Escherichia coli isolated from drinking water

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DA Ribeiro

2011-01-01

Full Text Available Enteric infections caused by the ingestion of contaminated water, especially by Escherichia coli, are important to define the virulence properties of these bacteria. Due to frequent infantile diarrhea in the city of Ouro Preto, Minas Gerais state, Brazil, the phenotypic and genotypic diarrheagenic properties of E. coli isolated from drinking water were studied. The culture supernatants of 39 (40% among a total of 97 E. coli isolates from drinking water were positive by suckling mouse assay and induced cytotoxic effects on Vero cells. The enterotoxic and cytotoxic activities were present in the fraction with less than 10 kDa and were not lost when heated up to 60°C and 100°C for 30 minutes. PCR assays showed that among these 39 Vero cytotoxigenic E. coli, four (10.2% were positive for ST II (estB and two (5% positive for αHly (hlyA. Gene amplification of SLT (stx 1, stx 2, ST I (estA, LT (eltI, eltII, EAST1 (astA, EHly (enhly and plasmid-encoded enterotoxin (pet were not observed. This heat-stable cytotoxic enterotoxin of E. coli is probably a new putative diarrheagenic virulence factor, as a toxin presenting these characteristics has not yet been described.

Oral Administration of Astrovirus Capsid Protein Is Sufficient To Induce Acute Diarrhea In Vivo

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Victoria A. Meliopoulos

2016-11-01

Full Text Available The disease mechanisms associated with the onset of astrovirus diarrhea are unknown. Unlike other enteric virus infections, astrovirus infection is not associated with an inflammatory response or cellular damage. In vitro studies in differentiated Caco-2 cells demonstrated that human astrovirus serotype 1 (HAstV-1 capsid protein alone disrupts the actin cytoskeleton and tight junction complex, leading to increased epithelial barrier permeability. In this study, we show that oral administration of purified recombinant turkey astrovirus 2 (TAstV-2 capsid protein results in acute diarrhea in a dose- and time-dependent manner in turkey poults. Similarly to that induced by infectious virus, TAstV-2 capsid-induced diarrhea was independent of inflammation or histological changes but was associated with increased intestinal barrier permeability, as well as redistribution of sodium hydrogen exchanger 3 (NHE3 from the membrane to the cytoplasm of the intestinal epithelium. Unlike other viral enterotoxins that have been identified, astrovirus capsid induces diarrhea after oral administration, reproducing the natural route of infection and demonstrating that ingestion of intact noninfectious capsid protein may be sufficient to provoke acute diarrhea. Based on these data, we hypothesize that the astrovirus capsid acts like an enterotoxin and induces intestinal epithelial barrier dysfunction.

Modeling the level of contamination of Staphylococcus aureus in ready-to-eat kimbab in Korea.

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Bahk, Gyung-Jin; Hong, Chong-Hae; Oh, Deog-Hwan; Ha, Sang-Do; Park, Ki-Hwan; Todd, Ewen C D

2006-06-01

The risk of Staphylococcus aureus in ready-to-eat kimbab (rice rolled in laver) sold in Korea was evaluated by a mathematical modeling approach. Four nodes were constructed from preparation at retail to consumption. A predictive microbial growth model and survey data were combined with probabilistic modeling to simulate the level of S. aureus in a single kimbab at the time of consumption. We estimated the mean level of S. aureus to be 2.92 log CFU/g for a typical kimbab (150 to 200 g each) at the time of consumption. Our model also showed that 29.73% of the kimbabs had > or = 100,000 S. aureus CFU/g, which poses some risk of illness, since some level of enterotoxin would be expected from toxigenic strains. However, because of the lack of dose-response models for staphylococcal enterotoxin, the final level of S. aureus in the kimbabs could not be used to estimate how many people would become ill from eating them. Correlation sensitivity results showed that consumer eating patterns and initial contamination levels at retail stores were the most significant risk factors for illness and that temperature control under 10 degrees C was a critical control point in kimbab retail establishments to prevent the growth of S. aureus.

Poor survival of Methicillin-resistant Staphylococcus aureus on inanimate objects in the public spaces.

Science.gov (United States)

Domon, Hisanori; Uehara, Yoshio; Oda, Masataka; Seo, Hiromi; Kubota, Noriko; Terao, Yutaka

2016-02-01

We investigated the prevalence of Staphylococcus aureus on shopping baskets in Osaka Prefecture, Japan. Multilocus sequence typing was performed to determine the genotypes of S. aureus isolates, and then a polymerase chain reaction method was used to detect staphylococcal enterotoxins and antibiotic resistance genes. In addition, desiccation tolerance of S. aureus isolates was evaluated in vitro. Forty-six (6.2%) S. aureus isolates were collected from 740 shopping baskets, though only one MRSA strain was identified. In multilocus sequence typing findings, ten sequence types and 24 singletons were classified, which were divided into ten clonal complexes and six singletons. The most frequent staphylococcal enterotoxin gene was seg (30.4%). Our in vitro findings demonstrated that 70% of the S. aureus isolates, including the MRSA strain, became undetectable at 12 h after desiccation at an appropriate cell density, while the others remained viable for up to 24 h. Thus, it is difficult for MRSA organisms to survive on dry surfaces found in public areas. We speculated that inanimate objects in the community are unlikely to be a potential source for transmission of MRSA and that S. aureus on such objects outside of hospital settings is not a public health threat. © 2015 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.

Balneotherapy of Psoriasis

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Golušin Zoran

2014-09-01

Full Text Available Application of different kinds of mineral waters and peloids on the skin exerts mechanical, thermal and chemical effects. Significant reduction of inflammation and increased differentiation of keratinocytes may explain why balneotherapy has positive clinical effects in psoriatic patients. In vitro models have shown that thermal water stimulates interleukin-2 production after cell stimulation by staphylococcal enterotoxin B, and reduces interleukin-4 secretion. After balneotherapy, a significant decrease in Psoriasis Area Severity Index (PASI, associated with a significant reduction of interleukin-8, Staphylococcus aureus colonization and enterotoxin N, have been reported in patients with psoriasis. Mineral water was found to have inhibitory in vitro effects on substance P, TNF-α release and antigen-induced cell degranulation. Immunomodulatory effects of water depend on its content. Sulfur waters have beneficial anti-inflammatory, keratolytic, and antipruriginous effects and also possess antibacterial and antifungal properties. The effectiveness of balneotherapy in the treatment of psoriasis has been reported in many studies conducted all over the world. The majority of studies were conducted at the Dead Sea coast. Investigations showed that balneotherapy factors are important therapeutic factors in the treatment of psoriatic patients. The first and only comparable study of this kind in Serbia, was conducted in Prolom Spa with satisfactory therapeutic results.

Isolation and identification of Staphylococcus sp. in powdered infant milk

Science.gov (United States)

Palilu, Prayolga Toban; Budiarso, Tri Yahya

2017-05-01

Staphylococcus sp. is one of the most dangerous bacteria that could cause food poisoning. It is a pathogenic bacterium which is able to produce enterotoxin in foods. Milk is an ideal growth medium for Staphylococcus sp., that may cause problem if it is to be consumed, especially by infant. It is the objective of this research to detect the presence of Staphylococcus sp. in powdered infant milk. As many as 14 samples obtained from market were used as samples for bacterial isolation. The isolation were done by employing enrichment step on BHI-broth, continued with Baird-Parker Agar which will produce a typical colony. It is then picked and grown on Mannitol Salt Agar, and gram staining, coagulase assay, and fermentation tests. The confirmation step was done by using API-Staph which gives the identification of Staphylococcus hemoliticus, Staphylococcus aureus and Staphylococcus epidermidis, with a percentage of identity ranging from 65.9-97.7%. Two isolates with the highest identification similarity values were then picked for molecular detection. A PCR primer pair targeting gene coding for enterotoxin A was used, and it gives positive result for the two isolates being tested. It is then concluded that the two isolates belong to Staphylococcus sp., and further research need to be done to correctly identify these isolates.

Presença de Staphylococcus spp. produtores de enterotoxinas e da toxina da síndrome do choque tóxico em manipuladores de queijo de cabra Presence of Staphylococcus strains producer of enterotoxins and toxic shock toxin syndrome isolated from goat's cheese handlers

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L.S. Rapini

2005-12-01

Full Text Available Agruparam-se 167 cepas de Staphylococcus, isoladas de diferentes sítios (fossas nasais, orofaringe, palma das mãos e leitos subungueais, de dois manipuladores de queijos de leite de cabra, em pools (45, de acordo com a similaridade da espécie e o local de origem, e testou-se o seu potencial de produção de enterotoxinas (SE A, B, C e D, e da toxina da síndrome do choque tóxico (TSST-1. Constatou-se que 62,2% dos pools foram capazes de produzir, de forma individual ou associada, SEA (33,3%; SEB (46,7%; SEC (8,9%; SED (4,4% e TSST-1 (4,4%. Dos pools enterotoxigênicos, 96,4% corresponderam a espécies coagulase negativa (Staphylococcus epidermidis e Staphylococcus cohnii. A capacidade de produção de SE e de TSST-1 pelas cepas de Staphylococcus spp. isoladas dos manipuladores, além de reafirmar sua importância na transmissão de intoxicações de origem alimentar, enfatiza a necessidade imprescindível da adoção de condutas higiênicas e sanitárias durante a manipulação do alimento.A total of 167 strains of Staphylococcus isolated from nasal cavities, oropharynx, palm of hands and subunguial of two goat's cheese handlers were collected. The strains were pooled (45 according to the species similarity and place of origin and tested for the production of enterotoxins (SE A, B, C, D and toxic shock toxin syndrome (TSST-1. It was observed that 62.2% of the pools presented the capacity to produce, individually or in association, SEA (33.3%; SEB (46.7%; SEC (8.9%; SED (4.4% and TSST-1 (4.4%. From the enterotoxigenic pools, 96.4% corresponded to species negative coagulase (Staphylococcus epidermidis and Staphylococcus cohnii. The capacity to produce SE and TSST-1 by Staphylococcus spp. strains isolated from food handlers reaffirms its hole on the transmission of food poisoning, beyond emphasizing the indispensable necessity of the adoption of satisfactory hygienic and sanitary procedures during the food manufacture.

Molecular characterization and antimicrobial susceptibility of Staphylococcus aureus from small-scale dairy systems in the highlands of Central México

OpenAIRE

Salgado-Ruiz, T. B.; Rodríguez González, Ana; Gutiérrez, Diana; Martínez Fernández, Beatriz; García Suárez, María Pilar; Arriaga-Jordán, C. M.

2015-01-01

Staphylococcus aureus is involved in human and animal infections. Because of mastitis in dairy cows, milk can be contaminated by enterotoxin-producing strains, which constitutes a food poisoning risk. Animal handlers can be asymptomatic carriers, becoming an additional source of contamination. This research aims to improve our understanding of Staphylococcus aureus in small-scale dairy systems in central Mexico. Samples were taken in 12 dairy farms and included composite milk (from the four t...

Bacillus cereus and related species.

OpenAIRE

Drobniewski, F A

1993-01-01

Bacillus cereus is a gram-positive aerobic or facultatively anaerobic spore-forming rod. It is a cause of food poisoning, which is frequently associated with the consumption of rice-based dishes. The organism produces an emetic or diarrheal syndrome induced by an emetic toxin and enterotoxin, respectively. Other toxins are produced during growth, including phospholipases, proteases, and hemolysins, one of which, cereolysin, is a thiol-activated hemolysin. These toxins may contribute to the pa...

Evaluación de estrategias de control frente a biopelículas de "Staphylococcus aureus" potencialmente presentes en la industria pesquera

OpenAIRE

Vázquez Sánchez, Daniel

2014-01-01

Staphylococcus aureus is one of the major bacterial agents causing foodborne diseases in humans worldwide, due to the ingestion of food containing staphylococcal enterotoxins. Spain is one of the largest producers and consumers of fishery products in the European Union. However, S. aureus is repeatedly detected in these products as a consequence of cross-contamination from food handlers and food contact surfaces. Biofilm formation also provides S. aureus a high tolerance to biocides allowing ...

Enhancement of antibiotic activity by Cordia verbenacea DC

OpenAIRE

Matias, Edinardo F.F.; Santos, Karla K. A.; Almeida, Thiago S.; Costa, José G.M. da; Coutinho, Henrique D.M.

2010-01-01

Escherichia coli is known to produce enterotoxins whose properties and its role in diarrheal disease has been extensively investigated. Some species of Staphylococcus are often recognized as etiological agents of many animal and human opportunistic infections. This study is the first test of change in resistance of antibiotic activity by Cordia verbenacea DC. against multiresistant strains of Escherichia coli and Staphylococcus aureus. In this study, the hexane and methanol extract of Cordia ...

Species Distribution and Prevalence of Putative Virulence Factors in Mesophilic Aeromonas spp. Isolated from Fresh Retail Sushi

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Sunniva Hoel

2017-05-01

Full Text Available Aeromonas spp. are ubiquitous bacteria that have received increasing attention as human pathogens because of their widespread occurrence in food, especially seafood and vegetables. The aim of this work was to assess the species identity and phylogenetic relationship of 118 Aeromonas strains isolated from fresh retail sushi from three producers, and to characterize the isolates with respect to genetic and phenotypic virulence factors. We also evaluate the potential hazard associated with their presence in ready-to-eat seafood not subjected to heat treatment. Mesophilic Aeromonas salmonicida was most prevalent (74%, followed by A. bestiarum (9%, A. dhakensis (5%, A. caviae (5%, A. media (4%, A. hydrophila (2%, and A. piscicola (1%. All isolates were considered potentially pathogenic due to the high prevalence of genes encoding hemolysin (hlyA (99%, aerolysin (aerA (98%, cytotoxic enterotoxin (act (86%, heat-labile cytotonic enterotoxin (alt (99%, and heat-stable cytotonic enterotoxin (ast (31%. The shiga-like toxins 1 and 2 (stx-1 and stx-2 were not detected. Moreover, there was heterogeneity in toxin gene distribution among the isolates, and the combination of act/alt/hlyA/aerA was most commonly detected (63%. β-hemolysis was species-dependent and observed in 91% of the isolates. All A. media and A. caviae strains were non-hemolytic. For isolates belonging to this group, lack of hemolysis was possibly related to the absence of the act gene. Swimming motility, linked to adhesion and host invasion, occurred in 65% of the isolates. Partial sequencing of the gyrB gene demonstrated its suitability as a genetic marker for Aeromonas species identification and for assessment of the phylogenetic relationship between the isolates. The gyrB sequence divergence within a given species ranged from 1.3 to 2.9%. A. bestiarum, A. salmonicida, and A. piscicola were the most closely related species; their sequences differed by 2.7–3.4%. The average gyrB sequence

Detection of the enterotoxigenic genes (sei,sej) in Staphylococcus aureus isolates from bovine mastitis milk in the West Azerbaijan of Iran

OpenAIRE

Ahmady, Malahat; Kazemi, Sahar

2012-01-01

Staphylococcus aureus is a major causative pathogen of clinical and subclinical mastitis of dairy domestic ruminants. This organism produces a variety of extracellular toxins and virulence factors such as enterotoxin SEI and SEJ that contribute to its pathogenic potential. In this study 25 S. aureus isolates obtained from four dairy herds of Urmia region which is located in West Azerbaijan province in Iran. The tested isolates were identified on the basis of the cultural and biochemical prope...

Use of the Real Time xCelligence System for Purposes of Medical Microbiology.

Science.gov (United States)

Junka, Adam Feliks; Janczura, Adriana; Smutnicka, Danuta; Mączyńska, Beata; Anna, Secewicz; Nowicka, Joanna; Bartoszewicz, Marzenna; Gościniak, Grażyna

2012-09-28

Roche's xCelligence impedance-measuring instrument is one of a few commercially available systems of such type. According to the best knowledge of authors, instrument was tested so far only for eukaryotic cell research. The aim of this work was to estimate xCELLigence suitability for the microbiological tests, including (i) measurement of morphological changes in eukaryotic cells as a result of bacterial toxin activity, (ii) measurement of bacterial biofilm formation and (iii) impact of antiseptics on the biofilm structure. To test the infuence of bacterial LT enterotoxin on eukaryotic cell lines, Chinese Hamster Ovary (CHO) cell line and reference strain Escherichia coli ATTC 35401 were used. To investigate Roche's instrument ability to measure biofilm formation and impact of antiseptics on its development, Staphylococcus aureus ATTC6538 reference strain was used. The data generated during the experiments indicate excellent ability of xCelligence instrument to detect cytopathic effect caused by bacterial LT endotoxin and to detect staphylococcal biofilm formation. However, interpretation of the results obtained during real-time measurement of antiseptic's bactericidal activity against staphylococcal biofilm, caused many difficulties. xCelligence instrument can be used for real-time monitoring of morphological changes in CHO cells treated with bacterial LT enterotoxin and for real-time measurement of staphylococcal biofilm formation in vitro. Further investigation is necessary to confirm suitability of system to analyze antiseptic's antimicrobial activity against biofilm in vitro.

EPIDEMIOLOGIA MOLECULAR APLICADA AO MONITORAMENTO DE ESTIRPES DE Staphylococcus aureus NA PRODUÇÃO DE QUEIJO MINAS FRESCAL

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Maria Izabel Merino de Medeiros

2013-03-01

Full Text Available We studied the molecular epidemiology of Staphylococcus aureus strains potentially toxigenic, isolated from the production process of Minas frescal cheese in a small dairy plant in the state of São Paulo. For this, samples were taken during the period from June 2008 to July 2009. Samples were collected from the surface of the receiving and storage tanks of raw milk, the surface of the balance tank of pasteurized milk, the water supply system, the pipes and equipments, the hands of the handler and from the packaged cheese, totaling 140 samples. The colonies isolated on Baird-Parker Agar confirmed as Gram positive and positive for catalase, coagulase and acetoin production, were submitted to extraction of bacterial DNA using the Invitek - Uniscience® kit. Confirmation of the isolated species and enterotoxins SEA, SEB, SEC, SED and TSST-1 toxin was carried out through the amplification of specific fragments of chromosomal DNA. Among the 74 strains of isolated coagulase-positive staphylococci, only 41 (55.4% strains were confirmed as Staphylococcus aureus, of which 25 (61.0% were positive to the presence of staphylococcal toxins. The most frequently identified enterotoxin was SEA. The toxigenic strains of Staphylococcus aureus were more frequently isolated from hands of the handler (16.0%, raw milk receiving tank (12.0%, pasteurized milk for cheese making (12.0% and fresh white cheese ready for consumption (12.0%.

Effect of packaging conditions on the growth of micro-organisms and the quality characteristics of fresh mushrooms (Agaricus bisporus) stored at inadequate temperatures.

Science.gov (United States)

González-Fandos, E; Giménez, M; Olarte, C; Sanz, S; Simón, A

2000-10-01

Mushrooms were packed in two polymeric films (perforated and non-perforated PVC) and stored at 17 degrees C and 25 degrees C. The carbon dioxide and oxygen content inside the packages, aerobic mesophiles, Pseudomonas spp., faecal coliforms, Escherichia coli, anaerobic spores and major sensory factors (colour, texture, development stage and presence of moulds) were determined. The non-perforated packages had the highest contents of CO2 (6-7%), the lowest contents of O2 (0.013-0.17%) and the most desirable quality parameters (texture, development stage and absence of moulds). Pseudomonas spp. counts were around 1 logarithmic unit lower in mushrooms packaged in non-perforated film as the O2 concentrations were lower than in perforated film. The mushrooms themselves were inoculated with an enterotoxin A-producing strain of Staphylococcus aureus, packaged in overwrapped trays and stored at 17 and 25 degrees C. Staphylococcus aureus did not grow in the samples stored at 17 degrees C. Only slight growth was observed in mushrooms packaged with non-perforated film after 1 day at 25 degrees C. No enterotoxin was detected in any package. Faecal coliform counts were <2 log cfu g(-1). Escherichia coli was not isolated in any of the samples. At 25 degrees C, counts of anaerobic spores of around 2 log cfu g(-1) were detected in those mushrooms packaged in non-perforated film.

Investigating an outbreak of staphylococcal food poisoning among travellers across two Australian states.

Science.gov (United States)

Fletcher, Stephanie; Boonwaat, Leng; Moore, Terry; Chavada, Ruchir; Conaty, Stephen

2015-01-01

Staphylococcus aureus is a common cause of staphylococcal food poisoning in Australia with several outbreaks associated with foods prepared by commercial caterers. Laboratory testing on cases of gastrointestinal illness caused by enterotoxin-producing S. aureus is not routinely done as this condition is self-limiting. Hence outbreaks of such illness may go undetected. A retrospective cohort study was conducted among a group of tourists who were hospitalized in Sydney shortly after flying from Queensland. The group had consumed food prepared by a restaurant on the Gold Coast before transit. Laboratory analyses on stool specimens were conducted in Sydney. An environmental assessment of the restaurant in the Gold Coast was conducted, and environmental specimens were assessed for contamination. Epidemiological investigations linked the outbreak to a restaurant in the Gold Coast where the suspected food was produced. Stool samples from two of the hospitalized cases were confirmed to have enterotoxin-producing S. aureus, and several environmental samples were found to be contaminated with S. aureus as well. Investigations suggested that absence of hand washing and other unhygienic food handling at the implicated restaurant was the likely cause of this outbreak. Food poisoning due to toxin-mediated S. aureus is frequently undetected and underreported. Public health units should consider toxin-producing pathogens such as S. aureus when investigating outbreaks where vomiting is the predominant symptom and occurs rapidly after consuming food.

A Chimeric protein of CFA/I, CS6 subunits and LTB/STa toxoid protects immunized mice against enterotoxigenic Escherichia coli.

Science.gov (United States)

Zeinalzadeh, Narges; Salmanian, Ali Hatef; Goujani, Goli; Amani, Jafar; Ahangari, Ghasem; Akhavian, Asal; Jafari, Mahyat

2017-07-01

Enterotoxigenic Escherichia Coli (ETEC) strains are the commonest bacteria causing diarrhea in children in developing countries and travelers to these areas. Colonization factors (CFs) and enterotoxins are the main virulence determinants in ETEC pathogenesis. Heterogeneity of CFs is commonly considered the bottleneck to developing an effective vaccine. It is believed that broad spectrum protection against ETEC would be achieved by induced anti-CF and anti-enterotoxin immunity simultaneously. Here, a fusion antigen strategy was used to construct a quadrivalent recombinant protein called 3CL and composed of CfaB, a structural subunit of CFA/I, and CS6 structural subunits, LTB and STa toxoid of ETEC. Its anti-CF and antitoxin immunogenicity was then assessed. To achieve high-level expression, the 3CL gene was synthesized using E. coli codon bias. Female BALB/C mice were immunized with purified recombinant 3CL. Immunized mice developed antibodies that were capable of detecting each recombinant subunit in addition to native CS6 protein and also protected the mice against ETEC challenge. Moreover, sera from immunized mice also neutralized STa toxin in a suckling mouse assay. These results indicate that 3CL can induce anti-CF and neutralizing antitoxin antibodies along with introducing CFA/I as a platform for epitope insertion. © 2017 The Societies and John Wiley & Sons Australia, Ltd.

SANITARY EVALUATION OF MILK PRODUCTS IN MOUNTAIN DAIRIES

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R. Mioni

2010-03-01

Full Text Available between 2006 and 2007 881 samples of “malga” (little mountain dairies milk products were analysed to estimate their hygienic characteristics. Several samples showed high counts for Escherichia coli and coagulase-positive staphylococci, while Salmonella spp. and Campylobacter spp. were absent in all of the samples; 0,9% of cheese samples, 4,1% of butter samples and 4,7% of “ricotta” samples were positive for Listeria monocytogenes, so as 14,7% of cheese samples for staphylococcal enterotoxins.

The application of alkaline and acidic electrolyzed water in the sterilization of chicken breasts and beef liver

OpenAIRE

Shimamura, Yuko; Shinke, Momoka; Hiraishi, Miki; Tsuchiya, Yusuke; Masuda, Shuichi

2015-01-01

Abstract The sterilization effect of a combination treatment with alkaline electrolyzed water (AlEW) and strong acidic electrolyzed water (StAEW) on fresh chicken breasts and beef liver was evaluated. Samples (1, 5, and 10?g) were inoculated with Salmonella Enteritidis NBRC3313, Escherichia coli ATCC 10798, Staphylococcus aureus FDA209P, and S.?aureus C?29 [staphylococcal enterotoxin A (SEA) productive strain] and subjected to a dipping combination treatment (4?C and 25?C for 3?min) with AlEW...

Effect of atopic skin stressors on natural moisturizing factors and cytokines in healthy adult epidermis

DEFF Research Database (Denmark)

Engebretsen, K A; Kezic, S; Jakasa, I

2018-01-01

: To explore the effect of selected exogenous skin stressors on NMF and skin cytokines levels in healthy adult epidermis. MATERIAL AND METHODS: 40 healthy volunteers (18-49 years) were exposed to hard, soft, and chlorinated water, 0.5% SLS, house dust mite, cat allergen, staphylococcal enterotoxin B (SEB...... of various skin cytokines in healthy individuals. Our data highlight environmental factors that might play a role in AD pathophysiology, but needs confirmation in AD patients. This article is protected by copyright. All rights reserved....

Investigating an outbreak of staphylococcal food poisoning among travellers across two Australian states

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Stephanie Fletcher

2015-05-01

Full Text Available Introduction: Staphylococcus aureus is a common cause of staphylococcal food poisoning in Australia with several outbreaks associated with foods prepared by commercial caterers. Laboratory testing on cases of gastrointestinal illness caused by enterotoxin-producing S. aureus is not routinely done as this condition is self-limiting. Hence outbreaks of such illness may go undetected. Methods: A retrospective cohort study was conducted among a group of tourists who were hospitalized in Sydney shortly after flying from Queensland. The group had consumed food prepared by a restaurant on the Gold Coast before transit. Laboratory analyses on stool specimens were conducted in Sydney. An environmental assessment of the restaurant in the Gold Coast was conducted, and environmental specimens were assessed for contamination. Results: Epidemiological investigations linked the outbreak to a restaurant in the Gold Coast where the suspected food was produced. Stool samples from two of the hospitalized cases were confirmed to have enterotoxin-producing S. aureus, and several environmental samples were found to be contaminated with S. aureus as well. Investigations suggested that absence of hand washing and other unhygienic food handling at the implicated restaurant was the likely cause of this outbreak. Conclusion: Food poisoning due to toxin-mediated S. aureus is frequently undetected and underreported. Public health units should consider toxin-producing pathogens such as S. aureus when investigating outbreaks where vomiting is the predominant symptom and occurs rapidly after consuming food.

Prevalence of enteropathogens and their antibiotic sensitivity pattern in puppies with hemorrhagic gastroenteritis

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A. Kokila Priya

2017-08-01

Full Text Available Aim: Hemorrhagic gastroenteritis (HGE ranging from mild to severe forms is commonly encountered in puppies. The aim of the study was to identify the prevalence of common enteropathogens and the antibiotic sensitivity pattern in puppies reported with HGE. Materials and Methods: The canine HGE activity index, with little modification, was adopted to identify Grade III/ severely affected puppies below 6 months of age. Fecal polymerase chain reaction (PCR assay was employed to screen and compare the enteropathogens in puppies with hemorrhagic diarrhea and healthy control. Results: Canine parvovirus 2b was identified in 90.3% of the diarrheic and 10% of the non-diarrheic healthy puppies. Clostridium difficile was identified in all the diarrheic puppies and in 80% of the healthy puppies. Among the diarrheic puppies, 17.7% were positive for Clostridium perfringens enterotoxin, 9.7% were positive for C. perfringens alpha toxin, 6.4% were positive for Escherichia coli shiga toxin, 6.4% were positive for E. coli enterotoxin (LT, and 3.2% were positive for canine distemper virus. Whereas, none of the healthy puppies were positive for these bacteria and toxins. Fecal antibiotic sensitivity test pattern revealed gentamicin to be sensitive in 95% of the cases, azithromycin in 50%, enrofloxacin in 25%, cefotaxime in 20%, and tetracycline in 5% of the cases. Conclusion: Parvoviral enteritis is predominant among puppies. Yet, bacteria and their toxins also play an important role in HGE. Gentamicin has higher sensitivity against the enteropathogens associated with the condition.

GENOTYPING OF CLOSTRIDIUM PERFRINGENS FROM FRESH WATER FISH AND FISH PICKLES

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Adarsh Jain

2012-08-01

Full Text Available This study aims to evaluate the genotypes of Clostridium perfringens in fish and fish based products from Tamil Nadu and Kerala states of India. A total of 301 samples consisting intestinal contents of freshwater fish (234 from various dams, freshwater lakes, ponds, retail shops and markets and fish pickles (67 obtained from randomly selected retail shops and supermarkets were investigated. Bacterial isolations, identifications and phenotypic characterization of virulence factors were carried out as per standard microbiological procedures. Genotyping of the C. perfringens isolates were done by amplifying four major lethal toxin genes namely- alpha toxin gene (cpa, beta toxin gene (cpb, epsilon toxin gene (etx, iota toxin gene (iA in a Thermal Cycler. Isolates were also screened for the presence of enterotoxin gene (cpe and beta2 toxin gene (cpb2 by single step PCR. Biochemical tests and phenotypic determination of virulence factors tentatively identified 82 (27.24% isolates of C. perfringens. In PCR assay, all 82 (100% isolates harbored cpa toxin genes of C. perfringens, however, 65 (79.26% isolates also carried additional cpb2 toxin genes. None of the isolates were found positive for beta, epsilon, iota and enterotoxin genes. Genotyping of the 82 isolates by PCR revealed that all the isolated bacteria were belonged to C. perfringens type A and both cpa and cpb2 toxin genes were prevalent among the isolates of C. perfringens type A, impending the risk of pathogenicity to human via freshwater fish and fish pickles.

Control of Virulence Gene Expression by the Master Regulator, CfaD, in the Prototypical Enterotoxigenic Escherichia coli Strain, H10407

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Carla Hodson

2017-08-01

Full Text Available Enterotoxigenic Escherichia coli (ETEC is the most common bacterial cause of diarrhea in children in developing countries, as well as in travelers to these countries. To cause disease, ETEC needs to produce a series of virulence proteins including enterotoxins, colonization factors and secretion pathways, which enable this pathogen to colonize the human small intestine and deliver enterotoxins to epithelial cells. Previously, a number of studies have demonstrated that CfaD, an AraC-like transcriptional regulator, plays a key role in virulence gene expression by ETEC. In this study, we carried out a transcriptomic analysis of ETEC strain, H10407, grown under different conditions, and determined the complete set of genes that are regulated by CfaD. In this way, we identified a number of new target genes, including rnr-1, rnr-2, etpBAC, agn43, flu, traM and ETEC_3214, whose expression is strongly activated by CfaD. Using promoter-lacZ reporters, primer extension and electrophoretic mobility shift assays, we characterized the CfaD-mediated activation of several selected target promoters. We also showed that the gut-associated environmental signal, sodium bicarbonate, stimulates CfaD-mediated upregulation of its virulence target operons. Finally, we screened a commercial small molecule library and identified a compound (CH-1 that specifically inhibited the regulatory function of CfaD, and by 2-D analoging, we identified a second inhibitor (CH-2 with greater potency.

Partial Sequencing of 16S rRNA Gene of Selected Staphylococcus aureus Isolates and its Antibiotic Resistance

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Harsi Dewantari Kusumaningrum

2016-08-01

Full Text Available The choice of primer used in 16S rRNA sequencing for identification of Staphylococcus species found in food is important. This study aimed to characterize Staphylococcus aureus isolates by partial sequencing based on 16S rRNA gene employing primers 16sF, 63F or 1387R. The isolates were isolated from milk, egg dishes and chicken dishes and selected based on the presence of sea gene that responsible for formation of enterotoxin-A. Antibiotic susceptibility of the isolates towards six antibiotics was also tested. The use of 16sF resulted generally in higher identity percentage and query coverage compared to the sequencing by 63F or 1387R. BLAST results of all isolates, sequenced by 16sF, showed 99% homology to complete genome of four S. aureus strains, with different characteristics on enterotoxin production and antibiotic resistance. Considering that all isolates were carrying sea gene, indicated by the occurence of 120 bp amplicon after PCR amplification using primer SEA1/SEA2,  the isolates were most in agreeing to S. aureus subsp. aureus ST288. This study indicated that 4 out of 8 selected isolates were resistant towards streptomycin. The 16S rRNA gene sequencing using 16sF is useful for identification of S. aureus. However, additional analysis such as PCR employing specific gene target, should give a valuable supplementary information, when specific characteristic is expected.

Antimicrobial resistance levels amongst staphylococci isolated from clinical cases of bovine mastitis in Kosovo.

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Mehmeti, Ibrahim; Behluli, Behlul; Mestani, Mergim; Ademi, Arsim; Nes, Ingolf F; Diep, Dzung B

2016-10-31

Mastitis is one of the most frequent and costly disease in cattle. We studied milk samples from cattle with mastitis from farms in Kosovo to identify mastitis-causing pathogens and possible effective antibiotics. Our ultimate goal is to help implement adequate antibiotic management and treatment practices in Kosovo METHODOLOGY: A total of 152 milk samples were collected from cows with clinical mastitis from different farms in Kosovo. After identification of microorganisms, antibiotic susceptibility and the occurrence of enterotoxins was investigated. Staphylococci were found in 89 samples, of which 58 were coagulase negative and 31 coagulase positive. S. aureus was isolated from 27 samples, S. epidermidis from 25, and S. chromogenes from 15, while other species of staphylococci were isolated from the remaining 22 isolates. Interestingly, the bacterial diversity was different between cows in different periods of lactation and among different breeds. Most of the isolates (76/89) were resistant to two or more antibiotics. The highest resistance was to penicillin and ampicillin (> 65%), followed by tetracycline, oxacillin, streptomycin, chloramphenicol (> 23%), and less than 3% to erythromycin. Of the 89 isolates, 40 produced enterotoxins that were most frequently typed as A and C. We detected human bacterial pathogens in the cultures of milk samples from cows with mastitis. The isolates demonstrated resistance to two or more antibiotics, some of which are frequently used to treat animal and human infections. We recommend increased control and more stringent use of antibiotics in veterinary as well as human medicine.

Antibiotic resistance and molecular analysis of Staphylococcus aureus isolated from cow's milk and dairy products in northeast Brazil.

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Silveira-Filho, Vladimir M; Luz, Isabelle S; Campos, Ana Paula F; Silva, Wellington M; Barros, Maria Paloma S; Medeiros, Elizabeth S; Freitas, Manuela F L; Mota, Rinaldo A; Sena, Maria J; Leal-Balbino, Tereza C

2014-04-01

This work aimed to assess the clonal distribution among 94 strains of Staphylococcus aureus isolated from cow's milk, raw cheese, and a milking machine in 12 dairy farms in northeast Brazil, by analyzing different typing methods and detecting resistance and toxigenic profiles. For the first time, isolates of this region were assessed simultaneously by the polymorphism of the 3'-end coa gene and 16S-23S rDNA, pulsed-field gel electrophoresis, antibiotic resistance phenotyping, and toxigenic arsenal. Although pulsed-field gel electrophoresis patterns showed a wider variation (discriminatory index 0.83) than the PCR-based methods, the internal transcribed spacer-PCR proved to be a useful and inexpensive procedure for conducting epidemiological surveys of S. aureus on a regional scale. Each dairy farm had its own resistance profile, and in two herds, 63% of the strains were multiresistant, probably due to the indiscriminate use of antibiotics in bovine mastitis treatment. No methicillin-resistant S. aureus strains were detected in this study; however, 93.6% of S. aureus strains harbored variable profiles of staphylococcal enterotoxin genes seg, seh, sei, and sej. Transcriptional analysis revealed that 53.3% of staphylococcal enterotoxin genes actually transcribed, pointing out the food poisoning risk of these dairy products to consumers in the region. Based on the detection of the most prevalent clones in a herd or region, appropriate antibiotic therapy and specific immunization can be used for the treatment and control of staphylococcal mastitis.

Virulence factors genes of Staphylococcus spp. isolated from caprine subclinical mastitis.

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Salaberry, Sandra Renata Sampaio; Saidenberg, André Becker Simões; Zuniga, Eveline; Melville, Priscilla Anne; Santos, Franklin Gerônimo Bispo; Guimarães, Ednaldo Carvalho; Gregori, Fábio; Benites, Nilson Roberti

2015-08-01

The aim of this study was to investigate genes involved in adhesion expression, biofilm formation, and enterotoxin production in isolates of Staphylococcus spp. from goats with subclinical mastitis and associate these results with the staphylococcal species. One hundred and twenty-four isolates were identified and polymerase chain reaction (PCR) was performed to detect the following genes: cna, ebpS, eno, fib, fnbA, fnbB, bap, sea, seb, sec, sed and see. The most commonly Staphylococcus species included S. epidermidis, S. lugdunensis, S. chromogenes, S. capitis ss capitis and S. intermedius. With the exception of fnbB, the genes were detected in different frequencies of occurrence in 86.3% of the Staphylococcus spp. isolates. Eno (73.2%) and bap (94.8%) were more frequently detected in coagulase-negative staphylococci (CNS); ebpS (76%), fib (90.9%) and fnbA (87%) were the most frequent genes in coagulase-positive staphylococci (CPS). Regarding enterotoxins, genes sed (28.2%) and see (24.2%) had a higher frequency of occurrence; sec gene was more frequently detected in CPS (58.8%). There was no association between the presence of the genes and the Staphylococcus species. Different virulence factors genes can be detected in caprine subclinical mastitis caused by CNS and CPS. The knowledge of the occurrence of these virulence factors is important for the development of effective control and prevention measures of subclinical mastitis caused by CNS and CPS in goats. Copyright © 2015 Elsevier Ltd. All rights reserved.

Genospecies and virulence factors of Aeromonas species in different sources in a North African country

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Khalifa Sifaw Ghenghesh

2014-09-01

Full Text Available Introduction: Aeromonads of medical importance have been reported from numerous clinical, food, and water sources, but identification of genospecies and virulence factors of Aeromonas species from countries in North Africa and the Middle East are few. Methods: In total 99 Aeromonas species isolates from different sources (diarrheal children [n=23], non-diarrheal children [n=16], untreated drinking water from wells [n=32], and chicken carcasses [n=28] in Tripoli, Libya, were included in the present investigation. Genus identification was confirmed by biochemical analysis, and genospecies were determined using a combination of 16S rDNA variable region and gyrB sequence analysis. Polymerase chain reaction (PCR was used to detect genes encoding toxins from 52 of the isolates. Results: We identified 44 isolates (44% as A. hydrophila (3 [3.0%] subspecies anaerogenes, 23 [23%] subspecies dhakensis, and 18 [18%] subspecies ranae; 27 isolates (27% as A. veronii; 23 isolates (23% as A. caviae; and 5 isolates (5.0% as other genospecies. The genes encoding aerolysin (aer, cytolytic enterotoxin (act, and A. hydrophila isolate SSU enterotoxin (ast were detected in 45 (87%, 4 (7.7%, and 9 (17% of the 52 isolates tested, respectively. The gene encoding an extracellular lipase (alt was not detected. Conclusion: The majority of aeromonads from Libya fall within three genospecies (i.e. A. hydrophila, A. veronii, and A. caviae, and genes coding for toxin production are common among them.

Assessment of hygienic quality of some types of cheese sampled from retail outlets.

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Prencipe, Vincenza; Migliorati, Giacomo; Matteucci, Osvaldo; Calistri, Paolo; Di Giannatale, Elisabetta

2010-01-01

The authors evaluated the prevalence of Listeria monocytogenes, Escherichia coli O157:H7, Salmonella spp. and staphylococcal enterotoxin, in 2,132 samples selected from six types of cheese on the basis of recorded consumption in Italy in 2004. In L. monocytogenes-positive samples the precise level of contamination was determined. To define the physical-chemical characteristics of the selected natural cheeses, the pH values, water activity and sodium chloride content were determined. The results suggest that blue and soft cheeses (Brie, Camembert, Gorgonzola and Taleggio) are more likely to be contaminated with L. monocytogenes. The mean prevalence of L. monocytogenes in the six types of cheese was 2.4% (from 0.2% in Asiago and Crescenza to 6.5% in Taleggio), with contamination levels of up to 460 MPN/g. No presence of Salmonella spp. and E. coli O157 was found in any sample. Staphylococcus enterotoxin was found in 0.6% of the samples examined. Physical and chemical parameter values confirmed that all types of cheese are considered capable of supporting the growth of L. monocytogenes. The study confirmed the need to apply effective control at production and sales levels to reduce the probability of contamination by L. monocytogenes. This micro-organism can attain high levels of contamination in food products, such as cheeses that have a long shelf-life when associated with difficulties of maintaining appropriate storage temperatures in both sales points and in the home.

Assessment of hygienic quality of some types of cheese sampled from retail outlets

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Elisabetta Di Giannatale

2010-06-01

Full Text Available The authors evaluated the prevalence of Listeria monocytogenes, Escherichia coli O157:H7, Salmonella spp. and Staphylococcus enterotoxin, in 2 132 samples selected from six types of cheese on the basis of recorded consumption in Italy in 2004. In L. monocytogenes-positive samples the precise level of contamination was determined. To define the physical-chemical characteristics of the selected natural cheeses, the pH values, water activity and sodium chloride content were determined. The results suggest that blue and soft cheeses (Brie, Camembert, Gorgonzola and Taleggio are more likely to be contaminated with L. monocytogenes. The mean prevalence of L. monocytogenes in the six types of cheese was 2.4% (from 0.2% in Asiago and Crescenza to 6.5% in Taleggio, with contamination levels of up to 460 MPN/g. No presence of Salmonella spp. and E. coli O157 was found in any sample. Staphylococcus enterotoxin was found in 0.6% of the samples examined. Physical and chemical parameter values confirmed that all types of cheese are considered capable of supporting the growth of L. monocytogenes. The study confirmed the need to apply effective control at production and sales levels to reduce the probability of contamination by L. monocytogenes. This micro-organism can attain high levels of contamination in food products, such as cheeses that have a long shelf-life when associated with difficulties of maintaining appropriate storage temperatures in both sales points and in the home.

Investigating potential exogenous tumor initiating and promoting factors for Cutaneous T-Cell Lymphomas (CTCL), a rare skin malignancy

DEFF Research Database (Denmark)

Litvinov, Ivan V.; Shtreis, Anna; Kobayashi, Kenneth

2016-01-01

-Cell lymphotropic virus type 1 (HTLV1), Epstein-Barr virus (EBV), and herpes simplex virus (HSV). In this report, we review recent evidence evaluating the involvement of these agents in cancer initiation/progression. Most importantly, recent molecular experimental evidence documented for the first time that S....... aureus can activate oncogenic STAT3 signaling in malignant T cells. Specifically, S. aureus Enterotoxin type A (SEA) was recently shown to trigger non-malignant infiltrating T cells to release IL-2 and other cytokines. These signals upon binging to their cognate receptors on malignant T cells...

Viability of Staphylococcus xylosus isolated from artisanal sausages for application as starter cultures in meat products.

Science.gov (United States)

Fiorentini, Angela Maria; Sawitzki, Maristela Cortez; Bertol, Teresinha Marisa; Sant'anna, Ernani S

2009-01-01

Viability of Staphylococcus xylosus isolated from artisanal sausages for application as starter cultures in meat products Viability of Staphylococcus xylosus strains AD1 and U5 isolated from natural fermented sausages was investigated as starter cultures in fermented sausages produced in the South Region of Brazil. The study demonstrated that the Staphylococcus xylosus strains AD1 and U5 showed significant growth during fermentation, stability over freeze-dried process, negative reaction for staphylococcal enterotoxins and viability for using as a single-strain culture or associated with lactic acid bacteria for production of fermented sausages.

Serologic test systems development. Progress report, July 1, 1976--September 30, 1977

Energy Technology Data Exchange (ETDEWEB)

Saunders, G.C.; Clinard, E.H.; Bartlett, M.L.; Petersen, P.M.; Sanders, W.M.; Payne, R.J.; Martinez, E.

1978-01-01

Work has continued on the development and application of the Enzyme-Labeled Antibody (ELA) test to the USDA needs. Results on trichinosis, brucellosis, and staphylococcal enterotoxin A detection are very encouraging. A field test for trichinosis detection is being worked out in cooperation with Food Safety and Quality Service personnel. Work is in progress with the Technicon Instrument Corporation to develop a modification of their equipment to automatically process samples by the ELA procedure. An automated ELA readout instrument for 96-well trays has been completed and is being checked out.

Enterotoxigenicity of Staphylococcus aureus isolated from traditional and commercial dairy products marketed in Iran

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Ebrahim Rahimi

2013-01-01

Full Text Available This study was conducted to determine the prevalence rate, enterotoxigenecity, and antimicrobial resistance of S. aureus isolated from dairy products in Iran. From September 2010 to July 2011, a total of 347 samples from various dairy products, traditional and commercial, were collected from randomly selected retail stores. Overall, 20 samples (5.8% were found to be contaminated with S. aureus. The highest prevalence of S. aureus was found in traditional cheese (11.1%, followed by traditional ice-cream (5.9%, cream (5.6%, and butter (5.3%. The ability to synthesize classical staphylococcal enterotoxins (SEA-E was determined in 7 of 20 (35% isolates by using ELISA. SE type C was the most common enterotoxin found in the isolated S. aureus (42.9%, followed by SE type A (28.6%, SEA+SEC and SE type D (14.3%. Of the 20 isolates, 16 (80.0% were positive for one or more entrotoxin genes and 8 different genotypes were observed. Susceptibilities of the isolates were determined for 14 antimicrobial drugs using the disk diffusion assay. Most of the isolates (95.0% were resistant to one or more two antimicrobial agent and 45.0% of the isolates were resistant to three or more of drugs. Resistance to ampicillin was the most common finding (55.0%, followed by tetracycline (40.0% and penicillin G (30.0%. The results of this study showed the wide spread of enterotoxigenic and multidrug-resistant S. aureus strains in traditional dairy products in Iran and highlighted their public health hazards.

Genotypes, antibiotic resistance, and virulence factors of staphylococci from ready-to-eat food.

Science.gov (United States)

Podkowik, Magdalena; Bystroń, Jarosław; Bania, Jacek

2012-01-01

Sixty-seven staphylococcal isolates belonging to 12 species were obtained from 70 ready-to-eat food products. Staphylococcus aureus (n=25), and Staphylococcus epidermidis (n=13) were dominant. Susceptibility to penicillin, oxacillin, tetracycline, clindamycin, gentamicin, erythromycin, ciprofloxacin, and vancomycin was determined. All investigated S. aureus isolates were resistant to at least one antibiotic, and fifteen isolates were resistant to four and more antibiotics. Thirty-eight coagulase-negative staphylococci (CNS) isolates were resistant to at least one antibiotic, and seventeen to four and more antibiotics. Fifteen CNS isolates were mecA positive, and grew in the presence of 6 μg/mL oxacillin. All S. aureus isolates were mecA-negative. Arginine catabolic mobile element (ACME) was found in seven S. epidermidis isolates. Five S. epidermidis isolates harbored ica operon, ACME and were able to form biofilm. Three of them also possessed IS256 element and were mecA-positive. The expression of icaA gene was comparable in five ica-positive S. epidermidis isolates. One of six mecA positive S. epidermidis isolates was classified as sequence type (ST)155, one as ST110, and two as ST88. Two methicillin-resistant Staphylococcus epidermis (MRSE) belonged to new STs, that is, ST362, and ST363. Enterotoxin genes were found in 92% of S. aureus isolates. No enterotoxin gene was detected in analyzed CNS population. We show that ready-to-eat products are an important source of antibiotic-resistant CNS and potentially virulent strains of S. epidermidis, including genotypes undistinguishable from hospital-adapted clones.

Rapid differentiation of Staphylococcus aureus isolates harbouring egc loci with pseudogenes psient1 and psient2 and the selu or seluv gene using PCR-RFLP.

Science.gov (United States)

Collery, Mark M; Smyth, Cyril J

2007-02-01

The egc locus of Staphylococus aureus harbours two enterotoxin genes (seg and sei) and three enterotoxin-like genes (selm, seln and selo). Between the sei and seln genes are located two pseudogenes, psient1 and psient2, or the selu or seluv gene. While these two alternative sei-seln intergenic regions can be distinguished by PCR, to date, DNA sequencing has been the only confirmatory option because of the very high degree of sequence similarity between egc loci bearing the pseudogenes and the selu or seluv gene. In silico restriction enzyme digestion of genomic regions encompassing the egc locus from the 3' end of the sei gene through the 5' first quarter of the seln gene allowed pseudogene- and selu- or seluv-bearing egc loci to be distinguished by PCR-RFLP. Experimental application of these findings demonstrated that endonuclease HindIII cleaved PCR amplimers bearing pseudogenes but not those with a selu or seluv gene, while selu- or seluv-bearing amplimers were susceptible to cleavage by endonuclease HphI, but not by endonuclease HindIII. The restriction enzyme BccI cleaved selu- or seluv-harbouring amplimers at a unique restriction site created by their signature 15 bp insertion compared with pseudogene-bearing amplimers, thereby allowing distinction of these egc loci. PCR-RFLP analysis using these restriction enzymes provides a rapid, easy to interpret alternative to DNA sequencing for verification of PCR findings on the nature of an egc locus type, and can also be used for the primary identification of the intergenic sei-seln egc locus type.

Detection of Methicillin Resistance and Various Virulence Factors in Staphylococcus aureus Strains Isolated from Nasal Carriers

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Hatice Türk Dağı

2015-06-01

Full Text Available Background: Staphylococus aureus can be found as a commensal on skin and nasal flora or it may cause local and invasive infections. S. aureus has a large number of virulence factors. Aims: To investigate the methicillin resistance and frequency of various virulence factors in S. aureus nasal isolates. Study Design: Descriptive study. Methods: Nasal samples collected from university students were cultured in media. S. aureus was identified by conventional methods and the Staphyloslide latex test (Becton Dickinson, Sparks, USA. Antibiotic susceptibility tests were conducted, and the methicillin resistance was determined. The mecA, nuc, pvl and staphylococcal toxin genes were examined by polymerase chain reaction (PCR. Results: S. aureus was isolated in 104 of 600 (17.3% nasal samples. In total, 101 (97.1% S. aureus isolates were methicillin-sensitive and the remaining 3 (2.9% were methicillin-resistant. Furthermore, all but five isolates carried at least one staphylococcal enterotoxin gene, with seg being predominant. The tst and eta genes were determined in 29 (27.9%, and 3 (2.9% isolates, respectively. None of the S. aureus isolates harbored see, etb, and pvl genes. Conclusion: A moderate rate of S. aureus carriage and low frequency of MRSA were detected in healthy students. S. aureus isolates had a high prevalence of staphylococcal enterotoxin genes and the tst gene. In this study, a large number of virulence factors were examined in S. aureus nasal isolates, and the data obtained from this study can be used for monitoring the prevalence of virulence genes in S. aureus strains isolated from nasal carriers.

Detecção da toxina-1 da síndrome do choque tóxico em amostras de Staphylococcus aureus isoladas de mastite bovina Detection of toxic shock syndrome toxin by Staphylococcus aureus strains isolated from bovine mastitis

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H.F.T. Cardoso

2000-02-01

Full Text Available Este trabalho teve por objetivo caracterizar a produção da toxina da síndrome do choque tóxico (TSST-1 e de enterotoxinas estafilocócicas (SE A, B, C e D em 127 amostras de S. aureus, isoladas de amostras de leite proveniente de vacas com mastite no Estado de Minas Gerais, entre 1994 e 1997. A verificação da produção de toxinas foi feita pela técnica de sensibilidade ótima em placa. Das 127 amostras testadas, 60 (47% eram produtoras de TSST-1 e 54 (43% produtoras de SE, 38 amostras produziram SED (30%, 24 SEB (19%, 8 SEC (6% e 4 SEA (3%. Estes resultados trazem preocupações quanto à saúde pública pela alta prevalência de amostras de S. aureus produtoras de TSST-1 e de enterotoxinas em isolamentos a partir de leite de vacas com mastite.A total of 127 strains of Staphylococcus aureus were examined for the production of toxic shock syndrome toxin (TSST-1 and staphylococcal enterotoxins (SE A, B, C and D. The strains were isolated from milk samples from cows with mastitis in dairy herds of Minas Gerais State, Brazil, from 1994 to 1997. The toxins were detected using the optimum-sensitivity plate method. Of 127 isolates, 60 (47% produced TSST-1 and 54 (43% produced SE, 38 (30% produced SED, 24 (19% SEB, 8 (6% SEC and 4 (3% enterotoxin A..

Molecular characterization and antibiotic resistance of enterotoxigenic and entero-aggregative Escherichia coli isolated from raw milk and unpasteurized cheeses

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Mojtaba Bonyadian

2014-04-01

Full Text Available The aim of this study was to determine the occurrence of enterotoxigenic and enteroaggregative Escherichia coli strains and antibiotic resistance of the isolates in raw milk and unpasteurized cheese. Out of 200 samples of raw milk and 50 samples of unpasteurized cheeses, 96 and 24 strains of E. coli were isolated, respectively. Polymerase chain reaction (PCR was used to detect the genes encoding heat-stable enterotoxin a (STa, heat-stable enterotoxin b (STb, heat labile toxin (LT and enteroaggregative heat-stable toxin1 (EAST1. Twelve out of 120 (10.00% isolates harbored the gene for EAST1, 2(1.66% isolates were detected as producing STb and LT toxins and 12 (10.00% strains contained STb and EAST1 genes. None of the strains contain the STa gene. All of the strains were tested for antibiotic resistance by disk diffusion method. Disks included: ciprofloxacin (CFN, trimetoprim-sulfamethoxazole (TSX, oxytetracycline (OTC, gentamicin (GMN, cephalexin (CPN, nalidixic acid (NDA and nitrofurantoin (NFN, ampicillin (AMP, neomycin (NEO and streptomycin (STM. Among 120 isolated strains of E. coli, the resistance to each antibiotics were as follows: OTC100%, CPN 86.00%, NDA 56.00%, NFN 42.00%, GMN 30.00%, TSX 28.00%, CFN 20%, AM 23.40% and STM 4.25%. None of the isolates were resistant to NEO. The present data indicate that different resistant E. coli pathogens may be found in raw milk and unpasteurized cheese. It poses an infection risk for human and transferring the resistant factors to microflora of the consumers gut.

IL-2 regulates SEB induced toxic shock syndrome in BALB/c mice.

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Aslam Ali Khan

2009-12-01

Full Text Available Toxic Shock Syndrome (TSS is characterized by fever, rash, hypotension, constitutional symptoms, and multi-organ involvement and is caused by Staphylococcus aureus enterotoxins such as Staphylococcal Enterotoxin B (SEB. SEB binds to the MHC-IIalpha chain and is recognized by the TCRbeta chain of the Vbeta8 TCR(+ T cells. The binding of SEB to Vbeta chain results in rapid activation of T cells and production of inflammatory cytokines, such as Interleukin-2 (IL-2, Interferon-gamma and Tumor Necrosis Factor-alpha which mediate TSS. Although IL2 was originally identified as the T cell growth factor and was proposed to contribute to T cell differentiation, its role in TSS remains unexplored.Mice were injected with D-Gal (25 mg/mouse. One hour after D-Galactosamine (D-Gal injection each mouse was injected with SEB (20 microg/mouse. Mice were then observed for 72 hrs and death was recorded at different times. We tested Interleukin-12, IFNgamma, and IL-2 deficient mice (IL-2(-/-, but only the IL-2 deficient mice were resistant to SEB induced toxic shock syndrome. More importantly reconstitution of IL-2 in IL-2 deficient mice restored the shock. Interestingly, SEB induced IL-2 production from T cells was dependent on p38MAPK activation in macrophages as inhibition of it in macrophages significantly inhibited IL-2 production from T cells.This study shows the importance of IL -2 in TSS which has not been previously explored and it also shows that regulating macrophages function can regulate T cells and TSS.

The Role of Intestinal Bacteria in Acute Diarrheal Diseases

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1981-01-01

another enteric pathogen, Vibrio cholerae , is present in high number in the cral cavity during acute and convalescent periods. Also, the buccal cells...h42 LT/ST + - - 2 K32&c4 AD) Kenya ... . .- r K32ScT All Kenya - LT - - - NT TD46ZcT ADY Mexfco 06:HT6 LT/ST - - - Z13 TD260cl AD Mexico 06:H16 LT...2 TD514c! AD Mexico .. . . NT TD472cI A Mexico LT/ST - - NT TD514cZ AD Mexico - NT "TABLE 1 CONTINUED 2 1 Enterotoxins MR-HA Pili 3 Human Strains

Staphylococcus aureus in Some Brazilian Dairy Industries: Changes of Contamination and Diversity

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Dittmann, Karen Kiesbye; Chaul, Luiza T.; Lee, Sarah H. I.

2017-01-01

Staphylococcus aureus, a major food-poisoning pathogen, is a common contaminant in dairy industries worldwide, including in Brazil. We determined the occurrence of S. aureus in five dairies in Brazil over 8 months. Of 421 samples, 31 (7.4%) were positive for S. aureus and prevalence varied from 0.......8% of strains being sensitive to all antibiotic classes and no Methicillin-resistant S. aureus (MRSA) strains were found. The enterotoxin-encoding genes involved in food-poisoning, e.g., sea, sed, see, and seg were targeted by PCR. The two toxin-encoding genes, sed and see, were not detected. Only three strains...... contamination....

Prevalence, virulence factor genes and antibiotic resistance of Bacillus cereus sensu lato isolated from dairy farms and traditional dairy products

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Owusu-Kwarteng, James; Wuni, Alhassan; Akabanda, Fortune

2017-01-01

of B. cereus sensu lato isolated from cattle grazing soils and dairy products in Ghana. A total of 114 samples made up of 25 soil collected from cattle grazing farm land, 30 raw milk, 28 nunu (yoghurt-like product) and 31 woagashie (West African soft cheese). Ninety-six B. cereus sensu lato isolates......%), oxacillin (92%), penicillin (100%), amoxicillin (100%), and cefepime (100%) but susceptible to other antibiotics tested. Conclusions: Bacillus cereus s. l. is prevalent in soil, raw milk and dairy products in Ghana. However, loads are at levels considered to be safe for consumption. Various enterotoxin...

PCR múltiple para la detección de los genes sea, seb, sec, sed y see de Staphylococcus aureus: Caracterización de aislamientos de origen alimentario Multiplex PCR for the detection of sea, seb, sec, sed and see genes of Staphylococcus aureus: Characterization of isolates from food

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E. A. Manfredi

2010-09-01

Full Text Available La presencia de Staphylococcus aureus en los alimentos representa un riesgo potencial para la salud pública; sus enterotoxinas son el principal factor de virulencia. La detección de las enterotoxinas de S. aureus puede realizarse por ELISA, aunque sólo es posible detectar el pool de enterotoxinas SEA, SEB, SEC, SED y SEE. Los objetivos del presente trabajo fueron optimizar dos técnicas de PCR múltiple para la detección de los genes sea, seb, sec, sed y see de S. aureus y caracterizar un conjunto de 115 aislamientos de Staphylococcus spp. asociados a intoxicaciones alimentarias provenientes de diferentes provincias de Argentina. La caracterización se realizó por pruebas bioquímicas, ELISA y PCR. Sesenta y ocho aislamientos (59,1% fueron positivos por ELISA, mientras que 61 (53% fueron positivos por PCR. De los aislamientos positivos por PCR, 34 (55,7% portaron el gen sea, 9 (14,8% el gen seb, 5 (8,1% el gen see, 4 (6,5% el gen sec, 6 (9,9% los genes sea y seb, 2 (3,3% los genes sea y sec, y 1 (1,7% los genes sea y sed. Este es el primer estudio de caracterización genotípica de aislamientos de S. aureus asociados con brotes de intoxicación alimentaria registrados en distintas provincias argentinas.The presence of Staphylococcus aureus in food represents a potential risk to public health, being its enterotoxins the major virulence factor. Enterotoxin detection can be determined by ELISA, but only for the pool of enterotoxins SEA, SEB, SEC, SED and SEE. The main aims of this study were to optimize two PCR techniques for detection of S. aureus sea, seb, sec, sed and see, and to characterize Staphylococcus spp. isolates associated with food intoxication. Two PCR techniques were optimized and 115 Staphylococcus spp. isolates from Ciudad Autónoma de Buenos Aires, and Buenos Aires, Córdoba, and Neuquén provinces were characterized. The characterization was performed by biochemical tests, ELISA and PCR. Sixty-eight isolates (59.1% were

Transcriptomic and metabolic responses of Staphylococcus aureus in mixed culture with Lactobacillus plantarum, Streptococcus thermophilus and Enterococcus durans in milk.

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Zdenkova, Kamila; Alibayov, Babek; Karamonova, Ludmila; Purkrtova, Sabina; Karpiskova, Renata; Demnerova, Katerina

2016-09-01

Staphylococcus aureus is a major food-borne pathogen due to the production of enterotoxin and is particularly prevalent in contaminated milk and dairy products. The lactic acid bacteria (LAB) are widely used as biocontrol agents in fermented foods which can inhibit pathogenic flora. In our work, we investigated the influence of three strains of LAB (Lactobacillus plantarum, Streptococcus thermophilus and Enterococcus durans) on the relative expression of three enterotoxin genes (sea, sec, sell) and eight virulence and/or regulatory genes (sarA, saeS, codY, srrA, rot, hld/RNAIII, agrA/RNAII, sigB) in two S. aureus strains (MW2 and Sa1612) in TSB and reduced-fat milk (1.5 %) at 30 °C over a 24-h period. The tested LAB and S. aureus strains proved to be mutually non-competitive or only slightly competitive during co-cultivation. In addition, under the above-mentioned conditions, differential gene expression between the S. aureus MW2 and Sa1612 strains was well documented. S. aureus growth was changed in mixed culture with LAB; however, its effect on the repression of sea and sec expression correlated with production of these virulence factors. In comparison, the presence of LAB strains generally inhibited the expression of sec, sell, sarA, seaS, agrA/RNAII and hld/RNAIII genes. The effect of LAB strains presence on the expression of sea, codY, srrA, rot and sigB genes was medium, time, LAB and S. aureus strain specific. SEA and SEC production was significantly reduced in milk compared to TSB in pure culture. After the 24-h cultivation, S. aureus MW2 and Sa1612 SEC production was 187 and 331 times lower in milk compared to TSB, respectively (0.07 and 0.39 ng/mL in milk, versus 13.1 and 129.2 ng/mL in TSB, respectively). At the same time S. aureus MW2 and Sa1612 SEA production was 77 and 68 times lower in milk compared to TSB, respectively (0.99 and 0.17 ng/mL in milk, versus 76.4 and 11.5 ng/mL in TSB, respectively). This study has revealed new insights into the

Genotypic and phenotypic characterization of Clostridium perfringens isolates from Darmbrand cases in post-World War II Germany.

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Ma, Menglin; Li, Jihong; McClane, Bruce A

2012-12-01

Clostridium perfringens type C strains are the only non-type-A isolates that cause human disease. They are responsible for enteritis necroticans, which was termed Darmbrand when occurring in post-World War II Germany. Darmbrand strains were initially classified as type F because of their exceptional heat resistance but later identified as type C strains. Since only limited information exists regarding Darmbrand strains, this study genetically and phenotypically characterized seven 1940s era Darmbrand-associated strains. Results obtained indicated the following. (i) Five of these Darmbrand isolates belong to type C, carry beta-toxin (cpb) and enterotoxin (cpe) genes on large plasmids, and express both beta-toxin and enterotoxin. The other two isolates are cpe-negative type A. (ii) All seven isolates produce highly heat-resistant spores with D(100) values (the time that a culture must be kept at 100°C to reduce its viability by 90%) of 7 to 40 min. (iii) All of the isolates surveyed produce the same variant small acid-soluble protein 4 (Ssp4) made by type A food poisoning isolates with a chromosomal cpe gene that also produce extremely heat-resistant spores. (iv) The Darmbrand isolates share a genetic background with type A chromosomal-cpe-bearing isolates. Finally, it was shown that both the cpe and cpb genes can be mobilized in Darmbrand isolates. These results suggest that C. perfringens type A and C strains that cause human food-borne illness share a spore heat resistance mechanism that likely favors their survival in temperature-abused food. They also suggest possible evolutionary relationships between Darmbrand strains and type A strains carrying a chromosomal cpe gene.

Evaluation of antimicrobial susceptibilities and virulence factors of Staphylococcus aureus strains isolated from community-acquired and health-care associated pediatric infections.

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Karbuz, Adem; Karahan, Zeynep Ceren; Aldemir-Kocabaş, Bilge; Tekeli, Alper; Özdemir, Halil; Güriz, Haluk; Gökdemir, Refik; İnce, Erdal; Çiftçi, Ergin

2017-01-01

Karbuz A, Karahan ZC, Aldemir-Kocabaş B, Tekeli A, Özdemir H, Güriz H, Gökdemir R, İnce E, Çiftçi E. Evaluation of antimicrobial susceptibilities and virulence factors of Staphylococcus aureus strains isolated from community-acquired and health-care associated pediatric infections. Turk J Pediatr 2017; 59: 395-403. The aim of this study was to investigate the enterotoxins and Panton-Valentine leukocidin (PVL) gene as virulence factor, identification if antimicrobial sensitivity patterns, agr (accessory gene regulator) types and sequence types and in resistant cases to obtain SCCmec (staphylococcal cassette chromosome mec) gene types which will be helpful to decide empirical therapy and future health politics for S. aureus species. Total of 150 isolates of S. aureus were isolated from the cultures of the child patients in January 2011 and December 2012. In this study, the penicillin resistance was observed as 93.8%. PVL and mecA was detected positive in 8.7% and in 6% of all S. aureus strains, respectively. Two MRSA (methicillin resistant S.aureus) strains were detected as SCCmec type III and SCCmec type V and five MRSA strains were detected as SCCmec type IV. SET-I and SET-G were the most common detected enterotoxins. In both community-associated and healthcare-associated MRSA strains, agr type 1 was detected most commonly. The most common sequence types were ST737 in 13 patients than ST22 in eight patients and ST121 in six patients. This study highlights a necessity to review the cause of small changes in the structural genes in order to determine whether it is a cause or outcome; community-acquired and healthcare associated strains overlap.

Virulence factors and genetic variability of Staphylococcus aureus strains isolated from raw sheep's milk cheese.

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Spanu, Vincenzo; Spanu, Carlo; Virdis, Salvatore; Cossu, Francesca; Scarano, Christian; De Santis, Enrico Pietro Luigi

2012-02-01

Contamination of dairy products with Staphylococcus aureus can be of animal or human origin. The host pathogen relationship is an important factor determining genetic polymorphism of the strains and their potential virulence. The aim of the present study was to carry out an extensive characterization of virulence factors and to study the genetic variability of S. aureus strains isolated from raw ewe's milk cheese. A total of 100 S. aureus strains isolated from cheese samples produced in 10 artisan cheese factories were analyzed for the presence of enterotoxins (sea-see) and enterotoxins-like genes (seh, sek, sel, sem, seo, sep), leukocidins, exfoliatins, haemolysins, toxic shock syndrome toxin 1 (TSST-1) and the accessory gene regulator alleles (agr). Strains were also typed using pulsed-field gel electrophoresis (PFGE). AMOVA analysis carried out on PFGE and PCR data showed that the major component explaining genetic distance between strains was the dairy of origin. Of the total isolates 81% had a pathogenicity profile ascribable to "animal" biovar while 16% could be related to "human" biovar. The biovar allowed to estimate the most likely origin of the contamination. Minimum inhibitory concentrations (MICs) of nine antimicrobial agents and the presence of the corresponding genes coding for antibiotic resistance was also investigated. 18 strains carrying blaZ gene showed resistance to ampicillin and penicillin and 6 strains carrying tetM gene were resistant to tetracycline. The presence of mecA gene and methicillin resistance, typical of strains of human origin, was never detected. The results obtained in the present study confirm that S. aureus contamination in artisan cheese production is mainly of animal origin. Copyright © 2011. Published by Elsevier B.V.

Genotyping of Staphylococcus aureus in bovine mastitis and correlation to phenotypic characteristics.

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Artursson, Karin; Söderlund, Robert; Liu, Lihong; Monecke, Stefan; Schelin, Jenny

2016-09-25

Reducing the prevalence of mastitis caused by Staphylococcus aureus (S. aureus) is essential to improve animal health and reduce economic losses for farmers. The clinical outcome of acute mastitis and risk of progression to persistent mastitis can, at least to some extent, be related to genetic variants of the strain causing the infection. In the present study we have used microarrays to investigate the presence of virulence genes in S. aureus isolates from dairy cows with acute clinical mastitis (n=70) and correlated the findings to other genotypic and phenotypic characteristics. Among the most commonly found virulence factors were genes encoding several hemolysin types, leukocidins D and lukM/lukF-P83, clumping factors A and B, fibrinogen binding protein and fibronectin-binding protein A. Some virulence factors e.g. fibronectin-binding protein B and Staphylococcus aureus surface protein G were less common. Genes coding for several staphylococcal enterotoxins and toxic shock syndrome toxin-1 (TSST-1) were commonly found, especially in one major pulsotype. No beta-lactamase genes were found in any common pulsotype, while present in some rare pulsotypes, indicated to be of human origin. Production of TSST-1, enterotoxins, hemolysins and beta-lactamase could all be positively correlated to presence of the corresponding genes. This study reveals a number of genotypic differences and similarities among common and rare pulsotypes of S. aureus from cases of mastitis in Sweden. The results could help the design of diagnostic tools to guide on-farm interventions according to the expected impact on udder health from a specific S. aureus genotype. Copyright © 2016 Elsevier B.V. All rights reserved.

Incidence of Aeromonas spp. infection in fish and chicken meat and its related public health hazards: A review

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Praveen Kumar Praveen

2016-01-01

Full Text Available Aeromonas is recognized to cause a variety of diseases in man. In humans, they are associated with intestinal and extraintestinal infections. With the growing importance of Aeromonas as an emerging pathogen, it is important to combat this organism. It is indisputable that Aeromonas strains may produce many different putative virulence factors such as enterotoxins, hemolysins or cytotoxins, and antibiotic resistance against different antibiotics. The ability of these bacteria to grow competitively at 5°C may be indicative of their potential as a public health hazard. Comprehensive enteric disease surveillance strategies, prevention and education are essential for meeting the challenges in the years ahead. It is important for us to promote the value of enteric cultures when patients have a gastrointestinal illness or bloody diarrhea or when multiple cases of enteric disease occur after a common exposure. With the growing importance of Aeromonas as an emerging pathogen, it is important to combat this organism. It is indisputable that Aeromonas strains may produce many different putative virulence factors, such as enterotoxins, hemolysins or cytotoxins. It has been established that aerolysin is a virulence factor contributing to the pathogenesis of Aeromonas hydrophila infection. Fish and chicken play an important role in the transmission of this pathogen to humans. In the present study, the high prevalence of toxin-producing strains was found among the Aeromonas isolates. The ability of these bacteria to grow competitively at 5°C may be indicative of their potential as a public health hazard. The present review was constructed with a view to highlight the zoonotic importance of Aeromonas pathogen in fish and chicken meat.

Growth of enterotoxigenic Staphylococcus aureus in povi masima, a traditional Pacific island food.

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Wong, T L; Whyte, R J; Graham, C G; Saunders, D; Schumacher, J; Hudson, J A

2004-01-01

To obtain preliminary data on the microbiology and hurdles to pathogen growth in the traditional Pacific Island food, povi masima, which is essentially beef brisket cured in brine. Six containers of povi masima were prepared and two were inoculated with five enterotoxigenic strains of Staphyloccocus aureus. The povi masima were divided into two lots each containing two uninoculated control and an inoculated container. Lot 1 was incubated at room temperature (20 degrees C) and lot 2 under refrigeration (4-5 degrees C) for up to 98 days. During storage, samples were removed and tested for aerobic plate count, coagulase-producing Staphylococci, Clostridium perfringens, staphylococcal enterotoxin and various chemical parameters of the food. Coagulase-producing Staphylococci and aerobic plate counts grew to high levels in both the inoculated and uninoculated lots stored at room temperature, but enterotoxin was only detected at one time point in these lots and this may represent a false positive result. The concentration of NaCl in the meat increased with time as concentrations equilibrated, and nitrite was rapidly lost in those lots stored at room temperature. Storage at 4-5 degrees C prevented proliferation of coagulase-producing Staphylococci. For safe curing and storage, this food should be kept under refrigeration as this prevented growth of staphylococci. Optimum storage would also be achieved with improved attempts to ensure equal distribution of NaCl prior to storage. Under conditions traditionally used to cure and store this food, enterotoxigenic staphylococci can grow to numbers where toxigenesis might occur, especially during the early stages of curing where the salt has not diffused from the brine into the meat.

Incidence of Aeromonas spp. infection in fish and chicken meat and its related public health hazards: A review.

Science.gov (United States)

Praveen, Praveen Kumar; Debnath, Chanchal; Shekhar, Shashank; Dalai, Nirupama; Ganguly, Subha

2016-01-01

Aeromonas is recognized to cause a variety of diseases in man. In humans, they are associated with intestinal and extra-intestinal infections. With the growing importance of Aeromonas as an emerging pathogen, it is important to combat this organism. It is indisputable that Aeromonas strains may produce many different putative virulence factors such as enterotoxins, hemolysins or cytotoxins, and antibiotic resistance against different antibiotics. The ability of these bacteria to grow competitively at 5°C may be indicative of their potential as a public health hazard. Comprehensive enteric disease surveillance strategies, prevention and education are essential for meeting the challenges in the years ahead. It is important for us to promote the value of enteric cultures when patients have a gastrointestinal illness or bloody diarrhea or when multiple cases of enteric disease occur after a common exposure. With the growing importance of Aeromonas as an emerging pathogen, it is important to combat this organism. It is indisputable that Aeromonas strains may produce many different putative virulence factors, such as enterotoxins, hemolysins or cytotoxins. It has been established that aerolysin is a virulence factor contributing to the pathogenesis of Aeromonas hydrophila infection. Fish and chicken play an important role in the transmission of this pathogen to humans. In the present study, the high prevalence of toxin-producing strains was found among the Aeromonas isolates. The ability of these bacteria to grow competitively at 5°C may be indicative of their potential as a public health hazard. The present review was constructed with a view to highlight the zoonotic importance of Aeromonas pathogen in fish and chicken meat.

[Carriage of Staphylococcus aureus among food service workers].

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Alarcón-Lavín, María Paula; Oyarzo, Carolina; Escudero, Carlos; Cerda-Leal, Fabiola; Valenzuela, Francisco J

2017-12-01

Background Staphylococcus aureus produces 11 serotypes of endotoxins that may cause food poisoning. Aim To determine the prevalence of type A enterotoxigenic Staphylococcus aureus carriage among food service workers in Chillan, Chile. Material and Methods Pharyngeal swabs were obtained from 100 food service workers and were cultured in Agar plates. After identifying the presence of Staphylococcus aureus, DNA was extracted to identify type A toxin by conventional PCR. Results Thirty eight percent of samples were colonized with Staphylococcus aureus. Among these, 26% were toxin A producers. Conclusions Half of the sampled workers carried Staphylococcus aureus and a quarter of these produced type A enterotoxin.

BACTERIA STAPHYLOCOCCUS SPP. IZOLATED FROM MASTITIS OF SHEEP AND THEIR ENTEROTOXIGENIC PROPERTIES

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František Zigo

2011-12-01

Full Text Available In our study was followed occurrence of staphylococcal mastitis in herd of 350 sheep during one lactation season. We found, the bacteria S. schleiferi was identified in 88 from all 204 isolates. In high number were identified also S. caprae (33, S. chromogenes (21, S. aureus (19, S. epidermidis (17, respectively. Important was occurrence S. intermedius, S. simulans, S. xylosus a S. warneri, too. The Staphylococcuss spp. caused latent and subclinical forms of mastitis predominantly, showed into subacute mastitis (26.5%. Acute mastitis was determined in 7.8 %. Eight bacteria S. aureus, two S. chromogenes, and two S. epidermidis produced staphylococcal enterotoxins.doi: 10.5219/171

The Effect Of PHA And SEA On Mitotic Index Of Lymphocyte Cell Of Macaca Fasciulare

International Nuclear Information System (INIS)

Lubis, Masnelli; Iwiq-Indrawati

2003-01-01

The observation of influences of PHA (phytohemagglutinin) and SEA (staphilucoccal enterotoxin A) on mitotic index of lymphocyte of Macaca Fascicularis had been done. Half milliliters of lymphocyte cells stimulated with PHA or SEA were cultured in 10 ml RPMI + 1.0 ml Fetal Bouvine Serum (FBS ) + 0.1 ml L-glutamine + 0.15 ml PHA or 0.1 ml SEA ( 0.5 μg/ml ) + 0.1 ml Colchisin on 37 degree C for 96 hours. The result demonstrated that the frequency of mitotic index stimulated with PHA was higher than that of SEA. The average of mitotic index with PHA was 18.56 %, and with SEA was 8.3 %. (author)

SCCmec Type IX Element in Methicillin Resistant Staphylococcusaureusspa Type t337 (CC9) Isolated from Pigs and Pork in Thailand

DEFF Research Database (Denmark)

Vestergaard, Martin; Cavaco, Lina M; Sirichote, Pantip

2012-01-01

blaZ, aac-aphD, vga(A), tetM, and a tet efflux marker, in all strains and additionally ermB and aadD, cat and fex(A) in the pork isolates. None of the isolates were found PVL-positive, but enterotoxins were identified in all isolates. To our knowledge, only a few descriptions of MRSA in livestock...... and food products in Thailand have been observed but this is the first observation of MRSA CC9 associated with SCCmec IX in pork. This study indicates a likely widespread distribution of MRSA in pig and pork in Thailand and further investigation on the prevalence and importance of livestock associated MRSA...

Genetic mapping of the regulator gene determining enterotoxin synthesis in Vibrio cholerae

International Nuclear Information System (INIS)

Smirnova, N.I.; Livanova, L.F.; Shaginyan, I.A.; Motin, V.L.

1986-01-01

Data on the genetic mapping of mutation tox-7 (the mutation affecting the synthesis of the cholera toxin) were obtained by conjugation crosses between the atoxigenic donor strain Vibrio cholerae Eltor and the toxigenic recipient strain V. cholera classica. The molecular and genetic analysis of the Tox - recombinants indicated that, when the synthesis of the cholera toxin is disrupted in these strains, the tox-7 mutation (which impairs the regulator gene tox) is gained. Close linkage between the tox-7 and pur-63 mutations was established (during the selection procedure there was 81.1% combined transfer with respect to marker pur-63 situated in the donor strain chromosome more proximal than mutation tox-7). The markers were localized in the following order in the region under investigation: asp-cys-nal-pur-61-trp-his-pur-63-tox-7-ile

Multilocus Sequence Typing and Virulence-Associated Gene Profile Analysis of Staphylococcus aureus Isolates From Retail Ready-to-Eat Food in China.

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Yang, Xiaojuan; Yu, Shubo; Wu, Qingping; Zhang, Jumei; Wu, Shi; Rong, Dongli

2018-01-01

The aim of this study was to characterize the subtypes and virulence profiles of 69 Staphylococcus aureus isolates obtained from retail ready-to-eat food in China. The isolates were analyzed using multilocus sequence typing (MLST) and polymerase chain reaction (PCR) analysis of important virulence factor genes, including the staphylococcal enterotoxin (SE) genes ( sea , seb , sec , sed , see , seg , seh , sei , sej ), the exfoliative toxin genes ( eta and etb ), the toxic shock syndrome toxin-1 gene ( tst ), and the Panton-Valentine leucocidin-encoding gene ( pvl ). The isolates encompassed 26 different sequence types (STs), including four new STs (ST3482, ST3484, ST3485, ST3504), clustered in three clonal complexes and 17 singletons. The most prevalent STs were ST1, ST6, and ST15, constituting 34.8% of all isolates. Most STs (15/26, 57.7%) detected have previously been associated with human infections. All 13 toxin genes examined were detected in the S. aureus isolates, with 84.1% of isolates containing toxin genes. The three most prevalent toxin genes were seb (36.2%), sea (33.3%), and seg (33.3%). The classical SE genes ( sea - see ), which contribute significantly to staphylococcal food poisoning (SFP), were detected in 72.5% of the S. aureus isolates. In addition, pvl , eta , etb , and tst were found in 11.6, 10.1, 10.1, and 7.2% of the S. aureus isolates, respectively. Strains ST6 carrying sea and ST1 harboring sec-seh enterotoxin profile, which are the two most common clones associated with SFP, were also frequently detected in the food samples in this study. This study indicates that these S. aureus isolates present in Chinese ready-to-eat food represents a potential public health risk. These data are valuable for epidemiological studies, risk management, and public health strategies.

Antimicrobial resistance and prevalence of CvfB, SEK and SEQ genes among Staphylococcus aureus isolates from paediatric patients with bloodstream infections.

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Liang, Bing-Shao; Huang, Yan-Mei; Chen, Yin-Shuang; Dong, Hui; Mai, Jia-Liang; Xie, Yong-Qiang; Zhong, Hua-Min; Deng, Qiu-Lian; Long, Yan; Yang, Yi-Yu; Gong, Si-Tang; Zhou, Zhen-Wen

2017-11-01

Staphylococcus aureus ( S. aureus ) is one of the most frequently isolated pathogens in neonatal cases of early and late-onset sepsis. Drug resistance profiles and carriage of toxin genes may affect the treatment and outcome of an infection. The present study aimed to determine the antimicrobial resistance patterns and frequencies of the toxin-associated genes conserved virulence factor B (CvfB), staphylococcal enterotoxin Q (SEQ) and staphylococcal enterotoxin K (SEK) among S. aureus isolates recovered from paediatric patients with bloodstream infections (BSIs) in Guangzhou (China). Of the 53 isolates, 43.4% were methicillin-resistant S. aureus (MRSA), and resistance rates to penicillin, erythromycin, clindamycin, trimethoprim/sulfamethoxazole, tetracycline, and ciprofloxacin of 92.5, 66.0, 62.3, 13.2, 20.8 and 1.9% were recorded, respectively. However, no resistance to nitrofurantoin, dalfopristin/quinupristin, rifampicin, gentamicin, linezolid or vancomycin was detected. Resistance to erythromycin, clindamycin and tetracycline in the MRSA group was significantly higher than that in the methicillin-susceptible S. aureus (MSSA) group. No significant differences in antimicrobial resistance patterns were noted between two age groups (≤1 year and >1 year). The proportion of S. aureus isolates positive for CvfB, SEQ and SEK was 100, 34.0 and 35.8%, respectively, with 24.5% (13/53) of strains carrying all three genes. Compared with those in MSSA isolates, the rates of SEK, SEQ and SEK + SEQ carriage among MRSA isolates were significantly higher. Correlations were identified between the carriage of SEQ, SEK and SEQ + SEK genes and MRSA (contingency coefficient 0.500, 0.416, 0.546, respectively; Pstudy clarified the characteristics of BSI-associated S. aureus and enhanced the current understanding of the pathogenicity and treatment of MRSA.

Multilocus Sequence Typing and Virulence-Associated Gene Profile Analysis of Staphylococcus aureus Isolates From Retail Ready-to-Eat Food in China

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Xiaojuan Yang

2018-03-01

Full Text Available The aim of this study was to characterize the subtypes and virulence profiles of 69 Staphylococcus aureus isolates obtained from retail ready-to-eat food in China. The isolates were analyzed using multilocus sequence typing (MLST and polymerase chain reaction (PCR analysis of important virulence factor genes, including the staphylococcal enterotoxin (SE genes (sea, seb, sec, sed, see, seg, seh, sei, sej, the exfoliative toxin genes (eta and etb, the toxic shock syndrome toxin-1 gene (tst, and the Panton-Valentine leucocidin-encoding gene (pvl. The isolates encompassed 26 different sequence types (STs, including four new STs (ST3482, ST3484, ST3485, ST3504, clustered in three clonal complexes and 17 singletons. The most prevalent STs were ST1, ST6, and ST15, constituting 34.8% of all isolates. Most STs (15/26, 57.7% detected have previously been associated with human infections. All 13 toxin genes examined were detected in the S. aureus isolates, with 84.1% of isolates containing toxin genes. The three most prevalent toxin genes were seb (36.2%, sea (33.3%, and seg (33.3%. The classical SE genes (sea–see, which contribute significantly to staphylococcal food poisoning (SFP, were detected in 72.5% of the S. aureus isolates. In addition, pvl, eta, etb, and tst were found in 11.6, 10.1, 10.1, and 7.2% of the S. aureus isolates, respectively. Strains ST6 carrying sea and ST1 harboring sec-seh enterotoxin profile, which are the two most common clones associated with SFP, were also frequently detected in the food samples in this study. This study indicates that these S. aureus isolates present in Chinese ready-to-eat food represents a potential public health risk. These data are valuable for epidemiological studies, risk management, and public health strategies.

Brote de intoxicación alimentaria asociado al consumo de leche ultrapasteurizada en la República del Paraguay Foodborne outbreak associated with consumption of ultrapasteurized milk in the Republic of Paraguay

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Natalie Weiler

2011-03-01

Full Text Available Durante marzo de 2007 ocurrió un brote epidémico asociado al consumo de leche ultrapasteurizada que afectó a las ciudades de San Lorenzo, Ciudad del Este y Asunción, de la República del Paraguay. Las personas afectadas fueron 400, de las cuales 60 requirieron hospitalización. Se aisló S. aureus subespecie aureus de 5 pacientes, 3 operarios y 3 muestras de leche. Todas las cepas fueron productoras de enterotoxinas. Las aislamientos de 3 pacientes, de un operario y de las muestras de leche portaron los genes que codifican las enterotoxinas C (sec y D (sed, y presentaron un patrón único de macrorrestricción (SmaI-PFGE. Se identificó a la leche como fuente de intoxicación y a un operario de la línea de producción como origen de la contaminación. Este es el primer brote de ETA denunciado en Paraguay en el cual se pudo aislar, caracterizar y subtipificar el agente etiológico en la planta de elaboración, en el alimento y en las personas afectadas.During March 2007 there was an epidemic outbreak associated with the consumption of ultrapasteurized milk. Four hundred people were affected and 60 required hospitalization. S. aureus subspecies aureus was isolated from 5 patients, 3 operators and 3 milk samples. All strains produced enterotoxins. Strains isolated from 3 patients, one operator and all the milk samples carried the genes encoding enterotoxins C (sec and D (sed, and showed an indistinguishable macrorestriction pattern (SmaI-PFGE. Milk was identified as the source of intoxication and a production line operator as the source of contamination. This is the first foodborne outbreak reported in Paraguay whose agent was isolated, characterized and subtypified in the production plant, the food and the affected people.

Heterogeneity and phylogenetic relationships of community-associated methicillin-sensitive/resistant Staphylococcus aureus isolates in healthy dogs, cats and their owners.

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Wan, M T; Fu, S Y; Lo, Y P; Huang, T M; Cheng, M M; Chou, C C

2012-01-01

To investigate the distribution of staphylococcal enterotoxin genes (se) and the molecular features of community-associated methicillin-sensitive/resistant Staphylococcus aureus (CA-MSSA/MRSA) isolates in the nostrils of healthy pets and their owners. A total of 114 Staph. aureus isolates were identified from 1563 nasal swab samples, and CA-MRSA accounted for 20·2% (n = 23) of the total identified isolates. CA-MRSA isolates (91·3%, 21/23) harboured higher percentage of se than did CA-MSSA isolates (58·2%, 53/91) (P human bond caused by CA-staphylococci in the commonwealth and the need to take cautions worldwide. © 2011 The Authors. Journal of Applied Microbiology © 2011 The Society for Applied Microbiology.

Technical University of Denmark Microbiological contaminants in food in the European Union in 2004-2009

DEFF Research Database (Denmark)

Helwigh, Birgitte; Korsgaard, Helle; Grønlund, Anne Christine Jørgensen

(Salmonella, Campylobacter, Listeria, Yersinia, verotoxigenic Escherichia coli (VTEC), Brucella)and non-zoonotic microbiological contaminants (histamine, Enterobacter sakazakii and staphylococcal enterotoxins) in food to the zoonoses database at the European Food Safety Authority (EFSA). Norway...... also been excluded from the detailed analyses of the microbiological contaminants in Chapter 3. Initially, 86% of the reported samples were assumed to be representative of the occurrence of microbiological contaminants in domestically produced food. However, when samples without specific sampling unit...... and figures alone. For some foodstuffs, the occurrence of microbiological contaminants increased along the food production chain, which indicates either cross-contamination during production or microbiological growth during shelf-life. For other foodstuffs, the occurrence of microbiological contaminants...

Comparison of antimicrobial resistance in E. coli isolated from rectal and floor samples in pens with diarrhoeic nursery pigs in Denmark

DEFF Research Database (Denmark)

Weber, Nicolai Rosager; Nielsen, Jens Peter; Jorsal, Sven Erik Lind

2017-01-01

and classified as ETEC when genes for adhesin factors and enterotoxins were detected. Minimum inhibitory concentrations of 13 antimicrobial agents were determined by the broth micro dilution method. Isolates were classified as resistant based on clinical breakpoints. Results. Based on logistic regression models......%: 2.87-18.10). The odds of an isolate being resistant were significantly higher in ETEC isolates compared to Non-ETEC isolates for ampicillin (p trimethoprim (p..., sulphamethoxazole, tetracycline and trimethoprim. Conclusions. We found that ETEC isolates were more resistant than Non-ETEC isolates. Furthermore, this study indicates that resistance testing of ETEC isolates from pen floor samples can be used as a convenient sampling method for resistance testing...

Contagem de Staphylococcus sp. e detecção de enterotoxinas estafilocócicas e toxina da síndrome do choque tóxico em amostras de leite cru refrigerado Staphylococcus sp. counting and detection of staphylococcal enterotoxins and toxic shock toxin syndrome from cooled raw milk

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H.C. Lamaita

2005-10-01

Full Text Available Analisaram-se 80 amostras de leite cru refrigerado a 4°C e estocado por 48 horas em tanques refrigeradores de propriedades rurais do estado de Minas Gerais quanto à contagem e identificação de Staphylococcus sp. e detecção de enterotoxinas estafilocócicas (SE e da toxina da síndrome do choque tóxico (TSST-1. Staphylococcus sp. foi detectado em 100% das amostras de leite de tanque refrigerador em contagens que variaram de 1,0 × 10(5 a 2,5 × 10(7 UFC/ml (média = 5,60 log UFC/ml; s = 0,53 e CV = 9,5%. Isolaram-se e identificaram-se 436 estirpes como: S. aureus, S hyicus, S. epidermidis, S. intermedius, S. cohnii, S. sciuri, S. schleirferi e S. delphini. As estirpes de mesmo perfil bioquímico, oriundas da mesma amostra, foram agrupadas (pools e induzidas a produzir SE e TSST-1. A detecção dessas enterotoxinas foi feita pelo método optimum sensitivity plate, usando-se técnica de celofane sobre ágar. Identificou-se a produção de SEA, SEB, SEC, SED e de TSST-1 em percentuais variados. Dos 138 pools preparados, 91 produziram, pelo menos, uma toxina isoladamente ou em associação a outras toxinas. Dos pools enterotoxigênicos, 24,6% eram coagulase positiva e 41,3%, coagulase negativa. A confirmação de estirpes enterotoxigênicas de Staphylococcus coagulase negativa isoladas de amostras de leite é importante em relação à saúde pública.In order to count and identify Staphylococcus sp., the detection of the Staphylococcal enterotoxins (SE and toxic shock toxin syndrome (TSST-1, 80 raw milk samples cooled at 4°C and stored in bulk tanks for 48 hours in different farms from Minas Gerais State were analyzed. Staphylococcus sp. was observed in all samples and the counts varied from 1.0 × 10(5 to 2.5 × 10(7 CFU/ml (mean = 5.60 log CFU/ml; sd = 0.53 and CV = 9.46 %. A total of 436 strains of Staphylococcus were isolated and identified as S. aureus, S hyicus, S. epidermidis, S. intermedius, S. cohnii, S. sciuri, S. schleirferi and S

Pilot Scale Production and Testing of a Recombinant Staphylococcal Enterotoxin (SEB) Triple Mutant

Science.gov (United States)

2017-09-01

Polymerase Chain Reaction . J. Food Prot. 2000, 63, (4), 479–488. 6. Ulrich, R.G.; Olson, M.A.; Bavari, S. Development of Engineered Vaccines...TR-1471 9. SPONSORING / MONITORING AGENCY NAME(S) AND ADDRESS(ES) Joint Program Executive Office for Chemical and Biological Defense, Joint Project...distribution unlimited. 13. SUPPLEMENTARY NOTES 14. ABSTRACT: The use of chemically or thermally inactivated biothreat toxins in detection

Molecular analysis of the interaction between Clostridium perfringens Enterotoxin and Claudins

OpenAIRE

Protze, Jonas

2015-01-01

Claudins are essential constituents of Tight Junctions (TJs) and responsible for maintenance of these cell-cell contacts. Binding of Clostridium perfringens Enterotoxin’s C-terminal domain (cCPE) to the extracellular loop 2 (EZS2) of claudins, especially Cld3, Cld4 and Cld6-Cld9 causes a reversible opening of TJs. Thus, a structure-function analysis of this system is relevant for biomedical application, since cCPE could be used to enhance paracellular drug uptake. Furthermore cCPE respectivel...

Stress-caused Anergy of Leukocytes towards Staphylococcal enterotoxin B and Exposure Transcriptome Signatures

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2015-05-28

concerted immune depletion was highly pronounced in the case of genes involved in synthesis /secretion of chemokines and antigen presentation. (c) The 21...biosynthetic process 48545 8.03E-03 12 Response to steroid hormone stimulus 16126 9.50E-03 5 Sterol biosynthetic process 6950 1.01E-02 44 Response to

Newer insights into the mechanism of action of Psidium guajava L. leaves in infectious diarrhoea.

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Birdi, Tannaz; Daswani, Poonam; Brijesh, S; Tetali, Pundarikakshudu; Natu, Arvind; Antia, Noshir

2010-06-28

Psidium guajava L., Myrtaceae, is used widely in traditional medicine for the treatment of diarrhoea, dysentery, gastroenteritis, stomachaches, and indigestion. However, the effect of the leaf extract of P. guajava on the pathogenesis of infectious diarrhoea has not been studied. The present study evaluates the effect of a hot aqueous extract (decoction) of dried leaves of P. guajava on parameters associated with pathogenicity of infectious diarrhoea. The aim was to understand its possible mechanism(s) of action in controlling infectious diarrhoea and compare it with quercetin, one of the most reported active constituents of P. guajava with antidiarrhoeal activity. The crude decoction and quercetin were studied for their antibacterial activity and effect on virulence features of common diarrhoeal pathogens viz. colonization of epithelial cells and production and action of enterotoxins. Colonization as measured by adherence of enteropathogenic Escherichia coli (EPEC) and invasion of enteroinvasive E. coli (EIEC) and Shigella flexneri was assessed using HEp-2 cell line. The production of E. coli heat labile toxin (LT) and cholera toxin (CT) and their binding to ganglioside monosialic acid (GM1) were studied by GM1-ELISA whereas the production and action of E. coli heat stable toxin (ST) was assessed by suckling mouse assay. The decoction of P. guajava showed antibacterial activity towards S. flexneri and Vibrio cholerae. It decreased production of both LT and CT and their binding to GM1. However, it had no effect on production and action of ST. The decoction also inhibited the adherence of EPEC and invasion by both EIEC and S. flexneri to HEp-2 cells. Quercetin, on the other hand, had no antibacterial activity at the concentrations used nor did it affect any of the enterotoxins. Although it did not affect adherence of EPEC, it inhibited the invasion of both EIEC and S. flexneri to HEp-2 cells. Collectively, the results indicate that the decoction of P. guajava leaves

Turn a diarrhoea toxin into a receptor-mediated therapy for a plethora of CLDN-4-overexpressing cancers

International Nuclear Information System (INIS)

Yao, Qin; Cao, Siyu; Li, Chun; Mengesha, Asferd; Low, Pauline; Kong, Beihua; Dai, Shuzhen; Wei, Mingqian

2010-01-01

Research highlights: → CLDN-4 is the high-affinity receptor for Clostridium perfringens enterotoxin (CPE). → The targeted toxin C-CPE-ETA' utilises the C-terminal fragment of CPE for binding. → C-CPE-ETA' rapidly binds to and internalises into CLDN-4 positive cancer cells. → C-CPE-ETA' has anti-cancer ability in a range of CLDN-4 positive cancers. -- Abstract: Molecular targeted therapy (MTT) represents the new generation of anti-cancer arsenals. In this study, we report an alternative approach using a hybrid toxin that utilises the high-affinity of receptor-binding fragment of Clostridium perfringens enterotoxin (CPE). CPE naturally binds to CLDN-4 through the C-terminal 30 amino acid. However, recent studies have shown that CLDN-4 is also overexpressed on a range of cancer cells. We thus constructed a cDNA comprising C-CPE and a well characterised toxic domain of Pseudomonas aeruginosa exotoxin A (C-CPE-ETA'). The recombinant C-CPE-ETA' fusion protein was shown to retain the specificity of binding to CLDN-4 and initiating rapid penetration into cytosol in five different CLDN-4 positive cancer cells (Breast-MCF7, Skin-A431, Colon-SW480, Prostate-PC3 and DU145) but not to CLDN-4 negative cells (Hela, HUVEC). C-CPE-ETA' was strongly cytotoxic towards CLDN-4 positive cancer cell, as opposed to cells lacking CLDN-4 expression. Furthermore, we demonstrated that the recombinant fusion protein had significant anti-cancer ability in CLDN-4 positive cancer models in vivo. Subcutaneously implanted MCF7 and SW480 xenograft tumours were significantly decreased or abolished after three repeated injection of the hybrid toxin. Taken together, our results convincingly show that the hybrid toxin targets CLDN-4 positive cancer through receptor-binding, and causes significant tumour cell apoptosis, suggesting its potential as an alternative molecular targeted therapy against a plethora of CLDN-4 positive cancers.

Staphylococcus aureus CC30 Lineage and Absence of sed,j,r-Harboring Plasmid Predict Embolism in Infective Endocarditis

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Jean-Philippe Rasigade

2018-06-01

Full Text Available Staphylococcus aureus induces severe infective endocarditis (IE where embolic complications are a major cause of death. Risk factors for embolism have been reported such as a younger age or larger IE vegetations, while methicillin resistance conferred by the mecA gene appeared as a protective factor. It is unclear, however, whether embolism is influenced by other S. aureus characteristics such as clonal complex (CC or virulence pattern. We examined clinical and microbiological predictors of embolism in a prospective multicentric cohort of 98 French patients with monomicrobial S. aureus IE. The genomic contents of causative isolates were characterized using DNA array. To preserve statistical power, genotypic predictors were restricted to CC, secreted virulence factors and virulence regulators. Multivariate regularized logistic regression identified three independent predictors of embolism. Patients at higher risk were younger than the cohort median age of 62.5 y (adjusted odds ratio [OR] 0.14; 95% confidence interval [CI] 0.05–0.36. S. aureus characteristics predicting embolism were a CC30 genetic background (adjusted OR 9.734; 95% CI 1.53–192.8 and the absence of pIB485-like plasmid-borne enterotoxin-encoding genes sed, sej, and ser (sedjr; adjusted OR 0.07; 95% CI 0.004–0.457. CC30 S. aureus has been repeatedly reported to exhibit enhanced fitness in bloodstream infections, which might impact its ability to cause embolism. sedjr-encoded enterotoxins, whose superantigenic activity is unlikely to protect against embolism, possibly acted as a proxy to others genes of the pIB485-like plasmid found in genetically unrelated isolates from mostly embolism-free patients. mecA did not independently predict embolism but was strongly associated with sedjr. This mecA-sedjr association might have driven previous reports of a negative association of mecA and embolism. Collectively, our results suggest that the influence of S. aureus genotypic features

Genotypic and Phenotypic Characterization of Enterotoxigenic Escherichia coli Strains Isolated from Peruvian Children ▿

Science.gov (United States)

Rivera, F. P.; Ochoa, T. J.; Maves, R. C.; Bernal, M.; Medina, A. M.; Meza, R.; Barletta, F.; Mercado, E.; Ecker, L.; Gil, A. I.; Hall, E. R.; Huicho, L.; Lanata, C. F.

2010-01-01

Enterotoxigenic Escherichia coli (ETEC) is a major cause of childhood diarrhea. The present study sought to determine the prevalence and distribution of toxin types, colonization factors (CFs), and antimicrobial susceptibility of ETEC strains isolated from Peruvian children. We analyzed ETEC strains isolated from Peruvian children between 2 and 24 months of age in a passive surveillance study. Five E. coli colonies per patient were studied by multiplex real-time PCR to identify ETEC virulence factors. ETEC-associated toxins were confirmed using a GM1-based enzyme-linked immunosorbent assay. Confirmed strains were tested for CFs by dot blot assay using 21 monoclonal antibodies. We analyzed 1,129 samples from children with diarrhea and 744 control children and found ETEC in 5.3% and 4.3%, respectively. ETEC was more frequently isolated from children >12 months of age than from children <12 months of age (P < 0.001). Fifty-two percent of ETEC isolates from children with diarrhea and 72% of isolates from controls were heat-labile enterotoxin (LT) positive and heat-stable enterotoxin (ST) negative; 25% and 19%, respectively, were LT negative and ST positive; and 23% and 9%, respectively, were LT positive and ST positive. CFs were identified in 64% of diarrheal samples and 37% of control samples (P < 0.05). The most common CFs were CS6 (14% and 7%, respectively), CS12 (12% and 4%, respectively), and CS1 (9% and 4%, respectively). ST-producing ETEC strains caused more severe diarrhea than non-ST-producing ETEC strains. The strains were most frequently resistant to ampicillin (71%) and co-trimoxazole (61%). ETEC was thus found to be more prevalent in older infants. LT was the most common toxin type; 64% of strains had an identified CF. These data are relevant in estimating the burden of disease due to ETEC and the potential coverage of children in Peru by investigational vaccines. PMID:20631096

Using Common Spatial Distributions of Atoms to Relate Functionally Divergent Influenza Virus N10 and N11 Protein Structures to Functionally Characterized Neuraminidase Structures, Toxin Cell Entry Domains, and Non-Influenza Virus Cell Entry Domains

Science.gov (United States)

Weininger, Arthur; Weininger, Susan

2015-01-01

The ability to identify the functional correlates of structural and sequence variation in proteins is a critical capability. We related structures of influenza A N10 and N11 proteins that have no established function to structures of proteins with known function by identifying spatially conserved atoms. We identified atoms with common distributed spatial occupancy in PDB structures of N10 protein, N11 protein, an influenza A neuraminidase, an influenza B neuraminidase, and a bacterial neuraminidase. By superposing these spatially conserved atoms, we aligned the structures and associated molecules. We report spatially and sequence invariant residues in the aligned structures. Spatially invariant residues in the N6 and influenza B neuraminidase active sites were found in previously unidentified spatially equivalent sites in the N10 and N11 proteins. We found the corresponding secondary and tertiary structures of the aligned proteins to be largely identical despite significant sequence divergence. We found structural precedent in known non-neuraminidase structures for residues exhibiting structural and sequence divergence in the aligned structures. In N10 protein, we identified staphylococcal enterotoxin I-like domains. In N11 protein, we identified hepatitis E E2S-like domains, SARS spike protein-like domains, and toxin components shared by alpha-bungarotoxin, staphylococcal enterotoxin I, anthrax lethal factor, clostridium botulinum neurotoxin, and clostridium tetanus toxin. The presence of active site components common to the N6, influenza B, and S. pneumoniae neuraminidases in the N10 and N11 proteins, combined with the absence of apparent neuraminidase function, suggests that the role of neuraminidases in H17N10 and H18N11 emerging influenza A viruses may have changed. The presentation of E2S-like, SARS spike protein-like, or toxin-like domains by the N10 and N11 proteins in these emerging viruses may indicate that H17N10 and H18N11 sialidase-facilitated cell

Tracing and inhibiting growth of Staphylococcus aureus in barbecue cheese production after product recall.

Science.gov (United States)

Johler, S; Zurfluh, K; Stephan, R

2016-05-01

Staphylococcal food poisoning is one of the most prevalent causes of foodborne intoxication worldwide. It is caused by ingestion of enterotoxins formed by Staphylococcus aureus during growth in the food matrix. Following a recall of barbecue cheese due to the detection of staphylococcal enterotoxins in Switzerland in July 2015, we analyzed the production process of the respective dairy. Although most cheese-making processes involve acidification to inhibit the growth of pathogenic bacteria, barbecue cheese has to maintain a pH >6.0 to prevent undesired melting of the cheese. In addition, the dairy decided to retain the traditional manual production process of the barbecue cheese. In this study, therefore, we aimed to (1) trace Staph. aureus along the barbecue cheese production process, and (2) develop a sustainable strategy to inhibit growth of Staph. aureus and decrease the risk of staphylococcal food poisoning without changing the traditional production process. To this end, we traced Staph. aureus in a step-wise blinded process analysis on 4 different production days using spa (Staphylococcus protein A gene) typing, DNA microarray profiling, and pulsed-field gel electrophoresis analysis. We subsequently selected a new starter culture and used a model cheese production including a challenge test assay to assess its antagonistic effect on Staph. aureus growth, as well as its sensory and technological implications. We detected Staph. aureus in 30% (37/124) of the collected samples taken from the barbecue cheese production at the dairy. This included detection of Staph. aureus in the final product on all 4 production days, either after enrichment or using quantitative detection. We traced 2 enterotoxigenic Staph. aureus strains (t073/CC45 and t282/CC45) colonizing the nasal cavity and the forearms of the cheesemakers to the final product. In the challenge test assay, we were able to show that the new starter culture inhibited growth of Staph. aureus while meeting

Clinical Signs, Staphylococcus and Atopic Eczema-Related Seromarkers

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Kam Lun Hon

2017-02-01

Full Text Available Childhood eczema or atopic dermatitis (AD is a distressing disease associated with pruritus, sleep disturbance, impaired quality of life and Staphylococcus aureus isolation. The pathophysiology of AD is complex and various seromarkers of immunity are involved. We investigated if anti-staphylococcal enterotoxin IgE (anti-SE, selected seromarkers of T regulatory (Treg, T helper (Th and antigen-presenting cells (APC are associated with clinical signs of disease severity and quality of life. Disease severity was assessed with the Scoring Atopic Dermatitis (SCORAD index, and quality of life with the Children’s Dermatology Life Quality Index (CDLQI in AD patients ≤18 years old. Concentrations of anti-staphylococcus enterotoxin A and B immunoglobulin E (anti-SEA and anti-SEB, selected Treg/Th/APC chemokines, skin hydration and transepidermal water loss (TEWL were measured in these patients. Forty patients with AD [median (interquartile range age of 13.1 (7.9 years were recruited. Backward stepwise linear regression (controlling for age, personal allergic rhinitis and asthma, and other blood markers showed the serum anti-SEB level was positively associated with S. aureus and S. epidermidis isolations, objective SCORAD, clinical signs and CDLQI. TNF-α (a Th1 cytokine was positively associated with objective SCORAD (B = 4.935, p = 0.010, TGF-β (a Treg cytokine negatively with disease extent (B = −0.015, p = 0.001, IL-18 (an APC cytokine positively with disease extent (B = 0.438, p = 0.001 and with TEWL (B = 0.040, p = 0.010, and IL-23 (an APC cytokine negatively with disease extent (B = −2.812, p = 0.006 and positively with pruritus (B = 0.387, p = 0.007. Conclusions: Blood levels of anti-SEB, Th1, Treg and APC cytokines are correlated with various clinical signs of AD. AD is a systemic immunologic disease involving Staphylococcus aureus, cellular, humoral, cytokine and chemokine pathophysiology.

Prevalence and Characterization of Staphylococcus aureus Strains in the Pork Chain Supply in Chile.

Science.gov (United States)

Velasco, Valeria; Vergara, José L; Bonilla, Ana M; Muñoz, Javier; Mallea, Alejandra; Vallejos, Diego; Quezada-Aguiluz, Mario; Campos, Jorge; Rojas-García, Pedro

2018-05-01

The detection of methicillin-resistant Staphylococcus aureus (MRSA) and other emerging strains in meat-producing animals and retail meat has increased the risk of contamination of food. The aim of this study was to determine the prevalence and characterize S. aureus strains isolated from the pork chain supply in Chile. A total of 487 samples were collected: 332 samples from pigs at farms and slaughterhouses (nasal, n = 155; skin, n = 177); 85 samples from carcasses at slaughterhouses; and 70 meat samples at supermarkets and retail stores. The isolation of S. aureus was carried out by selective enrichment and culture media. Biochemical testing (API ® Staph) and PCR (detection of the nuc and mecA genes) were used to confirm S. aureus and MRSA strains. The agglutination test was used to determine the protein PBP2'. Enterotoxins (SEA, SEB, SEC, SED) were determined by agglutination test and the se genes by PCR method. Oxacillin and cefoxitin susceptibility testing were carried out using the diffusion method. The overall prevalence of S. aureus in the pork meat supply was 33.9%. A higher prevalence was detected on carcasses (56.5%), in pigs sampled at farms (40.6%) than in pigs sampled at slaughterhouses (23.3%) and in nonpackaged retail meat (43.1%) than packaged retail meat (5.3%) (p ≤ 0.05). No significant differences (p > 0.05) were found between the prevalence in pigs (28.3%) and pork meat (32.9%) and between natural pig farming (33.3%) and conventional production (52.8%). The mecA gene and the protein PBP2' were not detected in S. aureus strains. Two S. aureus strains exhibited oxacillin and cefoxitin resistance, and one S. aureus strain was resistant to cefoxitin. One S. aureus strain isolated from a meat sample was positive for enterotoxin SEB. Although the mecA gene was not detected, oxacillin-resistant and seb-producing S. aureus strains were detected, which represent a risk in the pork chain supply.

Antimicrobial resistance of Aeromonas hydrophila isolated from different food sources: A mini-review

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Deyan Stratev

2016-09-01

Full Text Available Summary: Aeromonas hydrophila is a Gram-negative, oxidase-positive, facultative, anaerobic, opportunistic aquatic pathogen. A. hydrophila produces virulence factors, such as hemolysins, aerolysins, adhesins, enterotoxins, phospholipase and lipase. In addition to isolation from aquatic sources, A. hydrophila has been isolated from meat and meat products, milk and dairy products, and vegetables. However, various studies showed that this opportunistic pathogen is resistant to commercial antibiotics. This is attributed to factors such as the indiscriminate use of antibiotics in aquaculture, plasmids or horizontal gene transfer. In this report, we highlight the occurrence, prevalence and antimicrobial resistance of A. hydrophila isolated from different food samples. The presence of antimicrobial-resistant A. hydrophila in food poses threats to public and aquatic animal health. Keywords: A. hydrophila, Antimicrobial resistance, Microbial food safety

Role of the T cell receptor ligand affinity in T cell activation by bacterial superantigens

DEFF Research Database (Denmark)

Andersen, P S; Geisler, C; Buus, S

2001-01-01

Similar to native peptide/MHC ligands, bacterial superantigens have been found to bind with low affinity to the T cell receptor (TCR). It has been hypothesized that low ligand affinity is required to allow optimal TCR signaling. To test this, we generated variants of Staphylococcus enterotoxin C3...... (SEC3) with up to a 150-fold increase in TCR affinity. By stimulating T cells with SEC3 molecules immobilized onto plastic surfaces, we demonstrate that increasing the affinity of the SEC3/TCR interaction caused a proportional increase in the ability of SEC3 to activate T cells. Thus, the potency...... correlation between ligand affinity and ligand potency indicating that it is the density of receptor-ligand complexes in the T cell contact area that determines TCR signaling strength....

Diversity and enterotoxigenicity of Staphylococcus spp. associated with domiati cheese.

Science.gov (United States)

El-Sharoud, Walid M; Spano, Giuseppe

2008-12-01

A total of 87 samples of fresh and stored Domiati cheese (an Egyptian soft cheese) were examined for the presence of Staphylococcus spp. Fifteen Staphylococcus isolates identified as S. aureus (2 isolates), S. xylosus (4), S. caprae (4), and S. chromogenes (5) were recovered from 15 cheese samples. The S. aureus isolates were resistant to penicillin G and ampicillin, and one isolate was also resistant to tetracycline. S. aureus isolates harbored classical staphylococcal enterotoxin (SE) genes (sea and seb) and recently characterized SE-like genes (selg, seli, selm, and selo). One S. aureus isolate contained a single SE gene (sea), whereas another isolate contained five SE genes (seb, selg, seli, selm, and selo). These results suggest that Domiati cheese is a source for various Staphylococcus species, including S. aureus strains that could be enterotoxigenic.

Invasive Community-Acquired Methicillin-Resistant Staphylococcus aureus in a Japanese Girl with Disseminating Multiple Organ Infection: A Case Report and Review of Japanese Pediatric Cases

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Ryuta Yonezawa

2015-01-01

Full Text Available Pediatric invasive community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA infection is very serious and occasionally fatal. This infectious disease is still a relatively rare and unfamiliar infectious disease in Japan. We report a positive outcome in a 23-month-old Japanese girl with meningitis, osteomyelitis, fasciitis, necrotizing pneumonia, urinary tract infection, and bacteremia due to CA-MRSA treated with linezolid. PCR testing of the CA-MRSA strain was positive for PVL and staphylococcal enterotoxin b and negative for ACME. SCC mec was type IVa. This case underscores the selection of effective combinations of antimicrobial agents for its treatment. We need to be aware of invasive CA-MRSA infection, which rapidly progresses with a serious clinical course, because the incidence of the disease may be increasing in Japan.

Chemical composition of fennel essential oil and its impact on Staphylococcus aureus exotoxin production.

Science.gov (United States)

Qiu, Jiazhang; Li, Hongen; Su, Hongwei; Dong, Jing; Luo, Mingjing; Wang, Jianfeng; Leng, Bingfeng; Deng, Yanhong; Liu, Juxiong; Deng, Xuming

2012-04-01

In this study, fennel oil was isolated by hydrodistillation, and the chemical composition was determined by gas chromatography/mass spectral analysis. The antimicrobial activity of fennel oil against Staphylococcus aureus was evaluated by broth microdilution. A haemolysis assay, tumour necrosis factor (TNF) release assay, western blot, and real-time reverse transcription (RT)-PCR were applied to investigate the influence of fennel oil on the production of S. aureus virulence-related exoproteins. The data show that fennel oil, which contains a high level of trans-anethole, was active against S. aureus, with MICs ranging from 64 to 256 μg/ml. Furthermore, fennel oil, when used at subinhibitory concentrations, could dose-dependently decrease the expression of S. aureus exotoxins, including α-toxin, Staphylococcal enterotoxins (SEs) and toxic shock syndrome toxin 1 (TSST-1).

Monoclonal antibodies and toxins--a perspective on function and isotype.

Science.gov (United States)

Chow, Siu-Kei; Casadevall, Arturo

2012-06-01

Antibody therapy remains the only effective treatment for toxin-mediated diseases. The development of hybridoma technology has allowed the isolation of monoclonal antibodies (mAbs) with high specificity and defined properties, and numerous mAbs have been purified and characterized for their protective efficacy against different toxins. This review summarizes the mAb studies for 6 toxins--Shiga toxin, pertussis toxin, anthrax toxin, ricin toxin, botulinum toxin, and Staphylococcal enterotoxin B (SEB)--and analyzes the prevalence of mAb functions and their isotypes. Here we show that most toxin-binding mAbs resulted from immunization are non-protective and that mAbs with potential therapeutic use are preferably characterized. Various common practices and caveats of protection studies are discussed, with the goal of providing insights for the design of future research on antibody-toxin interactions.

Therapeutic Down-Modulators of Staphylococcal Superantigen-Induced Inflammation and Toxic Shock

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Teresa Krakauer

2010-07-01

Full Text Available Staphylococcal enterotoxin B (SEB and related superantigenic toxins are potent stimulators of the immune system and cause a variety of diseases in humans, ranging from food poisoning to toxic shock. These toxins bind directly to major histocompatibility complex (MHC class II molecules on antigen-presenting cells and specific Vb regions of T-cell receptors (TCR, resulting in hyperactivation of both monocytes/macrophages and T lymphocytes. Activated host cells produce massive amounts of proinflammatory cytokines and chemokines, activating inflammation and coagulation, causing clinical symptoms that include fever, hypotension, and shock. This review summarizes the in vitro and in vivo effects of staphylococcal superantigens, the role of pivotal mediators induced by these toxins in the pathogenic mechanisms of tissue injury, and the therapeutic agents to mitigate the toxic effects of superantigens.

Staphylococcus enterotoxin A modulates interleukin 15-induced signaling and mitogenesis in human T cells

DEFF Research Database (Denmark)

Gerwien, J; Kaltoft, K; Nielsen, M

1998-01-01

the anti-mitogenic effect of SEA on cytokine-induced proliferation and the pro-mitogenic effect of PMA. In contrast, inhibitors of PP1, PP2A, protein kinase C (PKC), phosphatidyl-inositol-3-kinase (PI-3K) and mammalian target of rapamycin (mTOR) are unable to inhibit the effects of SEA. In a SEA "non...

Molecular detection of Staphylococcus aureus resistant to temperature in milk and its products

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Sutejo, Stephani Valentina Harda; Amarantini, Charis; Budiarso, Tri Yahya

2017-11-01

Contamination of Staphylococcus aureus on milk can cause intoxication and infection by Staphylococcal enterotoxin. It has nuc gene, coding thermonuclease enzyme (TNase) that is responsible for nature of resistance in the heating process. This study was conducted to identify nuc gene of as S. aureus isolated from milk and its products like ultra-high temperature, sterile milk, sweetened condensed milk, formula milk, café/milk street traders and fresh milk. Biochemical identification was conducted by using carbohydrate fermentation tests and confirmed by API Staph. Molecular confirmation by amplification of nuc gene using PCR. Based on the results of confirmation using API Staph, all isolates were confirmed as S. aureus with index determinant percentage of 97%. An amplicon product of 270 bp was gained in all isolates. It is concluded that isolate of S. aureus has nuc gene.

Nonmalignant T cells stimulate growth of T-cell lymphoma cells in the presence of bacterial toxins

DEFF Research Database (Denmark)

Woetmann, Anders; Lovato, Paola; Eriksen, Karsten W

2007-01-01

Bacterial toxins including staphylococcal enterotoxins (SEs) have been implicated in the pathogenesis of cutaneous T-cell lymphomas (CTCLs). Here, we investigate SE-mediated interactions between nonmalignant T cells and malignant T-cell lines established from skin and blood of CTCL patients....... The malignant CTCL cells express MHC class II molecules that are high-affinity receptors for SE. Although treatment with SE has no direct effect on the growth of the malignant CTCL cells, the SE-treated CTCL cells induce vigorous proliferation of the SE-responsive nonmalignant T cells. In turn, the nonmalignant...... T cells enhance proliferation of the malignant cells in an SE- and MHC class II-dependent manner. Furthermore, SE and, in addition, alloantigen presentation by malignant CTCL cells to irradiated nonmalignant CD4(+) T-cell lines also enhance proliferation of the malignant cells. The growth...

HEALTH AND HYGIENIC CONDITIONS OF EWE'S MILK PROCESSING FROM THE ASPECT OF FOOD SAFETY

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Jana Pukáčová

2010-07-01

Full Text Available Totally, 47 strains of S. aureus and 578 coagulase negative staphylococci were detected in samples from raw ewe milk. The 35 out 47 isolates of S. aureus from ewe milk were positive for the presence of staphylococcal enterotoxin genes: sea(4 %, sec (48 % a sed (48 %. Staphylococcus epidermis (33.04%, Staphylococcus caprae (21.28% were more prevalent. Staphylococcus chromogenes (7.44 %, Staphylococcus hominis (7.09%, Staphylococcus xylosus (6,92 %, a Staphylococcus warneri (6.40 % were isolated also in ewes milk. Staphylococcus haemolyticus (3.11 %, Staphylococcus capitis (2.94 %, Staphylococcus simulans (2.08 % and Staphylococcus saprophyticus (1.73 % were isolated very rarely from the taken individual milk ewe samples. Sporadically, only in few cases, the others coagulase negative staphylococci were isolated (Staphylococcus cohnii cohnii, Staphylococcus sciuri, Staphylococcus closii, Staphylococcus lugdunensis, Staphylococcus auricularis and Staphylococcus equorum.   doi:10.5219/24

Detection of Different Genotypes of Clostridium perfringens in Feces of Healthy Dairy Cattle from China using Real-Time Duplex PCR Assay

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Guanghua Wang, Jizhang Zhou, Fuying Zheng, Guozhen Lin, Xiaoan Cao, Xiaowei Gong and Changqing Qiu*

2011-04-01

Full Text Available Dual-labeled fluorescence hybridization probe-based multiplex quantitative real-time polymerase chain reaction (qPCR assay was used for the detection of Clostridium perfringens toxin genes alpha (cpa, beta (cpb, iota (ia, epsilon (etx, beta2 (cpb2 and enterotoxin (cpe directly from the feces of cattle. Fecal samples from 261 lactating cattle, belonging to three dairy herds in Ningxia (China, were examined using the developed assays. The duplex qPCR assay revealed that cpa, etx, cpb2 and cpe toxin genes were detected in 176 (100%, 15 (8.5%, 142 (80.7% and 4 (2.3% of 176 PCR positive samples, respectively. The findings of this study revealed that C. perfringens beta2-toxin-producing strains were widely prevalent in lactating cows in Ningxia, possibly playing an important role in C. perfringens-associated diarrheal disease.

Automated 5 ' nuclease assay for detection of virulence factors in porcine Escherichia coli

DEFF Research Database (Denmark)

Frydendahl, K.; Imberechts, H.; Lehmann, S.

2001-01-01

(STa, STb, EAST1) and heat labile LT) enterotoxins and the verocytotoxin variant 2e (VT2e). To correctly identify false negative results, an endogenous internal control targeting the E. coil 16S rRNA gene was incorporated in each test tube. The assay was evaluated using a collection of E. coil...... reference strains which have previously been examined with phenotypical assays or DNA hybridization. Furthermore, the assay was evaluated by testing porcine E. coil field strains, previously characterized. The 5' nuclease assay correctly detected the presence of virulence genes in all reference strains....... When testing field strains there was generally excellent agreement with results obtained by laboratories in Belgium and Germany. In conclusion, the 5' nuclease assay developed is a fast and specific tool for detection of E. coli virulence genes in the veterinary diagnostic laboratory....

Monoclonal Antibodies and Toxins—A Perspective on Function and Isotype

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Siu-Kei Chow

2012-06-01

Full Text Available Antibody therapy remains the only effective treatment for toxin-mediated diseases. The development of hybridoma technology has allowed the isolation of monoclonal antibodies (mAbs with high specificity and defined properties, and numerous mAbs have been purified and characterized for their protective efficacy against different toxins. This review summarizes the mAb studies for 6 toxins—Shiga toxin, pertussis toxin, anthrax toxin, ricin toxin, botulinum toxin, and Staphylococcal enterotoxin B (SEB—and analyzes the prevalence of mAb functions and their isotypes. Here we show that most toxin-binding mAbs resulted from immunization are non-protective and that mAbs with potential therapeutic use are preferably characterized. Various common practices and caveats of protection studies are discussed, with the goal of providing insights for the design of future research on antibody-toxin interactions.

Monoclonal Antibodies and Toxins—A Perspective on Function and Isotype

Science.gov (United States)

Chow, Siu-Kei; Casadevall, Arturo

2012-01-01

Antibody therapy remains the only effective treatment for toxin-mediated diseases. The development of hybridoma technology has allowed the isolation of monoclonal antibodies (mAbs) with high specificity and defined properties, and numerous mAbs have been purified and characterized for their protective efficacy against different toxins. This review summarizes the mAb studies for 6 toxins—Shiga toxin, pertussis toxin, anthrax toxin, ricin toxin, botulinum toxin, and Staphylococcal enterotoxin B (SEB)—and analyzes the prevalence of mAb functions and their isotypes. Here we show that most toxin-binding mAbs resulted from immunization are non-protective and that mAbs with potential therapeutic use are preferably characterized. Various common practices and caveats of protection studies are discussed, with the goal of providing insights for the design of future research on antibody-toxin interactions. PMID:22822456

Molecular typing of Clostridium perfringens isolated from swine in slaughterhouses from São Paulo State, Brazil Tipagem molecular de Clostridium perfringens isolados de suínos em abatedouros do estado de São Paulo, Brasil

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Thais Sebastiana Porfida Ferreira

2012-08-01

Full Text Available Clostridium perfringens is an anaerobic Gram-positive bacterium known as common pathogen for humans, for domestic and wildlife animals. Although infections caused by C. perfringens type C and A in swine are well studied, just a few reports describe the genetic relationship among strains in the epidemiological chain of swine clostridioses, as well as the presence of the microorganism in the slaughterhouses. The aim of the present study was to isolate C. perfringens from feces and carcasses from swine slaughterhouses, characterize the strains in relation to the presence of enterotoxin, alpha, beta, epsilon, iota and beta-2 toxins genes, using polymerase chain reaction (PCR and comparing strains by means of Pulsed field gel electrophoresis (PFGE. Clostridium perfringens isolation frequencies in carcasses and finishing pig intestines were of 58.8% in both types of samples. According to the polymerase chain reaction assay, only alfa toxin was detected, being all isolates also negative to enterotoxin and beta2 toxin. Through PFGE technique, the strains were characterized in 35 pulsotypes. In only one pulsotype, the isolate from carcass sample was grouped with fecal isolate of the same animal, suggesting that the risk of cross-contamination was low. Despite the high prevalence of C. perfringens in swine carcasses from the slaughterhouses assessed, the risk of food poisoning to Brazilian pork consumers is low, since all strains were negative to cpe-gene, codifying enterotoxin.Clostridium perfringens é uma bactéria Gram positiva anaeróbica, conhecida por infectar os seres humanos, animais domésticos e de vida selvagem. Apesar de as infecções causadas por C. perfringens tipo C e A em suínos serem bastante estudadas, poucos relatos descrevem a relação genética entre as linhagens envolvidas na cadeia epidemiológica da clostridiose suína, bem como a presença do microorganismo em abatedouros. O objetivo do presente estudo foi isolar C

Inhibition of Biological Activity of Staphylococcal Enterotoxin A (SEA) by Apple Juice and Apple Polyphenols

Science.gov (United States)

The foodborne pathogen Staphylococcus aureus produces the virulent staphylococcal entertoxin A (SEA), a single-chain protein that consists of 233 amino acid residues with a molecular weight of 27 078 Da. SEA is a superantigen that is reported to contribute to animal (mastitis) and human (emesis, dia...

Staphylococcus aureus entrance into the dairy chain: Tracking S. aureus from dairy cow to cheese

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Judith Kümmel

2016-10-01

Full Text Available Staphylococcus aureus is one of the most important contagious mastitis pathogens in dairy cattle. Due to its zoonotic potential, control of S. aureus is not only of great economic importance in the dairy industry but also a significant public health concern. The aim of this study was to decipher the potential of bovine udder associated S. aureus as reservoir for S. aureus contamination in dairy production and processing. From 18 farms, delivering their milk to an alpine dairy plant for the production of smeared semi-hard and hard cheese. 1176 quarter milk (QM samples of all cows in lactation (n = 294 and representative samples form bulk tank milk (BTM of all farms were surveyed for coagulase positive (CPS and coagulase negative Staphylococci (CNS. Furthermore, samples from different steps of the cheese manufacturing process were tested for CPS and CNS. As revealed by chemometric-assisted FTIR spectroscopy and molecular subtyping (spa typing and multi locus sequence typing, dairy cattle represent indeed an important, yet underreported, entrance point of S. aureus into the dairy chain. Our data clearly show that certain S. aureus subtypes are present in primary production as well as in the cheese processing at the dairy plant. However, although a considerable diversity of S. aureus subtypes was observed in QM and BTM at the farms, only certain S. aureus subtypes were able to enter and persist in the cheese manufacturing at the dairy plant and could be isolated from cheese until day fourteen of ripening. Farm strains belonging to the FTIR cluster B1 and B3, which show genetic characteristics (t2953, ST8, enterotoxin profile: sea/sed/sej of the recently described S. aureus genotype B, most successfully contaminated the cheese production at the dairy plant. Thus our study fosters the hypothesis that genotype B S. aureus represent a specific challenge in control of S. aureus in the dairy chain that requires effective clearance strategies and hygienic

The bacteriological quality of goat and ovine milk

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Kateřina Bogdanovičová

2015-05-01

Full Text Available This study concentrates on information concerning the microbiological hazards that can be present in raw milk from animal species other than cows. A total of 54 (23 of ovine and 31 of goat bulk tank milk samples from 10 farms in the Czech Republic were collected in years 2013 - 2014. The sampling was done at regular time intervals during the whole year, with five to eight samples collected from each of the 10 dairy farms involved in the study. All milk samples were collected into sterile sampling bottles and transported in a cooler sampling case to the laboratory for immediate examination. Farms were randomly selected to cover the whole area of the Czech Republic. The prevalence and characteristic of Escherichia coli, Staphylococcus aureus, Salmonella spp., Campylobacter spp. and Listeria monocytogenes was studied. Raw cow's milk can be contaminated by E. coli intramammarily during clinical or subclinical mastitis and either directly through animal feces or indirectly during milk collection through farm employees or the milking equipment. E. coli was detected in 90.3% of the goat milk and 95.7% of the ovine milk samples. The genes encoding Shiga toxins 1 and 2- (stx1, stx2 were not detected and no STEC was identified. The Eae was the detected in 3 (4.6% isolates. S. aureus was detected in 9 (29.0% samples of goat milk and 8 (34.8% samples of ovine milk. A total 12 (57.1% enterotoxin positive S. aureus were obtained; 6 (28.6% were positive for the production of sec encoding enterotoxin SEC; in 4 (19.0% isolates the gene seh was detected; 2 (9.5% isolates were proven positive for seg (4.8% and combination seg and sei (4.8%. The presence of MRSA was not detected in the tested samples in our study. L. monocytogenes was detected in 1 (3.2% samples of goat milk and 1 (4.3% samples of ovine milk. The serotype (1/2a, 1/2b was detected in our study. Campylobacter spp. and Salmonella spp. were not isolated from any of the samples. These results form

Detection and Characterization of Staphylococcus aureus and Methicillin-Resistant S. aureus in Foods Confiscated in EU Borders

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David Rodríguez-Lázaro

2017-07-01

Full Text Available The aim of the study was to evaluate the potential role of the illegal entry of food in UE in the Methicillin-resistant S. aureus (MRSA spread. We studied the prevalence and characteristics of Staphylococcus aureus and MRSA isolated from foods of animal origin confiscated from passengers on flights from 45 non-EU countries from 2012 to 2015 by the Border Authorities at Bilbao International Airport (Spain and Vienna International Airport (Austria, as well as foods from open markets close to EU land borders. Of 868 food samples tested (diverse meat samples including antelope, duck, guinea pig, pork, rodents, turkey, dairy products, and eggs, 136 (15.7% were positive for S. aureus and 26 (3.0% for MRSA. All MRSA strains were mecA-positive. The prevalence of S. aureus-positive dairy samples among food confiscated at Bilbao International Airport was 64.6%, and this airport also had the highest value (11.8% for MRSA-positive samples. The predominant sequence type was ST5 (30.8%, followed by ST8, ST1649, ST1, and other lineages were found to a lesser extent (ST7, ST22, ST72, ST97, and ST398. Six isolates tested positive for luk-PVL genes (SCCmec IV subtypes IVc and IVe. Enterotoxin profiling revealed that 19 MRSA strains were enterotoxigenic, harboring one or more se genes. The MRSA isolates positive for luk-PVL genes were not enterotoxigenic, and none of the isolates tested positive for enterotoxin E. We found 14 resistance profiles, and more than 69% of the MRSA isolates were resistant to three or more types of antimicrobial agents. This finding reveals both the wide diversity of the antimicrobial resistance found in the strains and the capacity to resist not only to beta-lactam drugs. One MRSA strain showed unusual characteristics: it was oxacillin-susceptible, harbored SCCmec V, and was positive for sed, seg, and sej but negative for PVL virulence factors. This study shows the presence of enterotoxigenic HA-, CA-, and LA-MRSA in foods illegally

Molecular Characterization of Methicillin-Resistant Staphylococcus aureus from Outpatients in Northern Japan: Increasing Tendency of ST5/ST764 MRSA-IIa with Arginine Catabolic Mobile Element.

Science.gov (United States)

Aung, Meiji Soe; Kawaguchiya, Mitsuyo; Urushibara, Noriko; Sumi, Ayako; Ito, Masahiko; Kudo, Kenji; Morimoto, Shigeo; Hosoya, Shino; Kobayashi, Nobumichi

2017-07-01

Arginine catabolic mobile element (ACME) is a genomic island of staphylococcus and is considered to confer enhanced ability to survive and growth on host bacterial cells. ACME has been typically identified in Panton-Valentine Leukocidin (PVL)-positive ST8 methicillin-resistant Staphylococcus aureus (MRSA) with SCCmec type IVa (USA300 clone), and it is also found in other lineages at low frequency. Prevalence and molecular characteristics of PVL + and/or ACME + MRSA were investigated for 624 clinical isolates collected from outpatients in northern Japan from 2013 to 2014. Both PVL genes and ACME type I were detected in nine isolates (1.4%), which were ST8-MRSA-SCCmec IVa/spa type t008/agr-I; whereas solely PVL genes were positive in two isolates, ST30-MRSA-SCCmec IV and ST59-MRSA-SCCmec V. ACME type II' (previously referred to as ACME ΔII) was detected in 36 isolates (5.8%) with SCCmec II and V (32 and 4 isolates, respectively), exhibiting an increased rate within SCCmec II-MRSA (7.1%) compared with our previous studies (0.86-4.5%, 2008-2011). ACME II'-positive MRSA strains were classified into ST5-SCCmec IIa/V or ST764-SCCmec IIa belonging to five different spa types, with t002 being dominant. They harbored mostly enterotoxin gene clusters (seg-sei-sem-sen-seo-seu) and some more enterotoxin genes (seb1, seb2, sec3, sel, sep), showing resistance to more antimicrobials than ST8-MRSA-SCCmec IVa. ACME-SCCmec composite island (CI) of the 36 ACME II'-positive MRSA was classified into five types (ii)-(vi), among which type (ii) (orfX-ΨSCC ΔJ1 SCCmec I -ACME II'-SCCmec II) was dominant and subdivided into the A3 variant and the less common A2 variant. CI types (v) and (vi) were considered novel genetic organizations having speG (acetyltransferase genes for polyamines) in inserted SCC4610/SCC266-like genetic elements. The present study revealed increased prevalence and genetic diversity of the ST5/ST764-MRSA-SCCmec II with ACME II' in northern Japan.

Efficient protein immobilization on polyethersolfone electrospun nanofibrous membrane via covalent binding for biosensing applications

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Mahmoudifard, Matin [Institute for Nanoscience and Nanotechnology, Sharif University of Technology, Tehran (Iran, Islamic Republic of); Soudi, Sara [Stem Cell Biology Department, Stem Cell Technology Research Center, Tehran (Iran, Islamic Republic of); Department of Immunology, Faculty of Medical Sciences, Tarbiat Modares University, Tehran (Iran, Islamic Republic of); Soleimani, Masoud [Department of Hematology, Faculty of Medical Sciences, Tarbiat Modares University, Tehran (Iran, Islamic Republic of); Hosseinzadeh, Simzar [Nanotechnology and Tissue Engineering Department, Stem Cell Technology Research Center, Tehran (Iran, Islamic Republic of); School of Advanced Medical Technologies, Tehran University of Medical Sciences, Tehran (Iran, Islamic Republic of); Esmaeili, Elaheh [Institute for Nanoscience and Nanotechnology, Sharif University of Technology, Tehran (Iran, Islamic Republic of); Department of Hematology, Faculty of Medical Sciences, Tarbiat Modares University, Tehran (Iran, Islamic Republic of); Vossoughi, Manouchehr, E-mail: vosoughi@sharif.edu [Institute for Nanoscience and Nanotechnology, Sharif University of Technology, Tehran (Iran, Islamic Republic of); Chemical and Petroleum Engineering Department, Sharif University of Technology, Tehran (Iran, Islamic Republic of)

2016-01-01

In this paper we introduce novel strategy for antibody immobilization using high surface area electrospun nanofibrous membrane based on ethyl-3-(3-dimethylaminopropyl)-carbodiimide/N-hydroxysuccinimide (EDC/NHS) coupling chemistry. To present the high performance of proposed biosensors, anti-staphylococcus enterotoxin B (anti-SEB) was used as a model to demonstrate the utility of our proposed system. Polymer solution of polyethersolfone was used to fabricate fine nanofibrous membrane. Moreover, industrial polyvinylidene fluoride membrane and conventional microtiter plate were also used to compare the efficiency of antibody immobilization. Scanning electron microscopy images were taken to study the morphology of the membranes. The surface activation of nanofibrous membrane was done with the help of O{sub 2} plasma. PES nanofibrous membrane with carboxyl functional groups for covalent attachment of antibodies were treated by EDC/NHS coupling agent. The quantity of antibody immobilization was measured by enzyme-linked immuno sorbent assay (ELISA) method. Attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR) spectroscopy was performed to confirm the covalent immobilization of antibody on membrane. Atomic force microscopy, scanning electron microscopy and invert fluorescence microscopy were used to analyze the antibody distribution pattern on solid surfaces. Results show that oxygen plasma treatment effectively increased the amount of antibody immobilization through EDC/NHS coupling chemistry. It was found that the use of nanofibrous membrane causes the improved detection signal of ELISA based biosensors in comparison to the standard assay carried out in the 96-well microtiter plate. This method has the potential to improve the ELISA-based biosensor and we believe that this technique can be used in various biosensing methods. - Highlights: • Introduction of novel strategy for antibody immobilization using high surface area electrospun

Newer insights into the mechanism of action of Psidium guajava L. leaves in infectious diarrhoea

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Natu Arvind

2010-06-01

Full Text Available Abstract Background Psidium guajava L., Myrtaceae, is used widely in traditional medicine for the treatment of diarrhoea, dysentery, gastroenteritis, stomachaches, and indigestion. However, the effect of the leaf extract of P. guajava on the pathogenesis of infectious diarrhoea has not been studied. The present study evaluates the effect of a hot aqueous extract (decoction of dried leaves of P. guajava on parameters associated with pathogenicity of infectious diarrhoea. The aim was to understand its possible mechanism(s of action in controlling infectious diarrhoea and compare it with quercetin, one of the most reported active constituents of P. guajava with antidiarrhoeal activity. Methods The crude decoction and quercetin were studied for their antibacterial activity and effect on virulence features of common diarrhoeal pathogens viz. colonization of epithelial cells and production and action of enterotoxins. Colonization as measured by adherence of enteropathogenic Escherichia coli (EPEC and invasion of enteroinvasive E. coli (EIEC and Shigella flexneri was assessed using HEp-2 cell line. The production of E. coli heat labile toxin (LT and cholera toxin (CT and their binding to ganglioside monosialic acid (GM1 were studied by GM1-ELISA whereas the production and action of E. coli heat stable toxin (ST was assessed by suckling mouse assay. Results The decoction of P. guajava showed antibacterial activity towards S. flexneri and Vibrio cholerae. It decreased production of both LT and CT and their binding to GM1. However, it had no effect on production and action of ST. The decoction also inhibited the adherence of EPEC and invasion by both EIEC and S. flexneri to HEp-2 cells. Quercetin, on the other hand, had no antibacterial activity at the concentrations used nor did it affect any of the enterotoxins. Although it did not affect adherence of EPEC, it inhibited the invasion of both EIEC and S. flexneri to HEp-2 cells. Conclusion Collectively

Crystallization and preliminary crystallographic analysis of the NheA component of the Nhe toxin from Bacillus cereus

International Nuclear Information System (INIS)

Phung, Danh; Ganash, Magdah; Sedelnikova, Svetlana E.; Lindbäck, Toril; Granum, Per Einar; Artymiuk, Peter J.

2012-01-01

The NheA component of the B. cereus Nhe toxin was overexpressed in E. coli, purified and crystallized. Diffraction data were collected and processed to 2.05 Å resolution. The nonhaemolytic enterotoxin (Nhe) of Bacillus cereus plays a key role in cases of B. cereus food poisoning. The toxin is comprised of three different proteins: NheA, NheB and NheC. Here, the expression in Escherichia coli, purification and crystallization of the NheA protein are reported. The protein was crystallized by the sitting-drop vapour-diffusion method using PEG 3350 as a precipitant. The crystals of NheA diffracted to 2.05 Å resolution and belonged to space group C2, with unit-cell parameters a = 308.7, b = 58.2, c = 172.9 Å, β = 110.6°. Calculation of V M values suggests that there are approximately eight protein molecules per asymmetric unit

Study of the enterotoxigenicity of enterobacteria and their resistance to antibiotics in postirradiation abacteriosis

International Nuclear Information System (INIS)

Korshunov, V.M.; Bodryagina, A.B.; Pinegin, B.V.

1981-01-01

Qualitative and quantitative composition of Enterobacteriaceal family was studied and the percentage of enterotoxigenic and antibiotic resistant strains was determined in the large intestine cavity of intact mice of CBA line as well as in the cavity and wall of the small intestine of those animals on the 7 day after γ-irradiation (900 R). It was ascertained that postirradiation abacteriosis was characterized by the penetration in quantity of Esub(coli,Pr. morganii) into the small intestine and by the emergence in the small intestine of non-determinable in the intestinal tract intact animals: Pr. vulgaris, Pr. mirabilis, Pr. rettgeri, Ent. aerogenes, Ent. cloacae, Ent. hafniae, Citrobacter freundii, Providencia alcalifaciens; all revealed representatives of enterobacteria were capable of establishing associative bond with the mucous membrane of the intestine. Enterobacteria synthesizing resistant to antibiotics enterotoxins were revealed in quantity in the wall and lumen of the small intestine of irradiated mice [ru

Toxin genotyping of Clostridium perfringens field strains isolated from healthy and diseased chickens

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Luca Bano

2010-01-01

Full Text Available Clostridium perfringens is well known as the aetiological agent of necrotic enteritis in chicken. Type A and type C are considered the C. perfringens toxin types responsible for this disease. The aim of this study was to determine the presence of genes coding for α, β, ε, ι, β2 and enterotoxin in C. perfringens field strains collected from healthy and diseased chickens. Thirty-seven C. perfringens field strains were toxin typed: all strains resulted to be toxin type A and 3 of these tested positive for the presence of the toxin β2 coding gene. Four isolates showed the cpa gene with the insertion of a group II intron. Our findings confirm the most recent results reported from different countries and the data suggest that the role of C. perfringens type C should be revaluated in the etiopathogenesis of necrotic enteritis.

Production of vaccines for treatment of infectious diseases by transgenic plants

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Kristina LEDL

2016-04-01

Full Text Available Since the first pathogen antigen was expressed in transgenic plants with the aim of producing edible vaccine in early 1990s, transgenic plants have become a well-established expression system for production of alternative vaccines against various human and animal infectious diseases. The main focus of plant expression systems in the last five years has been on improving expression of well-studied antigens such as porcine reproductive and respiratory syndrome (PRRSV, bovine viral diarrhea disease virus (BVDV, footh and mouth disease virus (FMDV, hepatitis B surface antigen (HBsAg, rabies G protein, rotavirus, Newcastle disease virus (NDV, Norwalk virus capsid protein (NVCP, avian influenza virus H5N1, Escherichia coli heat-labile enterotoxin subunit B (LT-B, cholera toxin B (CT-B, human immunodeficiency virus (HIV, artherosclerosis, ebola and anthrax. Significant increases in expression have been obtained using improved expression vectors, different plant species and transformation methods.

Yersinia enterocolitica: Mode of Transmission, Molecular Insights of Virulence, and Pathogenesis of Infection

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Yeasmin Sabina

2011-01-01

Full Text Available Although Yersinia enterocolitica is usually transmitted through contaminated food and untreated water, occasional transmission such as human-to-human, animal-to-human and blood transfusion associated transmission have also identified in human disease. Of the six Y. enterocolitica biotypes, the virulence of the pathogenic biotypes, namely, 1B and 2–5 is attributed to the presence of a highly conserved 70-kb virulence plasmid, termed pYV/pCD and certain chromosomal genes. Some biotype 1A strains, despite lacking virulence plasmid (pYV and traditional chromosomal virulence genes, are isolated frequently from humans with gastrointestinal diseases similar to that produced by isolates belonging known pathogenic biotypes. Y. enterocolitica pathogenic biotypes have evolved two major properties: the ability to penetrate the intestinal wall, which is thought to be controlled by plasmid genes, and the production of heat-stable enterotoxin, which is controlled by chromosomal genes.

Dietary n-3 PUFAs augment caspase 8 activation in Staphylococcal aureus enterotoxin B stimulated T-cells

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Gill, R. [Department of Immunology and Microbiology, Wayne State University, Detroit, MI (United States); Jen, K.L. [Department of Nutrition and Food Science, Wayne State University, Detroit, MI (United States); Center for Urban Responses to Environmental Stressors (CURES), Wayne State University, Detroit, MI (United States); McCabe, M.J.J. [Department of Environmental Medicine, University of Rochester, Rochester, NY (United States); Rosenspire, A., E-mail: arosenspire@wayne.edu [Department of Immunology and Microbiology, Wayne State University, Detroit, MI (United States); Center for Urban Responses to Environmental Stressors (CURES), Wayne State University, Detroit, MI (United States)

2016-10-15

Epidemiological studies have linked consumption of n-3 PUFAs with a variety of beneficial health benefits, particularly with respect to putative anti-inflammatory effects. Unfortunately, many of these results remain somewhat controversial because in most instances there has not been a linkage to specific molecular mechanisms. For instance, dietary exposure to low levels of mercury has been shown to be damaging to neural development, but concomitant ingestion of n-3 PUFAs as occurs during consumption of fish, has been shown to counteract the detrimental effects. As the mechanisms mediating the neurotoxicity of environmental mercury are not fully delineated, it is difficult to conceptualize a testable molecular mechanism explaining how n-3 PUFAs negate its neurotoxic effects. However, environmental exposure to mercury also has been linked to increased autoimmunity. By way of a molecular understanding of this immuno-toxic association, disruption of CD95 signaling is well established as a triggering factor for autoimmunity, and we have previously shown that environmentally relevant in vitro and dietary exposures to mercury interfere with CD95 signaling. In particular we have shown that activation of caspase 8, as well as downstream activation of caspase 3, in response to CD95 agonist stimulation is depressed by mercury. More recently we have shown in vitro that the n-3 PUFA docosahexaenoic acid counteracts the negative effect of mercury on CD95 signaling by restoring caspase activity. We hypothesized that concomitant ingestion of n-3 PUFAs with mercury might be protective from the immuno-toxic effects of mercury, as it is with mercury's neuro-toxic effects, and in the case of immuno-toxicity this would be related to restoration of CD95 signal strength. We now show that dietary ingestion of n-3 PUFAs generally promotes CD95 signaling by upregulating caspase 8 activation. Apart from accounting for the ability of n-3 PUFAs to specifically counteract autoimmune sequelae of mercury exposure, this novel finding for the first time suggests a testable molecular mechanism explaining the overall anti-inflammatory properties of n-3 PUFAs. - Highlights: • Dietary n-3 PUFAs counter Hg{sup 2+} immunotoxicity • Hg{sup 2+} interference with SEB-mediated signal transduction is ameliorated by n-3 PUFA rich diets. • Dietary n-3 PUFAs augment SEB-mediated activation of caspase 8 in vivo.

Staphylococcal enterotoxin-A directly stimulates signal transduction and interferon-gamma production in psoriatic T-cell lines

DEFF Research Database (Denmark)

Nielsen, M B; Odum, N; Gerwien, J

1998-01-01

class II. Here we address the question of whether SEA can directly activate psoriatic T cells in the absence of professional antigen-presenting cells. We show that SEA induces i) tyrosine phosphorylation of several proteins, ii) downregulation of the T-cell receptor (TCR), and iii) production......-mediated proliferation. In contrast, SEA with a mutation in the MHC class II alpha-binding site induces IFN-gamma and a qualitatively changed tyrosine phosphorylation profile. Both mutations delete the co-stimulatory effect on cytokine-mediated proliferation. This suggests that both MHC class II binding sites...

Dietary n-3 PUFAs augment caspase 8 activation in Staphylococcal aureus enterotoxin B stimulated T-cells

International Nuclear Information System (INIS)

Gill, R.; Jen, K.L.; McCabe, M.J.J.; Rosenspire, A.

2016-01-01

Epidemiological studies have linked consumption of n-3 PUFAs with a variety of beneficial health benefits, particularly with respect to putative anti-inflammatory effects. Unfortunately, many of these results remain somewhat controversial because in most instances there has not been a linkage to specific molecular mechanisms. For instance, dietary exposure to low levels of mercury has been shown to be damaging to neural development, but concomitant ingestion of n-3 PUFAs as occurs during consumption of fish, has been shown to counteract the detrimental effects. As the mechanisms mediating the neurotoxicity of environmental mercury are not fully delineated, it is difficult to conceptualize a testable molecular mechanism explaining how n-3 PUFAs negate its neurotoxic effects. However, environmental exposure to mercury also has been linked to increased autoimmunity. By way of a molecular understanding of this immuno-toxic association, disruption of CD95 signaling is well established as a triggering factor for autoimmunity, and we have previously shown that environmentally relevant in vitro and dietary exposures to mercury interfere with CD95 signaling. In particular we have shown that activation of caspase 8, as well as downstream activation of caspase 3, in response to CD95 agonist stimulation is depressed by mercury. More recently we have shown in vitro that the n-3 PUFA docosahexaenoic acid counteracts the negative effect of mercury on CD95 signaling by restoring caspase activity. We hypothesized that concomitant ingestion of n-3 PUFAs with mercury might be protective from the immuno-toxic effects of mercury, as it is with mercury's neuro-toxic effects, and in the case of immuno-toxicity this would be related to restoration of CD95 signal strength. We now show that dietary ingestion of n-3 PUFAs generally promotes CD95 signaling by upregulating caspase 8 activation. Apart from accounting for the ability of n-3 PUFAs to specifically counteract autoimmune sequelae of mercury exposure, this novel finding for the first time suggests a testable molecular mechanism explaining the overall anti-inflammatory properties of n-3 PUFAs. - Highlights: • Dietary n-3 PUFAs counter Hg 2+ immunotoxicity • Hg 2+ interference with SEB-mediated signal transduction is ameliorated by n-3 PUFA rich diets. • Dietary n-3 PUFAs augment SEB-mediated activation of caspase 8 in vivo

Bacillus anthracis Edema Toxin Inhibits Staphylococcus aureus Enterotoxin B Effects in Vitro: A Potential Protein Therapeutic?

Science.gov (United States)

2005-10-01

5). Inherent characteristics of edema toxin and other procaryotic adenylate cyclases from Bordetella pertussis, Pseudomonas aeruginosa, and Yersinia...by mouse peritoneal macrophages: the role of cellular cyclic AMP. Immunology 64:719–724. 12. Krakauer, T. 1999. Induction of CC chemokines in human

Inhibition of toxicogenic Bacillus cereus in rice-based foods by enterocin AS-48.

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Grande, Maria J; Lucas, Rosario; Abriouel, Hikmate; Valdivia, Eva; Omar, Nabil Ben; Maqueda, Mercedes; Martínez-Bueno, Manuel; Martínez-Cañamero, Magdalena; Gálvez, Antonio

2006-02-01

The antimicrobial effect of the broad-spectrum bacteriocin enterocin AS-48 against the toxicogenic psychrotrophic strain Bacillus cereus LWL1 has been investigated in a model food system consisting of boiled rice and in a commercial infant rice-based gruel dissolved in whole milk stored at temperatures of 37 degrees C, 15 degrees C and 6 degrees C. In food samples supplemented with enterocin AS-48 (in a concentration range of 20-35 mug/ml), viable cell counts decreased rapidly over incubation time, depending on the bacteriocin concentration, the temperature of incubation and the food sample. Enterotoxin production at 37 degrees C was also inhibited. Heat sensitivity of endospores increased markedly in food samples supplemented with enterocin AS-48: inactivation of endospores was achieved by heating for 1 min at 90 degrees C in boiled rice or at 95 degrees C in rice-based gruel. Activity of enterocin AS-48 in rice gruel was potentiated by sodium lactate in a concentration-dependent way.

Phenotypic and Genotypic Characterization of Staphylococcus aureus Strains from Italian Dairy Products

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Stefano Morandi

2009-01-01

Full Text Available Staphylococcus aureus is a known major cause of foodborne illnesses, and milk and dairy products are often contaminated by enterotoxigenic strains of this bacterium. In the present study, 122 S. aureus isolates collected from different dairy products were characterised by phenotypic properties, by the distribution of genes encoding staphylococcal enterotoxins (sea, sec, sed, seg, seh, sei, sej, and sel and by randomly amplified polymorphic DNA PCR (RAPD-PCR. Moreover, strain resistance to vancomycin and methicillin (oxacillin was studied. The differences in the RAPD-PCR profiles obtained with the primers M13 and AP4 revealed the presence of a great genetic heterogeneity among the different S. aureus strains. Using the primer AP4 and M13, eight groups were distinguished by RAPD-PCR cluster analysis, although, except in few cases, it was not possible to correlate the isolates of different animal species (cow or ovine with the presence of se genes. None of the isolates showed resistance to vancomycin or methicillin.

Superantigens Are Critical for Staphylococcus aureus Infective Endocarditis, Sepsis, and Acute Kidney Injury

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Salgado-Pabón, Wilmara; Breshears, Laura; Spaulding, Adam R.; Merriman, Joseph A.; Stach, Christopher S.; Horswill, Alexander R.; Peterson, Marnie L.; Schlievert, Patrick M.

2013-01-01

ABSTRACT Infective endocarditis and kidney infections are serious complications of Staphylococcus aureus sepsis. We investigated the role of superantigens (SAgs) in the development of lethal sepsis, infective endocarditis, and kidney infections. SAgs cause toxic shock syndrome, but it is unclear if SAgs contribute to infective endocarditis and kidney infections secondary to sepsis. We show in the methicillin-resistant S. aureus strain MW2 that lethal sepsis, infective endocarditis, and kidney infections in rabbits are critically dependent on high-level SAgs. In contrast, the isogenic strain lacking staphylococcal enterotoxin C (SEC), the major SAg in this strain, is attenuated in virulence, while complementation restores disease production. SAgs’ role in infective endocarditis appears to be both superantigenicity and direct endothelial cell stimulation. Maintenance of elevated blood pressure by fluid therapy significantly protects from infective endocarditis, possibly through preventing bacterial accumulation on valves and increased SAg elimination. These data should facilitate better methods to manage these serious illnesses. PMID:23963178

Optical Properties and Immunoassay Applications of Noble Metal Nanoparticles

International Nuclear Information System (INIS)

Zhu, S.; Zhou, W.

2010-01-01

Noble metal, especially gold (Au) and silver (Ag) nanoparticles exhibit unique and tunable optical properties on account of their surface plasmon resonance (SPR). In this paper, we mainly discussed the theory background of the enhanced optical properties of noble metal nanoparticles. Mie theory, transfer matrix method, discrete dipole approximation (DDA) method, and finite-difference time domain (FDTD) method applied brute-force computational methods for different nanoparticles optical properties. Some important nanostructure fabrication technologies such as nanosphere lithography (NSL) and focused ion beam (FIB) are also introduced in this paper. Moreover, these fabricated nanostructures are used in the plasmonic sensing fields. The binding signal between the antibody and antigen, amyloid-derived diffusible ligands (ADDLs)-potential Alzheimer's disease (AD) biomarkers, and staphylococcal enterotoxin B (SEB) in nano-Moore per liter (nM) concentration level are detected by our designed nanobiosensor. They have many potential applications in the biosensor, environment protection, food security, and medicine safety for health, and so forth, fields.

MacA, a periplasmic membrane fusion protein of the macrolide transporter MacAB-TolC, binds lipopolysaccharide core specifically and with high affinity.

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Lu, Shuo; Zgurskaya, Helen I

2013-11-01

The Escherichia coli MacAB-TolC transporter has been implicated in efflux of macrolide antibiotics and secretion of enterotoxin STII. In this study, we found that purified MacA, a periplasmic membrane fusion protein, contains one tightly bound rough core lipopolysaccharide (R-LPS) molecule per MacA molecule. R-LPS was bound specifically to MacA protein with affinity exceeding that of polymyxin B. Sequence analyses showed that MacA contains two high-density clusters of positively charged amino acid residues located in the cytoplasmic N-terminal domain and the periplasmic C-terminal domain. Substitutions in the C-terminal cluster reducing the positive-charge density completely abolished binding of R-LPS. At the same time, these substitutions significantly reduced the functionality of MacA in the protection of E. coli against macrolides in vivo and in the in vitro MacB ATPase stimulation assays. Taken together, our results suggest that R-LPS or a similar glycolipid is a physiological substrate of MacAB-TolC.

Incidence of Clostridium perfringens in Broiler Chickens in the Czech Republic

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I. Svobodová

2007-01-01

Full Text Available Clostridium perfringens is a causative agent of human and animal foodborne diseases. It is known as a normal inhabitant of the intestinal tract of chickens as well as a potential pathogen causing necrotic enteritis. The aim of the present study was to detect the incidence of C. perfringens in healthy broiler chickens. From May 2005 to September 2006, 609 samples of caecal content from broilers from 23 intensive poultry farms were analyzed. The samples were cultivated on TSC and blood agar, typical colonies were identified and biochemically confirmed. the total number of positive samples was 112 (18.39%. the samples were processed by the multiplex polymerase chain reaction method (PCR for toxin genotyping. The presence of alpha, beta, beta2 and enterotoxin gene was detected. All C. perfringens isolates were classified as type A, four isolates had the cpb2 gene. In conclusion the prevalence of C. perfringens-positive farms is approximately 74% and the amount ranges about 104 cfu/g of caecal content.

Evaluation of Cholera Toxin Expression in Acidic, Alkaline and Neutral Conditions

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Narges Rahimi

2015-02-01

Full Text Available Background: Cholera is a severe disease which is caused by Vibrio cholerae and it is typically transmitted by either contaminated food or water particularly in developing countries. The most important virulence factor of this bacterium is an enterotoxin called cholera toxin which is a protein complex secreted by the Vibrio cholerae. Objectives: In this project, we determined the production of cholera toxin at different pH values. Materials and Methods: Two standard strain of Vibrio cholerae O1 biovar EL Tor N16961 and Vibrio cholerae O1 biovar Classic ATCC 14035 were used. After overnight cultivation of both the strains the total mRNA extracted and converted to total cDNA. Results: By Relative Real-Time PCR analysis the most cholera toxin production in classical and El Tor strains was at pH 8.5 and 8, respectively. Conclusions: Therefore, We may conclude that use of acidic diet will help in reduction of cholera toxin production.

A novel Hsp70 inhibitor prevents cell intoxication with the actin ADP-ribosylating Clostridium perfringens iota toxin

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Ernst, Katharina; Liebscher, Markus; Mathea, Sebastian; Granzhan, Anton; Schmid, Johannes; Popoff, Michel R.; Ihmels, Heiko; Barth, Holger; Schiene-Fischer, Cordelia

2016-01-01

Hsp70 family proteins are folding helper proteins involved in a wide variety of cellular pathways. Members of this family interact with key factors in signal transduction, transcription, cell-cycle control, and stress response. Here, we developed the first Hsp70 low molecular weight inhibitor specifically targeting the peptide binding site of human Hsp70. After demonstrating that the inhibitor modulates the Hsp70 function in the cell, we used the inhibitor to show for the first time that the stress-inducible chaperone Hsp70 functions as molecular component for entry of a bacterial protein toxin into mammalian cells. Pharmacological inhibition of Hsp70 protected cells from intoxication with the binary actin ADP-ribosylating iota toxin from Clostridium perfringens, the prototype of a family of enterotoxins from pathogenic Clostridia and inhibited translocation of its enzyme component across cell membranes into the cytosol. This finding offers a starting point for novel therapeutic strategies against certain bacterial toxins. PMID:26839186

Survey and properties of Staphylococcus aureus isolated from Japanese-style desserts.

Science.gov (United States)

Shimamura, Yuko; Kidokoro, Shiho; Murata, Masatsune

2006-07-01

We surveyed the contamination of 315 Japanese- and western-style desserts and 247 human hands by Staphylococcus aureus and other staphylococcal bacteria. The most frequently isolated staphylococcal bacterium was S. warneri, followed by S. aureus. Only 1.9% of western-style desserts were contaminated by S. aureus strains, while 19.4% and 13.0% of Japanese-style desserts and human hands respectively were contaminated. Ninety-four isolates of S. aureus were characterized as to their biological properties and enterotoxigenicity. Although staphylococcal enterotoxins (SEs) were detected by enzyme-linked immunosorbent assay in the cultured broth of all S. aureus isolates, the reversed passive latex agglutination method and the polymerase chain reaction showed only 39 (41.5%) and 40 (42.6%) samples respectively as SE-positive. The predominant type of SE was SEB (67.5%), and eight strains produced SEA. None of the S. aureus strains had penicillin-binding protein 2', showing that methicillin-resistant S. aureus was not present in the samples.

The application of alkaline and acidic electrolyzed water in the sterilization of chicken breasts and beef liver.

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Shimamura, Yuko; Shinke, Momoka; Hiraishi, Miki; Tsuchiya, Yusuke; Masuda, Shuichi

2016-05-01

The sterilization effect of a combination treatment with alkaline electrolyzed water (AlEW) and strong acidic electrolyzed water (StAEW) on fresh chicken breasts and beef liver was evaluated. Samples (1, 5, and 10 g) were inoculated with Salmonella Enteritidis NBRC3313, Escherichia coli ATCC 10798, Staphylococcus aureus FDA209P, and S. aureus C-29 [staphylococcal enterotoxin A (SEA) productive strain] and subjected to a dipping combination treatment (4°C and 25°C for 3 min) with AlEW and StAEW. Combination treatment with AlEW and StAEW significantly reduced the bacteria, and reduction of more than 1 log colony-forming units (CFU)/g was achieved. Furthermore, this combination treatment significantly decreased the SEA gene expression level in samples. Some quality variables of the meat samples such as pH, lipid oxidation, color, amino-acid content, texture, and sensory characteristics showed no significant differences between the combination treatment with AlEW and StAEW and the untreated control.

A large foodborne outbreak on a small Pacific island.

Science.gov (United States)

Thein, C C; Trinidad, R M; Pavlin, B I

2010-04-01

On March 25, 2009, the Ebeye Leroj Kitlang Memorial Health Center on the island of Ebeye in the Republic of the Marshall Islands was overwhelmed with over 100 patients presenting for vomiting and diarrhea. Epidemiologic investigation revealed that there were 174 cases among 187 attendees at a local funeral earlier in the day. Most cases had eaten served sandwiches containing egg products that had undergone severe time-temperature abuse. While no causal agents were identified, the epidemiology and clinical presentation is compatible with foodborne toxins, most likely enterotoxins of either Staphylococcus aureus or Bacillus cereus. Mitigation measures undertaken by public health centered on education of food preparers and the general public regarding safe food preparation practices. This large outbreak serves to remind us that, while there are simple and highly effective measures to prevent such foodborne disease outbreaks, we in the public health sector have a duty to improve the community's knowledge and understanding of these measures.

Aeromonas caviae strain induces Th1 cytokine response in mouse intestinal tract

Energy Technology Data Exchange (ETDEWEB)

Hayes, S L; Lye, D J; McKinstry, Craig A.; Vesper, Sephen J.

2010-01-01

Aeromonas caviae has been associated with human gastrointestinal disease. Strains of this species typically lack virulence factors (VFs) such as enterotoxins and hemolysins that are produced by other human pathogens of the Aeromonas genus. Microarray profiling of murine small intestinal extracts, 24 hours after oral infection with an A. caviae strain, provides evidence of a Th1 type immune response. A large number of gamma-interferon (γ-IFN) induced genes are up-regulated as well as several tumor necrosis factor-alpha (TNF-α) transcripts. A. caviae has always been considered as opportunistic pathogen because it lacks obvious virulence factors. This current effort suggests that an A. caviae strain can colonize the murine intestinal tract and cause what has been described by others as a dysregulatory cytokine response. This response could explain why a number of diarrheal waterborne disease cases have been attributed to A. caviae even though it lacks obvious enteropathogenic properties.

Prevalence and characterization of Vibrio cholerae isolated from shrimp products imported into Denmark

DEFF Research Database (Denmark)

Dalsgaard, A.; Bjergskov, T.; Jeppesen, V.F.

1996-01-01

A total of 3,555 metric tonnes of warm water shrimp were imported into Denmark from December 1994 to July 1995. V. cholerae O1 was not detected in any of the 748 samples analyzed. Non-Ol V. cholerae was found in a single (0.1%) cooked frozen shrimp product and in five (0.7%) raw frozen products......, all originating from shrimp produced in aquaculture. Six isolated strains agglutinated in polyvalent O antisera, but did not agglutinate in Ogawa or Inaba antisera. The six strains were resistant to colistin and sulfisoxazole; three strains also showed resistance to ampicillin. None of the strains...... contained plasmids or genes encoding cholera toxin (CT) or heat-stable enterotoxin (NAG-ST), The absence of V. cholerae O1 and the low number of samples containing CT and NAG-ST negative non-Ol strains in imported shrimp suggest that I! cholerae in such products may not constitute a public health problem....

Molecular typing and antimicrobial susceptibility of Clostridium perfringens from broiler chickens.

Science.gov (United States)

Gharaibeh, Saad; Al Rifai, Rami; Al-Majali, Ahmad

2010-12-01

Clostridium perfringens (Cp) causes necrotic enteritis disease in commercial poultry. Antimicrobials are used to control and treat this disease and sometimes clinical outbreaks do not respond well to certain treatments. This study was designed to isolate Cp from clinical cases, type these isolates by multiplex PCR, and determine their antimicrobial susceptibility by micro-dilution method. A total of 67 Cp isolates were obtained from 155 broiler chicken flocks. All isolates were classified as type A and non-enterotoxin producers. Lincomycin, erythromycins, and tilmicosin showed very high minimal inhibitory concentration (MIC) 50 of ≥256 μg/ml. However, tylosin, amoxicillin, ampicillin, penicillin, florfenicol, danofloxacin, enrofloxacin, chlortetracycline, doxycycline, and oxytetracycline had variable MIC₅₀ of 64, 0.5, 1, 1, 8, 4, 8, 4, 8, 0.5 μg/ml, respectively. It is recommended that Cp infections in Jordan be treated with either penicillins or tetracyclines especially amoxicillin and oxytetracycline. Copyright © 2010 Elsevier Ltd. All rights reserved.

Toxins as biological weapons for terror-characteristics, challenges and medical countermeasures: a mini-review.

Science.gov (United States)

Berger, Tamar; Eisenkraft, Arik; Bar-Haim, Erez; Kassirer, Michael; Aran, Adi Avniel; Fogel, Itay

2016-01-01

Toxins are hazardous biochemical compounds derived from bacteria, fungi, or plants. Some have mechanisms of action and physical properties that make them amenable for use as potential warfare agents. Currently, some toxins are classified as potential biological weapons, although they have several differences from classic living bio-terror pathogens and some similarities to manmade chemical warfare agents. This review focuses on category A and B bio-terror toxins recognized by the Centers for Disease Control and Prevention: Botulinum neurotoxin, staphylococcal enterotoxin B, Clostridium perfringens epsilon toxin, and ricin. Their derivation, pathogenesis, mechanism of action, associated clinical signs and symptoms, diagnosis, and treatment are discussed in detail. Given their expected covert use, the primary diagnostic challenge in toxin exposure is the early detection of morbidity clusters, apart from background morbidity, after a relatively short incubation period. For this reason, it is important that clinicians be familiar with the clinical manifestations of toxins and the appropriate methods of management and countermeasures.

Detection of Ampicillin Resistance Genes (bla in Clinical Isolates of Escherichia coli with Polymerase Chain Reaction Method

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Tiana Milanda

2014-09-01

Full Text Available Escherichia coli is a rod negative Gram which could be pathogenic, if its value increases or located in outer gastrointestinal tract. Pathogenic E. coli will produce enterotoxin which will cause diarrhoea or infection in urine tract. Ampicilin was one of particular antibiotics to overcome infection. Ampicilin nowadays is no longer used as primary medicine, because of its resistance case. The aim of this research is to detect the presence of gene which is responsible to ampicilin resistant E. coli. We used isolated midstream urine from cystitis object in Hasan Sadikin Hospital (RSHS as samples. Polymerase Chain Reaction (PCR method (colony-PCR and DNA-PCR were done to invenstigate the antibiotic resistency. Based on the result of antibiotic susceptibility testing to ampicillin, E. coli samples were resistant to ampicilin. Elektroforegram products of colony-PCR and DNA-PCR showed that the resistance case of ampicilin caused by bla gene (199 bp. Selective and rational antibiotic treatment is required to prevent ampicillin resistance in patients with symptoms

Genomic Insights and Its Comparative Analysis with Yersinia enterocolitica Reveals the Potential Virulence Determinants and Further Pathogenicity for Foodborne Outbreaks.

Science.gov (United States)

Gnanasekaran, Gopalsamy; Na, Eun Jung; Chung, Han Young; Kim, Suyeon; Kim, You-Tae; Kwak, Woori; Kim, Heebal; Ryu, Sangryeol; Choi, Sang Ho; Lee, Ju-Hoon

2017-02-28

Yersinia enterocolitica is a well-known foodborne pathogen causing gastrointestinal infections worldwide. The strain Y. enterocolitica FORC_002 was isolated from the gill of flatfish (plaice) and its genome was sequenced. The genomic DNA consists of 4,837,317 bp with a GC content of 47.1%, and is predicted to contain 4,221 open reading frames, 81 tRNA genes, and 26 rRNA genes. Interestingly, genomic analysis revealed pathogenesis and host immune evasion-associated genes encoding guanylate cyclase (Yst), invasin (Ail and Inv), outer membrane protein (Yops), autotransporter adhesin A (YadA), RTX-like toxins, and a type III secretion system. In particular, guanylate cyclase is a heat-stable enterotoxin causing Yersinia -associated diarrhea, and RTX-like toxins are responsible for attachment to integrin on the target cell for cytotoxic action. This genome can be used to identify virulence factors that can be applied for the development of novel biomarkers for the rapid detection of this pathogen in foods.

New tetrameric forms of the rotavirus NSP4 with antiparallel helices.

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Kumar, Sushant; Ramappa, Raghavendra; Pamidimukkala, Kiranmayee; Rao, C D; Suguna, K

2018-06-01

Rotavirus nonstructural protein 4, the first viral enterotoxin to be identified, is a multidomain, multifunctional glycoprotein. Earlier, we reported a Ca 2+ -bound coiled-coil tetrameric structure of the diarrhea-inducing region of NSP4 from the rotavirus strains SA11 and I321 and a Ca 2+ -free pentameric structure from the rotavirus strain ST3, all with a parallel arrangement of α-helices. pH was found to determine the oligomeric state: a basic pH favoured a tetramer, whereas an acidic pH favoured a pentamer. Here, we report two novel forms of the coiled-coil region of NSP4 from the bovine rotavirus strains MF66 and NCDV. These crystallized at acidic pH, forming antiparallel coiled-coil tetrameric structures without any bound Ca 2+ ion. Structural and mutational studies of the coiled-coil regions of NSP4 revealed that the nature of the residue at position 131 (Tyr/His) plays an important role in the observed structural diversity.

Prevalence and Characterization of Staphylococcus aureus Cultured From Raw Milk Taken From Dairy Cows With Mastitis in Beijing, China

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Wei Wang

2018-06-01

Full Text Available The colonization of dairy herds and subsequent contamination of raw milk by Staphylococcus aureus (S. aureus, especially those expressing a multi-drug resistance (MDR, biofilm and toxins producing ability, remains an important issue for both the dairy producer and public health. In this study, we investigated the prevalence, antimicrobial resistance, virulence, and genetic diversity of S. aureus in raw milk taken from 2 dairy farms in Beijing, China. Ninety (46.2%, 90/195 samples were positive for S. aureus. Resistant to penicillin (PEN (31.3%, ciprofloxacin (18.8% and enrofloxacin (15.6% were the most often observed. Isolates cultured from farm B showed significantly higher resistance to penicillin (73.9%, ciprofloxacin (34.8%, enrofloxacin (34.8%, tilmicosin (17.4%, and erythromycin (17.4% than those from farm A (p < 0.05. Totally, 94.8% S. aureus harbored at least one virulence gene and the pvl (93.8%, sec (65.6%, and sea (60.4% genes were the most frequently detected. The pvl and sec genes were more often detected in isolates from farm A (97.3% and 84.9% respectively than those from farm B (p < 0.05. Of all 77 staphylococcus enterotoxin (SE-positive isolates, more than 90% could produce enterotoxins and 70.1% could produce two types. Biofilm related genes (icaA/D, clf/B, can, and fnbA were detected in all96 isolates. All 96 isolates could produce biofilm with 8.3, 70.8, and 18.8% of the isolates demonstrating weak, moderate and strong biofilm formation, respectively. A total of 5 STs, 7 spa types (1 novel spa type t17182, 3agr types (no agrII, and 14 SmaI-pulso-types were found in this study. PFGE cluster II-CC1-ST1-t127-agr III was the most prevalent clone (56.3%. Isolates of agr III (PFGE Cluster I/II-CC1-ST1-t127/2279 had higher detection of virulence genes than those of agr I and agr IV. TheMSSA-ST398-t1456-agr I clone expressed the greatest MDRbut with no virulence genes and weakly biofilm formation. Our finding indicated a relatively

Influence of temperature shifts on survival, growth, and toxin production by psychrotrophic and mesophilic strains of Bacillus cereus in potatoes and chicken gravy.

Science.gov (United States)

Mahakarnchanakul, W; Beuchat, L R

1999-03-15

A study was done to determine the influence of temperature on growth and toxin production characteristics of psychrotrophic and mesophilic strains of Bacillus cereus when inoculated into mashed potatoes and chicken gravy containing various concentrations of sodium chloride and held at temperatures different from those at which cells had been cultured. Logarithmic growth phase cells (10 h, 30 degrees C) of psychrotrophic (F3802A/84) and mesophilic (B4ac-1) strains of Bacillus cereus were inoculated into rehydrated commercially processed instant mashed potatoes and chicken gravy supplemented with 0, 2, or 4% sodium chloride. Growth, survival, and diarrheal toxin production in potatoes and gravy held at 30, 37, and 10 degrees C (strain F3802A/84) or 30, 40, and 10 degrees C (strain B4ac-1) were monitored. Both strains grew in both foods containing no added sodium chloride or 2% sodium chloride when held at 30, 37, or 40 degrees C for 2 days. Strain B4ac-1 grew better than strain F3802A/84 in foods containing 4% sodium chloride. Maximum amounts of enterotoxin (1024 ng/g) were produced by strain B4ac-1 in chicken gravy held at 30 and 40 degrees C. Strain F3802A/84 grew to populations of 7 log10 CFU/g in foods containing no added sodium chloride or 2% sodium chloride at 10 degrees C. Strain F3802A/84 produced the highest amount of enterotoxin (1024 ng/g) at 30 degrees C in chicken gravy containing 0.7 or 2% sodium chloride; however, little or low amounts of toxin (4-16 ng/g) were produced in chicken gravy at 10 degrees C. Compared to strain B4ac-1, cells of strain F3802A/84 subjected to a downward shift in incubation temperature (10 degrees C) grew more rapidly in chicken gravy. Strain B4ac-1 produced the highest amount of toxin (1024 ng/g) at 30 degrees C in gravy containing 4% sodium chloride and at 40 degrees C in gravy containing 0.7% sodium chloride. Toxin was not detected in inoculated mashed potatoes. Results of this study indicate that shifts in incubation

Effect of gamma radiation on the resistance of Staphylococcus aureus (Rosembach, 1884) and in the physical-chemical and sensory properties of 'minas frescal' cheese

International Nuclear Information System (INIS)

Gurgel, Maria Sylvia de Campos Carvalho do Amaral

2000-01-01

Among food poisoning of microbial origin, S. aureus stands as one of the most important, being a pathogenic bacteria for as human kind and of important occurrence in food products. Although 'Minas Frescal' cheese is typically Brazilian, there aren't in this country technical rules for its preparation and appropriate inspection. In the present work the effect of gamma radiation on the resistance of S. aureus and its ability to produce toxins in 'Minas Frescal' cheese were evaluated. The effects of the radiation in 'Minas Frescal' cheese physical-chemical and sensory properties will be determined, with the objective of studying the efficiency of the gamma radiation as method of conservation of this product. During the fabrication of 'Minas Frescal' cheese in the Laboratory of Irradiation of Foods (CENA/USP), S. aureus strains (ATCC 13565, ATCC 14458, ATCC 19095) with approximate count of 106 UFC/ml were inoculated. After irradiation with doses of O(control); 1; 2; 3 and 4 kGy, the cheeses were stored under refrigeration (+- 5 deg C) and analyzed at 1, 7 and 14 days. After irradiation with doses of O(control); 1; 2; 3 and 4 kGy, the cheeses were stored under refrigeration (+- 5 deg C) and analyzed at 1,7 and 14 days. 'Minas Frescal' cheese was evaluated through the determination of parameters of acidity, pH, moisture and lever of extension proteolysis, according to methodology of Association of Official Analytical Chemists (AO.AC.),1995, and sensory analysis by the ADQ (Quantitative Descriptive Analysis) method. The microbiological analysis determined the survival of S. aureus in the Baird-Parker a selective medium and its ability of producing enterotoxin by the Passive Reverse Agglutination with Latex method. the results analyzed through randomized blocks in a factorial design (5x3), revealed that 'Minas Frescal' cheese did not present significant differences among the radiation doses in relation to its physical-chemical and sensory properties. Dose of 3 kGy was

Diversity and Impact of Prokaryotic Toxins on Aquatic Environments: A Review

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Rogério Tenreiro

2010-10-01

Full Text Available Microorganisms are ubiquitous in all habitats and are recognized by their metabolic versatility and ability to produce many bioactive compounds, including toxins. Some of the most common toxins present in water are produced by several cyanobacterial species. As a result, their blooms create major threats to animal and human health, tourism, recreation and aquaculture. Quite a few cyanobacterial toxins have been described, including hepatotoxins, neurotoxins, cytotoxins and dermatotoxins. These toxins are secondary metabolites, presenting a vast diversity of structures and variants. Most of cyanobacterial secondary metabolites are peptides or have peptidic substructures and are assumed to be synthesized by non-ribosomal peptide synthesis (NRPS, involving peptide synthetases, or NRPS/PKS, involving peptide synthetases and polyketide synthases hybrid pathways. Besides cyanobacteria, other bacteria associated with aquatic environments are recognized as significant toxin producers, representing important issues in food safety, public health, and human and animal well being. Vibrio species are one of the most representative groups of aquatic toxin producers, commonly associated with seafood-born infections. Some enterotoxins and hemolysins have been identified as fundamental for V. cholerae and V. vulnificus pathogenesis, but there is evidence for the existence of other potential toxins. Campylobacter spp. and Escherichia coli are also water contaminants and are able to produce important toxins after infecting their hosts. Other bacteria associated with aquatic environments are emerging as toxin producers, namely Legionella pneumophila and Aeromonas hydrophila, described as responsible for the synthesis of several exotoxins, enterotoxins and cytotoxins. Furthermore, several Clostridium species can produce potent neurotoxins. Although not considered aquatic microorganisms, they are ubiquitous in the environment and can easily contaminate drinking

Enterotoxigenicidade de Staphylococcus spp. isolados de leite in natura Enterotoxigenicity of Staphylococcus spp. isolated of milk in natura

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Tânia Lúcia Montenegro Stamford

2006-03-01

agents, which can take to manifested diseases by action of pathogenic microorganisms or their toxins. It was researched the occurrence of strains of Staphylococcus and your capacity in producing enterotoxins in milk in natura, that have been produced or commercialized in the State Pernambuco, Brazil. 109 strains of Staphylococus positive and negative coagulase of milk in natura were selected. The identification of the isolated strains was accomplished through morphologic and biochemical tests as: catalase, coagulase, haemolysins, DNAse, thermonuclease, acetoin production (VP and carbohydrates metabolism (glucose, maltose and mannitol. From the 77 coagulase positive strains 30 were identified as S. aureus, 3 as S. hyicus, 16 as S. intermedius, 13 as S. aureus identification presumptive and 15 as SCP. Among 32 coagulase negative strains 2 were identified as S. capitis, 1 as S. carnosus, 6 as S. chromogenes, 1 as S. hyicus, 1 as S. schleiferi and 21 as SCN. Fourty-three strains that presented very evident thermonuclease reaction, were selected in order to perform for staphylococcal enterotoxins analysis by the immuno enzimatic test (ELFA. 10 strains showed negative reaction for enterotoxins: S. aureus (4, S. carnosus (1, S. chromogenes (2, S. hyicus (2 and S. intermedius (1. Strains that gave positive results, were S. aureus (17, S. chromogenes (2, S. hyicus (1, S. intermedius (8, S. aureus identified presumptively (2 and of the groups SCP (1 and SCN (2. The species that presented larger number of enterotoxigenics strains were S. aureus and S. intermedius. Results can be attributed to the inadequate manipulation or food recontamination during the storage and distribution.

Staphylococcal Enterotoxins, Toxic Shock Syndrome Toxin-I, and Streptococcal Pyrogenic Exotoxins: Some Basic Biology of Bacterial Superantigens

National Research Council Canada - National Science Library

Krakauer, Teresa

2003-01-01

Staphylococcus aureus and Streptococcus pyogenes are facultative gram-positive cocci that play an important role in a myriad of human illnesses, including food poisoning, skin infections, pharyngitis...

Influence of water temperature and salinity on seasonal occurrences of Vibrio cholerae and enteric bacteria in oyster-producing areas of Veracruz, México.

Science.gov (United States)

Castañeda Chávez, Maria del Refugio; Pardio Sedas, Violeta; Orrantia Borunda, Erasmo; Lango Reynoso, Fabiola

2005-12-01

The influence of temperature and salinity on the occurrence of Vibrio cholerae, Escherichia coli and Salmonella spp. associated with water and oyster samples was investigated in two lagoons on the Atlantic Coast of Veracruz, Mexico over a 1-year period. The results indicated that seasonal salinity variability and warm temperatures, as well as nutrient influx, may influence the occurrence of V. cholera. non-O1 and O1. The conditions found in the Alvarado (31.12 degrees C, 6.27 per thousand, pH=8.74) and La Mancha lagoons (31.38 degrees C, 24.18 per thousand, pH=9.15) during the rainy season 2002 favored the occurrence of V. cholera O1 Inaba enterotoxin positive traced in oysters. Vibrio alginolyticus was detected in Alvarado lagoon water samples during the winter season. E. coli and Salmonella spp. were isolated from water samples from the La Mancha (90-96.7% and 86.7-96.7%) and Alvarado (88.6-97.1% and 88.6-100%) lagoons. Occurrence of bacteria may be due to effluents from urban, agricultural and industrial areas.

Algevir: An Expression System for Microalgae Based on Viral Vectors

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Bernardo Bañuelos-Hernández

2017-06-01

Full Text Available The use of recombinant algae for the production of valuable compounds is opening promising biotechnological applications. However, the development of efficient expression approaches is still needed to expand the exploitation of microalgae in biotechnology. Herein, the concept of using viral expression vectors in microalgae was explored for the first time. An inducible geminiviral vector leading to Rep-mediated replication of the expression cassette allowed the production of antigenic proteins at high levels. This system, called Algevir, allows the production of complex viral proteins (GP1 from Zaire ebolavirus and bacterial toxin subunits (B subunit of the heat-labile Escherichia coli enterotoxin, which retained their antigenic activity. The highest achieved yield was 1.25 mg/g fresh biomass (6 mg/L of culture, which was attained 3 days after transformation. The Algevir system allows for a fast and efficient production of recombinant proteins, overcoming the difficulties imposed by the low yields and unstable expression patterns frequently observed in stably transformed microalgae at the nuclear level; as well as the toxicity of some target proteins.

Cholera toxin B subunits assemble into pentamers--proposition of a fly-casting mechanism.

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Jihad Zrimi

Full Text Available The cholera toxin B pentamer (CtxB(5, which belongs to the AB(5 toxin family, is used as a model study for protein assembly. The effect of the pH on the reassembly of the toxin was investigated using immunochemical, electrophoretic and spectroscopic methods. Three pH-dependent steps were identified during the toxin reassembly: (i acquisition of a fully assembly-competent fold by the CtxB monomer, (ii association of CtxB monomer into oligomers, (iii acquisition of the native fold by the CtxB pentamer. The results show that CtxB(5 and the related heat labile enterotoxin LTB(5 have distinct mechanisms of assembly despite sharing high sequence identity (84% and almost identical atomic structures. The difference can be pinpointed to four histidines which are spread along the protein sequence and may act together. Thus, most of the toxin B amino acids appear negligible for the assembly, raising the possibility that assembly is driven by a small network of amino acids instead of involving all of them.

Prevalence of Staphylococcus aureus in Shrimps in Tehran during 2013

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Mohammad Mehdi Soltan Dallal

2015-12-01

Full Text Available Background During fishing and transport, preservation and quality of fish products are importantas well as storage to prevent the growth of pathogenic and toxin producing bacteria.Staphylococcus aureus is one of the most common causes of sea food-borne diseases worldwidedue to contamination of food by preformed enterotoxins. The aim of this study was to compare theprevalence and contamination of S. aureus in marine and farmed shrimps in Tehran fishery center.Methods: A total of 300 samples, including 150 marine, 150 farmed shrimps were selected duringSeptember 2013 to December 2013. Isolation and identification of S. aureus from isolated sampleswere carried out according to conventional methods, and antibiotic susceptibility test wasperformed by modified Kirby-Bauer disc diffusion methodResults: The results of this study showed that 30% of marine and 20% off armed shrimps werecontaminated with S. aureus. The highest resistance was observed with penicillin and ampicillin,whereas 100% were sensitive to vancomycin, clindamycin, ciprofloxacin, and rifampin.Conclusions: Due to relatively high contamination of shrimp by S. aureus more attention shouldbe given during processing and manufacturing.

Prevalence of Staphylococcus aureus in Shrimps in Tehran during 2013

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Mohammad Mehdi Soltan Dallal

2016-02-01

Full Text Available Background During fishing and transport, preservation and quality of fish products are importantas well as storage to prevent the growth of pathogenic and toxin producing bacteria.Staphylococcus aureus is one of the most common causes of sea food-borne diseases worldwidedue to contamination of food by preformed enterotoxins. The aim of this study was to compare theprevalence and contamination of S. aureus in marine and farmed shrimps in Tehran fishery center.Methods: A total of 300 samples, including 150 marine, 150 farmed shrimps were selected duringSeptember 2013 to December 2014. Isolation and identification of S. aureus from isolated sampleswere carried out according to conventional methods, and antibiotic susceptibility test wasperformed by modified Kirby-Bauer disc diffusion method.Results: The results of this study showed that 30% of marine and 20% off armed shrimps werecontaminated with S. aureus. The highest resistance was observed with penicillin and ampicillin,whereas 100% were sensitive to vancomycin, clindamycin, ciprofloxacin, and rifampin.Conclusions: Due to relatively high contamination of shrimp by S. aureus more attention shouldbe given during processing and manufacturing.

Multidrug resistance in Clostridium perfringens isolated from diarrheal neonatal piglets in Thailand.

Science.gov (United States)

Ngamwongsatit, Bhinyada; Tanomsridachchai, Wimonrat; Suthienkul, Orasa; Urairong, Supanee; Navasakuljinda, Wichian; Janvilisri, Tavan

2016-04-01

Clostridium perfringens causes diarrhea in neonatal piglets, thereby affecting commercial swine farming. The objective of this study was to determine the prevalence and characterize antimicrobial resistance in C. perfringens isolated from diarrheal neonatal piglets in Thailand. A total of 260 rectal swab samples were collected from 13 farms and were subjected to C. perfringens isolation. A total of 148 samples were PCR-positive for C. perfringens toxin genes, from which 122 were recovered. All isolates were cpb2-encoding C. perfringens type A and enterotoxin gene negative. Most of the isolates were susceptible to ampicillin, bacitracin, chlorotetracycline, doxycycline, and oxytetracycline with MIC50 values ranging from 0.32 to 8 μg/ml. The high resistance rates were observed for ceftiofur, enrofloxacin, erythromycin, lincomycin, and tylosin. Among resistant isolates, 82% were resistant to more than one type of antibiotics. The distinct pattern of multiple drug resistance in C. perfringens was observed in different regions, potentially reflecting the farm specific usage of these agents. Copyright © 2016 Elsevier Ltd. All rights reserved.

Short communication: Characterization of methicillin-resistant Staphylococcus aureus isolated from raw milk fresh cheese in Colombia.

Science.gov (United States)

Herrera, Fanny C; García-López, María-Luisa; Santos, Jesús A

2016-10-01

The aim of this study was the characterization of a collection of 8 methicillin-resistant Staphylococcus aureus (MRSA) isolates, obtained from samples of fresh cheese (Doble Crema) produced from raw cow milk in small dairies in Colombia. All the isolates harbored the mecA and Panton-Valentine leukocidin (PVL) genes, presented with SCCmec type IV, and belonged to multilocus sequence type 8 and spa type 024. Seven isolates presented 3 closely related pulsed-field gel electrophoresis profiles. Three of them carried the staphylococcal enterotoxin B gene. The isolates were resistant to cefoxitin, oxacillin, penicillin, and ampicillin and susceptible to all non-β-lactams antibiotics tested, with minimum inhibitory concentration values for oxacillin of 4 to 8mg/L. The isolates belonged to the community-acquired MRSA group, suggesting a human source of contamination. The risk of human infection by MRSA via contaminated foods is considered low, but contaminated food commodities can contribute to the worldwide dissemination of clones of community-acquired MRSA. Copyright © 2016 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Removal of Cholera Toxin from Aqueous Solution by Probiotic Bacteria

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Jussi A. O. Meriluoto

2012-06-01

Full Text Available Cholera remains a serious health problem, especially in developing countries where basic hygiene standards are not met. The symptoms of cholera are caused by cholera toxin, an enterotoxin, which is produced by the bacterium Vibrio cholerae. We have recently shown that human probiotic bacteria are capable of removing cyanobacterial toxins from aqueous solutions. In the present study we investigate the ability of the human probiotic bacteria, Lactobacillus rhamnosus strain GG (ATCC 53103 and Bifidobacterium longum 46 (DSM 14583, to remove cholera toxin from solution in vitro. Lactobacillus rhamnosus strain GG and Bifidobacterium longum 46 were able to remove 68% and 59% of cholera toxin from aqueous solutions during 18 h of incubation at 37 °C, respectively. The effect was dependent on bacterial concentration and L. rhamnosus GG was more effective at lower bacterial concentrations. No significant effect on cholera toxin concentration was observed when nonviable bacteria or bacterial supernatant was used.

Algevir: An Expression System for Microalgae Based on Viral Vectors

Science.gov (United States)

Bañuelos-Hernández, Bernardo; Monreal-Escalante, Elizabeth; González-Ortega, Omar; Angulo, Carlos; Rosales-Mendoza, Sergio

2017-01-01

The use of recombinant algae for the production of valuable compounds is opening promising biotechnological applications. However, the development of efficient expression approaches is still needed to expand the exploitation of microalgae in biotechnology. Herein, the concept of using viral expression vectors in microalgae was explored for the first time. An inducible geminiviral vector leading to Rep-mediated replication of the expression cassette allowed the production of antigenic proteins at high levels. This system, called Algevir, allows the production of complex viral proteins (GP1 from Zaire ebolavirus) and bacterial toxin subunits (B subunit of the heat-labile Escherichia coli enterotoxin), which retained their antigenic activity. The highest achieved yield was 1.25 mg/g fresh biomass (6 mg/L of culture), which was attained 3 days after transformation. The Algevir system allows for a fast and efficient production of recombinant proteins, overcoming the difficulties imposed by the low yields and unstable expression patterns frequently observed in stably transformed microalgae at the nuclear level; as well as the toxicity of some target proteins. PMID:28713333

Staphylococcus aureus food-poisoning outbreak associated with the consumption of ice-cream.

Science.gov (United States)

Fetsch, A; Contzen, M; Hartelt, K; Kleiser, A; Maassen, S; Rau, J; Kraushaar, B; Layer, F; Strommenger, B

2014-09-18

In April 2013, a food poisoning outbreak caused by staphylococcal enterotoxins (SEs) in ice-cream occurred in Freiburg, Germany, among the 31 participants of a christening party. Of the 13 cases, seven were hospitalized or obtained ambulatory treatment. Different types of ice-cream, which was freshly produced at the hotel where the party took place, were found to contain SE and high amounts of coagulase positive staphylococci. Enterotoxigenic Staphylococcus aureus strains isolated from ice-cream and human cases were of the same spa-type (t127), harboured the sea gene and displayed identical phenotypic resistance-, Fourier transform infrared spectroscopy- (FT-IR) and microarray-profiles. Despite the strong microbiological and epidemiological evidence of ice-cream being the incriminated food vehicle of the outbreak, a common source of S. aureus from the ice-cream could not be deduced. As none of the employees carried the outbreak strain, either the equipment used for the production of the ice-cream or a contaminated ingredient is the most likely introduction source. Copyright © 2014 Elsevier B.V. All rights reserved.

[A study on the construction, expression and immunosterility of Lagurus laguru zona pellucida 3 DNA vaccine pVAX1-sig-LTB-lZP3-C3d3].

Science.gov (United States)

Li, Chen-Chen; Yu, Ji-Yun; Jiang, Min; Tu, Yi-Xian; Ma, Xiao-Lin; Zhang, Fu-Chun

2011-09-01

To enhance the immunocontraceptive effect of Lagurus lagurus zona pellucida 3 DNA vaccine, and to achieve the prospect of application through the pVAX1-sig-LTB-lZP3-C3d3 different immunity pathway. Two adjuvant molecules were constructed into the recombinant plasmid pVAX1-sig-LTB-lZP3-C3d3 as DNA vaccine which contains Escherichia coli heat-labile enterotoxin B subunit and the molecular adjuvant 3 copies of C3d. The results of RT-PCR and western blot showed that the DNA vaccine was expressed in mRNA and protein level. The female C57BL/6 mice were immunized by three ways: intramuscular injection, intranasal or oral route.Antibody levels and types were detected by ELISA. ELISA results showed that recombinant plasmid pVAX1-sig-LTB-lZP3-C3d3 immunization induced specific IgG, IgA levels were significantly different comparing with control (Psig-LTB-lZP3-C3d3 can induce the specific immune response efficiently and enhance the immunocontraceptive effects.

Array biosensor for detection of toxins

Science.gov (United States)

Ligler, Frances S.; Taitt, Chris Rowe; Shriver-Lake, Lisa C.; Sapsford, Kim E.; Shubin, Yura; Golden, Joel P.

2003-01-01

The array biosensor is capable of detecting multiple targets rapidly and simultaneously on the surface of a single waveguide. Sandwich and competitive fluoroimmunoassays have been developed to detect high and low molecular weight toxins, respectively, in complex samples. Recognition molecules (usually antibodies) were first immobilized in specific locations on the waveguide and the resultant patterned array was used to interrogate up to 12 different samples for the presence of multiple different analytes. Upon binding of a fluorescent analyte or fluorescent immunocomplex, the pattern of fluorescent spots was detected using a CCD camera. Automated image analysis was used to determine a mean fluorescence value for each assay spot and to subtract the local background signal. The location of the spot and its mean fluorescence value were used to determine the toxin identity and concentration. Toxins were measured in clinical fluids, environmental samples and foods, with minimal sample preparation. Results are shown for rapid analyses of staphylococcal enterotoxin B, ricin, cholera toxin, botulinum toxoids, trinitrotoluene, and the mycotoxin fumonisin. Toxins were detected at levels as low as 0.5 ng mL(-1).

Immunogenicity of nuclear-encoded LTB:ST fusion protein from Escherichia coli expressed in tobacco plants.

Science.gov (United States)

Rosales-Mendoza, Sergio; Soria-Guerra, Ruth E; Moreno-Fierros, Leticia; Govea-Alonso, Dania O; Herrera-Díaz, Areli; Korban, Schuyler S; Alpuche-Solís, Ángel G

2011-06-01

Enterotoxigenic Escherichia coli (ETEC) is one of the main causative agents of diarrhea in infants and for travelers. Inclusion of a heat-stable (ST) toxin into vaccine formulations is mandatory as most ETEC strains can produce both heat-labile (LT) and ST enterotoxins. In this study, a genetic fusion gene encoding for an LTB:ST protein has been constructed and transferred into tobacco via Agrobacterium tumefaciens-mediated transformation. Transgenic tobacco plants carrying the LTB:ST gene are then subjected to GM1-ELISA revealing that the LTB:ST has assembled into pentamers and displays antigenic determinants from both LTB and ST. Protein accumulation of up to 0.05% total soluble protein is detected. Subsequently, mucosal and systemic humoral responses are elicited in mice orally dosed with transgenic tobacco leaves. This has suggested that the plant-derived LTB:ST is immunogenic via the oral route. These findings are critical for the development of a plant-based vaccine capable of eliciting broader protection against ETEC and targeting both LTB and ST. Features of this platform in comparison to transplastomic approaches are discussed.

Hsp70 facilitates trans-membrane transport of bacterial ADP-ribosylating toxins into the cytosol of mammalian cells.

Science.gov (United States)

Ernst, Katharina; Schmid, Johannes; Beck, Matthias; Hägele, Marlen; Hohwieler, Meike; Hauff, Patricia; Ückert, Anna Katharina; Anastasia, Anna; Fauler, Michael; Jank, Thomas; Aktories, Klaus; Popoff, Michel R; Schiene-Fischer, Cordelia; Kleger, Alexander; Müller, Martin; Frick, Manfred; Barth, Holger

2017-06-02

Binary enterotoxins Clostridium (C.) botulinum C2 toxin, C. perfringens iota toxin and C. difficile toxin CDT are composed of a transport (B) and a separate non-linked enzyme (A) component. Their B-components mediate endocytic uptake into mammalian cells and subsequently transport of the A-components from acidic endosomes into the cytosol, where the latter ADP-ribosylate G-actin resulting in cell rounding and cell death causing clinical symptoms. Protein folding enzymes, including Hsp90 and peptidyl-prolyl cis/trans isomerases facilitate transport of the A-components across endosomal membranes. Here, we identified Hsp70 as a novel host cell factor specifically interacting with A-components of C2, iota and CDT toxins to facilitate their transport into the cell cytosol. Pharmacological Hsp70-inhibition specifically prevented pH-dependent trans-membrane transport of A-components into the cytosol thereby protecting living cells and stem cell-derived human miniguts from intoxication. Thus, Hsp70-inhibition might lead to development of novel therapeutic strategies to treat diseases associated with bacterial ADP-ribosylating toxins.

Analysis of plasmid profiling as a method for rapid differentiation of food-associated Clostridium perfringens strains.

Science.gov (United States)

Jones, M K; Iwanejko, L A; Longden, M S

1989-09-01

Plasmid analysis of over 120 strains of Clostridium perfringens, isolated during food-poisoning incidents and from animal carcasses and food constituents with no association with food poisoning, showed the potential of plasmid profiling as a means of differentiating epidemiologically related strains. On average 65% of freshly isolated strains contained one or more plasmids which could be used in the analysis. Comparison of profiles of strains from unrelated sources or unrelated strains from the same source showed a particularly wide variety of plasmid profiles. Thus the possibility that epidemiologically-unrelated strains might possess similar profiles appears to be very low in this organism. Analysis of serologically-related strains from the same source revealed similar plasmid profiles in all the plasmid-bearing strains examined. A high proportion (71%) of fresh and well-characterized food-poisoning strains possessed plasmids of 6.2 kb in size (compared with 19% of non-food-poisoning strains). The possible role of these plasmids is discussed, since the structural gene encoding the enterotoxin type A was not present on any of the plasmids in the food-poisoning strains tested.

Molecular characterization of the VP4, VP6, VP7, and NSP4 genes of lapine rotaviruses identified in italy: emergence of a novel VP4 genotype

International Nuclear Information System (INIS)

Martella, Vito; Ciarlet, Max; Camarda, Antonio; Pratelli, Annamaria; Tempesta, Maria; Greco, Grazia; Cavalli, Alessandra; Elia, Gabriella; Decaro, Nicola; Terio, Valentina; Bozzo, Giancarlo; Camero, Michele; Buonavoglia, Canio

2003-01-01

The genes encoding the glycoprotein VP7, the VP8* trypsin-cleavage product of the protein VP4, a fragment of the protein VP6 associated with subgroup (SG) specificity, and the enterotoxin NSP4 of rotavirus strains identified in diarrheic fecal samples of rabbits in Italy were sequenced. The Italian lapine rotavirus (LRV) strains possessed a G3 VP7, SG I VP6, and KUN-like NSP4, a gene constellation typical of LRVs. One LRV strain (30/96), isolated in 1996, shared the closest amino acid (aa) identity (87-96%) with the P[14] genotype, composed of human and LRV strains. Conversely, three LRV strains (160/01, 229/01, and 308/01), identified in 2001, were highly identical (90-95%) among each other, but showed low aa identity (34-77%) to the VP8* genotype-specific sequences of representative rotavirus strains of all remaining P genotypes. This report confirms the worldwide genetic constellations of LRVs and identifies a novel VP4 genotype in rabbits, tentatively proposed as genotype P[22

Food compounds inhibit Staphylococcus aureus bacteria and the toxicity of Staphylococcus Enterotoxin A (SEA) associated with atopic dermatitis

Science.gov (United States)

Atopic dermatitis or eczema is characterized by skin rashes and itching is an inflammatory disease that affects 10-20% of children and 1-3% of adults. Staphylococcus aureus bacteria are present on the skin of nearly all patients with atopic dermatitis. Antibiotics that suppress colonization of S. au...

Detection of Staphylococcus Aureus Enterotoxin A and B Genes with PCR-EIA and a Hand-Held Electrochemical Sensor

National Research Council Canada - National Science Library

Aitichou, Mohamed; Henkins, Robert; Sultana, Afroz M; Ulrich, Robert G; Ibrahim, M. S

2004-01-01

... S. aureus DNA, and genomic DNA from Alcaligens, Bacillus, Bacteroides, Bordetella, Burkholderia, Clostridium, Comanonas, Enterobacter, Enterococcus, Escherichia, Francisella, Haemophilus, Klebsiella...

Staphylococcus aureus enterotoxin A (SEA) stimulates STAT3 activation and IL-17 expression in cutaneous T-cell lymphoma

DEFF Research Database (Denmark)

Willerslev-Olsen, Andreas; Krejsgaard, Thorbjørn; Lindahl, Lise Maria

2016-01-01

cells. The response is induced via IL-2 receptor common g chain cytokines and a Janus kinase 3 (JAK3)-dependent pathway in malignant T cells, and blocked by tofacitinib, a clinical-grade JAK3 inhibitor. In conclusion, we demonstrate that SEA induces cell cross talk-dependent activation of STAT3...

Monoclonal antibodies protect from Staphylococcal Enterotoxin K (SEK) induced toxic shock and sepsis by USA300 Staphylococcus aureus.

Science.gov (United States)

Aguilar, Jorge L; Varshney, Avanish K; Pechuan, Ximo; Dutta, Kaushik; Nosanchuk, Joshua D; Fries, Bettina C

2017-08-18

Staphylococcus aureus is a leading infectious cause of life-threatening disease in humans, yet there is currently no vaccine to combat this bacterium. The pathogenesis of S. aureus is mediated by a diverse array of protein toxins including a large family of secreted pyrogenic superantigens. Neutralization of superantigens, including SEB and TSST-1, has proven to be protective in several animal models of toxic shock and sepsis. We demonstrate, for the first time, that a far more prevalent staphylococcal superantigen, SEK, can also induce lethal shock in mice. Additionally, we describe monoclonal antibodies (mAbs) that inhibit SEK-induced mitogenicity as well as protect against SEK-induced lethality, and enhance survival from S. aureus septicemia in murine models. MAb-4G3 (IgG2b), mAb-5G2 (IgG1), and mAb-9H2 (IgG1), all inhibit SEK-induced proliferation and cytokine production of human immune cells. We then demonstrate that passive immunization with a combination of mAb-4G3 and mAb-5G4, 2 mAbs that do not compete for epitope(s) on SEK, significantly enhance survival in a murine model of SEK-induced toxic shock (p = 0.006). In the setting of sepsis, passive immunization with this combination of mAbs also significantly enhances survival in mice after challenge with CA-MRSA strain USA300 (p = 0.03). Furthermore, septic mice that received mAb treatment in conjunction with vancomycin exhibit less morbidity than mice treated with vancomycin alone. Taken together, these findings suggest that the contribution of SEK to S. aureus pathogenesis may be greater than previously appreciated, and that adjunctive therapy with passive immunotherapy against SEs may be beneficial.

Descripción de un brote de intoxicación alimentaria estafilocócica ocurrido en Las Rosas, Provincia de Santa Fe, Argentina

Directory of Open Access Journals (Sweden)

Aníbal A. Brizzio

2011-03-01

the foodborne outbreak revealed the presence of coagulase positive S. aureus subspecies aureus in samples from ingested and raw food, and from manipulators. Indicator microorganisms did not show significant levels and no other foodborne pathogen was isolated. Presence of staphylococcal enterotoxin-producing genes was subsequently investigated, and a positive result for enterotoxin B was shown in S. aureus strains isolated from a food handler as well as from food linked to the outbreak Moreover, these isolates showed 100% similarity by SmaI-PFGE. Timely notification together with coordinated sanitary measures and the availability of appropriate laboratory tools allowed to interrupt the chain of disease transmission by identifying risk and protective factors.

Use of single-enzyme amplified fragment length polymorphism for typing Clostridium perfringens isolated from diarrheic piglets Uso do polimorfismo do comprimento de fragmentos amplificados para tipagem de Clostridium perfringens isolados de suínos com diarréia

Directory of Open Access Journals (Sweden)

Luciane Tieko Shinya

2006-09-01

Full Text Available Clostridium perfringens is an important pathogen in human and veterinary medicine. In swine, the agent is responsible for necrotic enteritis and enterotoxemia characterized by diarrhea, weight loss, delayed development and, in some cases, death. In the present study amplified fragment length polymorphism analyses (AFLP was used to characterize 54 C. perfringens strains isolated from swine presenting diarrhea. Analysis of the results showed 29 distinct profiles with discriminatory index equal to 0.97. Partial correlation between the origin of the isolates and groups was drawn, and correlation was possible in only 18.5% of the samples. Characterization of the strains in biotypes (A, B, C, D and E, production of beta-2 toxin and enterotoxin were performed by means of the polymerase chain reaction (PCR. Biotypes A, C and D were observed among the strains analyzed. All samples were positive for presence of the gene encoding beta-2 toxin and negative for the gene encoding enterotoxin. AFLP have shown to be a simple, fast, low cost method with high discriminative power and good reproducibility, presenting a great potential in epidemiological studies involving C. perfringens strains of animal origin.Clostridium perfringens é um importante agente infeccioso em medicina veterinária e humana. Em suínos, o agente é responsável pela enterite necrótica e enterotoxemia, caracterizadas por diarréia, perda de peso, atraso no desenvolvimento e morte. No presente estudo foi utilizado o polimorfismo do comprimento de fragmentos amplificados (AFLP, para caracterizar 54 isolados de C. perfringens obtidos de suínos com diarréia. A análise dos resultados do AFLP demonstrou 29 perfis distintos com índice discriminatório igual a 0,97. A correlação entre a origem dos isolados e os agrupamentos obtidos foi parcial, sendo apenas possível a correlação total de 18,5% das amostras estudadas. A caracterização das cepas em biotipos (A, B, C, D e E, produ

Fatores de virulência de Bacillus thuringiensis: o que existe além das proteínas Cry

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Gislayne Vilas-Bôas

2012-03-01

Virulence Factors of Bacillus thuringiensis Berliner: Something Beyond of Cry Proteins? Abstract. The Cry proteins produced by the entomopathogenic bacterium Bacillus thuringiensis Berliner are widely known due to its high toxicity against a variety of insects. The mode of action of these proteins is specific and becomes B. thuringiensis-based products the most used in biological control programs of insect pests in agriculture and of important human disease vectors. However, while the Cry proteins are the best-known insect-specific virulence factor, strains of B. thuringiensis show also a wide range of other virulence factors, which allow the bacteria to achieve the hemolymph and colonize efficiently the insect host. Among these factors, we highlight the Vip proteins, Cyt, enterotoxins, hemolysins, phospholipases, proteases and enzymes of degradation, in addition to the recently described parasporin. This review explores the action of these virulence factors, as well as, the characterization and control of expression of their genes. Additionally, we discuss aspects related to the ecological niche of the bacteria with emphasis on the characteristics involved in the biosafety of the use of B. thuringiensis-based products for biological control of target insects.

Determination of enterotoxigenic Clostridium perfringens by detecting of the cpa and cpe genes in stool samples of human origin, associated to gastrointestinal disease

International Nuclear Information System (INIS)

Oropeza Barrios, Gletty

2014-01-01

A molecular methodology is provided to the Centro Nacional de Referencia de Bacteriologia (CNRB) of the Instituto Costarricense de Investigacion y Ensenanza en Nutricion y Salud. An opportune diagnosis is realized of enterotoxigenic Clostridium perfringens in stool samples of sporadic cases and cases associated to foodborne disease outbreaks. DNA extraction of the white microorganism was performed through the methodology implemented in the CNRB. The technique of polymerase chain reaction (PCR) were adapted and standardized to establish the identification of C. perfringens to species level and detection of cpe gene coding for enterotoxin. The sensitivity of the method was determined in a selective culture medium for C. perfringens (Tryptose sulfite cycloserine Agar). A detection limit of about 2,3 x 10 4 CFU/ml was reached for the cpe gene and at least 2,8 x 10 2 CFU/ml for the cpa gene. Retrospective analysis of 61 samples of diarrheal stool suspicious by C. perfringens is performed to evaluate the efficacy of the technique. Three outbreaks caused by C. perfringens were identified and a 10% of positivity in the samples were obtained analyzed during the period between July 2012-March 2014 [es

Screening of virulence genes in Staphylococcus aureus isolates from rabbits

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David Viana Martín

2015-09-01

Full Text Available Staphylococcus aureus is a versatile pathogen able to cause disease in both humans and animals. In rabbits, this bacterium infects animals of different ages, producing several purulent lesions. The ability of S. aureus to cause disease depends on a combination of virulence factors. The aim of this study was therefore to investigate the distribution of bacterial virulence determinants in 69 S. aureus isolates from rabbits. Some virulence factors (7 adhesins, 1 toxin and 1 protease were positive in all rabbit S. aureus isolates analysed, while others (1 adhesin and 10 toxins were always negative. The remaining virulence factors were more variable among isolates. An association between genotype and the different profiles of virulence factors was observed, but not with the type of lesion (P<0.05. One strain of each genotype was further analysed by multilocus sequence typing, generating ST121, ST96 and ST2951, determining a greater number of enterotoxins in ST121 isolates compared to ST96 and ST2951 isolates, which could justify the different pathogenicity between strains. 

Determination of staphylococcal exotoxins, SCCmec types, and genetic relatedness of Staphylococcus intermedius group isolates from veterinary staff, companion animals, and hospital environments in Korea

Science.gov (United States)

Youn, Jung-Ho; Ahn, Kuk Ju; Lim, Suk-Kyung

2011-01-01

The Staphylococcus (S.) intermedius group (SIG) has been a main research subject in recent years. S. pseudintermedius causes pyoderma and otitis in companion animals as well as foodborne diseases. To prevent SIG-associated infection and disease outbreaks, identification of both staphylococcal exotoxins and staphylococcal cassette chromosome mec (SCCmec) types among SIG isolates may be helpful. In this study, it was found that a single isolate (one out of 178 SIG isolates examined) harbored the canine enterotoxin SEC gene. However, the S. intermedius exfoliative toxin gene was found in 166 SIG isolates although the S. aureus-derived exfoliative toxin genes, such as eta, etb and etd, were not detected. SCCmec typing resulted in classifying one isolate as SCCmec type IV, 41 isolates as type V (including three S. intermedius isolates), and 10 isolates as non-classifiable. Genetic relatedness of all S. pseudintermedius isolates recovered from veterinary staff, companion animals, and hospital environments was determined by pulsed-field gel electrophoresis. Strains having the same band patterns were detected in S. pseudintermedius isolates collected at 13 and 18 months, suggesting possible colonization and/or expansion of a specific S. pseudintermedius strain in a veterinary hospital. PMID:21897094

Cloned polynucleotide and synthetic oligonucleotide probes used in colony hybridization are equally efficient in the identification of enterotoxigenic Escherichia coli

International Nuclear Information System (INIS)

Sommerfelt, H.; Kalland, K.H.; Raj, P.; Moseley, S.L.; Bhan, M.K.; Bjorvatn, B.

1988-01-01

Restriction endonuclease-generated polynucleotide and synthetically produced oligonucleotide gene probes used in colony hybridization assays proved to be efficient for the detection and differentiation of enterotoxigenic Escherichia coli. To compare their relative efficiencies, these two sets of probes were radiolabeled with 32 P and were applied to 74 strains of E. coli with known enterotoxin profiles and to 156 previously unexamined E. coli isolates. The enterotoxigenic bacteria Vibrio cholerae O1, Vibrio cholerae non-O1 (NAG), Yersinia enterocolitica, and E. coli harboring the plasmid vectors of the polynucleotide gene probes were examined for further evaluation of probe specificity. The two classes of probes showed a perfect concordance in their specific detection and differentiation of enterotoxigenic E. coli. In the analysis of six strains, the signal strength on autoradiography after hybridization with oligonucleotides was weaker than that obtained after hybridization with polynucleotide probes. The probes did not hybridize with DNA from V. cholerae O1, V. cholerae non-O1 (NAG), or Y. enterocolitica. The strains of E. coli harboring the plasmid vectors of the polynucleotide gene probes were, likewise, negative in the hybridization assays

Enterotoxigenic and non-enterotoxigenic Bacteroides fragilis from fecal microbiota of children

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Aline Ignacio

2015-01-01

Full Text Available Enterotoxigenic Bacteroides fragilis (ETBF is an important part of the human and animal intestinal microbiota and is commonly associated with diarrhea. ETBF strains produce an enterotoxin encoded by the bft gene located in the B. fragilispathogenicity island (BfPAI. Non-enterotoxigenic B. fragilis(NTBF strains lack the BfPAI and usually show two different genetic patterns, II and III, based on the absence or presence of a BfPAI-flanking region, respectively. The incidence of ETBF and NTBF strains in fecal samples isolated from children without acute diarrhea or any other intestinal disorders was determined. All 84 fecal samples evaluated were B. fragilis-positive by PCR, four of them harbored the bft gene, 27 contained the NTBF pattern III DNA sequence, and 52 were considered to be NTBF pattern II samples. One sample was positive for both ETBF and NTBF pattern III DNA sequences. All 19 B. fragilis strains isolated by the culture method were bft-negative, 9 belonged to pattern III and 10 to pattern II. We present an updated overview of the ETBF and NTBF incidence in the fecal microbiota of children from Sao Paulo City, Brazil.

Case report of a pustular dermatitis outbreak in sheep: Clinical and food safety considerations

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Mariana Roccaro

2018-04-01

Full Text Available The objective of this report is to describe an outbreak of pustular dermatitis in a flock of about 200 sheep, its clinical evolution and food safety implications. The onset of the symptoms was sudden and the lesions spread very quickly from ewe to ewe, so that in about 3 days almost all of the lactating sheep were stricken. Pustules from 5 different animals, six milk samples, two cheese samples, teat cup samples from the milking machine and farmer’s hands were analysed. A pure culture of Staphylococcus aureus, producing staphylococcal enterotoxin (SE C, was isolated from pustules. Milk and cheese showed a contamination by coagulase positive staphylococci <15 and 30 colony forming units respectively and the absence of SE. Farmer’s hands and teat cups samples resulted negative for coagulase positive staphylococci. Therapy with daily topical medicaments was prescribed and a prophylactic intervention was suggested by the administration of an autovaccine. The low level of milk and cheese contamination and the absence of SE in cheese supported the decision to not advise the farmer to recall cheese produced with milk from affected animals.

Functional insight into the C-terminal extension of halolysin SptA from haloarchaeon Natrinema sp. J7.

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Zhisheng Xu

Full Text Available Halolysin SptA from haloarchaeon Natrinema sp. J7 consists of a subtilisin-like catalytic domain and a C-terminal extension (CTE containing two cysteine residues. In this report, we have investigated the function of the CTE using recombinant enzymes expressed in Haloferax volcanii WFD11. Deletion of the CTE greatly reduced but did not abolish protease activity, which suggests that the CTE is not essential for enzyme folding. Mutational analysis suggests that residues Cys303 and Cys338 within the CTE form a disulfide bond that make this domain resistant to autocleavage and proteolysis under hypotonic conditions. Characterization of full-length and CTE-truncation enzymes indicates the CTE not only confers extra stability to the enzyme but also assists enzyme activity on protein substrates by facilitating binding at high salinities. Interestingly, homology modeling of the CTE yields a β-jelly roll-like structure similar to those seen in Claudin-binding domain of Clostridium perfringens enterotoxin (clostridial C-CPE and collagen binding domain (CBD, and the CTE also possesses collagen-binding activity, making it a potential candidate as an anchoring unit in drug delivery systems.

Zonula occludens toxin structure-function analysis. Identification of the fragment biologically active on tight junctions and of the zonulin receptor binding domain.

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Di Pierro, M; Lu, R; Uzzau, S; Wang, W; Margaretten, K; Pazzani, C; Maimone, F; Fasano, A

2001-06-01

Zonula occludens toxin (Zot) is an enterotoxin elaborated by Vibrio cholerae that increases intestinal permeability by interacting with a mammalian cell receptor with subsequent activation of intracellular signaling leading to the disassembly of the intercellular tight junctions. Zot localizes in the bacterial outer membrane of V. cholerae with subsequent cleavage and secretion of a carboxyl-terminal fragment in the host intestinal milieu. To identify the Zot domain(s) directly involved in the protein permeating effect, several zot gene deletion mutants were constructed and tested for their biological activity in the Ussing chamber assay and their ability to bind to the target receptor on intestinal epithelial cell cultures. The Zot biologically active domain was localized toward the carboxyl terminus of the protein and coincided with the predicted cleavage product generated by V. cholerae. This domain shared a putative receptor-binding motif with zonulin, the Zot mammalian analogue involved in tight junction modulation. Amino acid comparison between the Zot active fragment and zonulin, combined with site-directed mutagenesis experiments, confirmed the presence of an octapeptide receptor-binding domain toward the amino terminus of the processed Zot.

GAMMA Radiation Effect On Staphylococcus aureus (ATCC 19095) in Cheese MINAS FRESCALIRRADIATED

International Nuclear Information System (INIS)

Amaral Gurgel, M.S.C.C. DO; Spoto, M.H.F.; Domarco, R.E.

1999-01-01

Milk is an excellent medium of culture for development of Staphylococcus aureus. Gamma radiation can be an alternative method to guarantee the safety of the contaminated cheeses. The objective of this research was determine the effects of the gamma radiation on the resistance of S.aureus (ATCC 19095) in cheese M inas Frescal i rradiated. The cheeses elaborated in the Laboratory of Food Irradiation of CENA/USP, were contaminated during their production with 10 6 CFU/mL of culture of S.aureus (ATCC 19095). The cheeses were irradiated with 0; 1; 2; 3 and 4 kGy, maintained under refrigeration condition (5 0C ) and analyzed at 1, 7 and 14 days of storage. The evaluation microbiology was made through the S.aureus survival analysis using Baird Parker selective medium and confirmative test of coagulase, catalase and fermentation aerobics of the manitol. The capacity of enterotoxins production by irradiated S.aureus was detected by the method of Passive Reverse Agglutination Latex. Results showed that 3 kGy is enough to destroy the S.aureus and 2 kGy to inhibited its toxins production

Cloned polynucleotide and synthetic oligonucleotide probes used in colony hybridization are equally efficient in the identification of enterotoxigenic Escherichia coli

Energy Technology Data Exchange (ETDEWEB)

Sommerfelt, H.; Kalland, K.H.; Raj, P.; Moseley, S.L.; Bhan, M.K.; Bjorvatn, B.

1988-11-01

Restriction endonuclease-generated polynucleotide and synthetically produced oligonucleotide gene probes used in colony hybridization assays proved to be efficient for the detection and differentiation of enterotoxigenic Escherichia coli. To compare their relative efficiencies, these two sets of probes were radiolabeled with /sup 32/P and were applied to 74 strains of E. coli with known enterotoxin profiles and to 156 previously unexamined E. coli isolates. The enterotoxigenic bacteria Vibrio cholerae O1, Vibrio cholerae non-O1 (NAG), Yersinia enterocolitica, and E. coli harboring the plasmid vectors of the polynucleotide gene probes were examined for further evaluation of probe specificity. The two classes of probes showed a perfect concordance in their specific detection and differentiation of enterotoxigenic E. coli. In the analysis of six strains, the signal strength on autoradiography after hybridization with oligonucleotides was weaker than that obtained after hybridization with polynucleotide probes. The probes did not hybridize with DNA from V. cholerae O1, V. cholerae non-O1 (NAG), or Y. enterocolitica. The strains of E. coli harboring the plasmid vectors of the polynucleotide gene probes were, likewise, negative in the hybridization assays.

Ant-infecting Ophiocordyceps genomes reveal a high diversity of potential behavioral manipulation genes and a possible major role for enterotoxins

NARCIS (Netherlands)

De Bekker, Charissa; Ohm, Robin A.; Evans, Harry C.; Brachmann, Andreas; Hughes, David P

2017-01-01

Much can be gained from revealing the mechanisms fungal entomopathogens employ. Especially intriguing are fungal parasites that manipulate insect behavior because, presumably, they secrete a wealth of bioactive compounds. To gain more insight into their strategies, we compared the genomes of five

Toxin genotyping of Clostridium perfringens strains using a polymerase chain reaction protocol

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Elisabetta Di Giannatale

2010-03-01

Full Text Available A polymerase chain reaction protocol consisting of a multiplex to identify the cpa, cpb1, cpetx, cpi genes and a duplex to identify the cpe and cpb2 genes encoding for a, b1, e, i, enterotoxin and b2 toxins, respectively, was applied to DNA extracted from two collections of Clostridium perfringens strains. The first collection involved 19 isolates from rabbits. The second collection of 41 isolates came from routine necropsies. The cpa gene alone, or in association with the cpb2 gene, was detected in all DNA samples examined. The cpa gene, together with cpb2 gene, were detected in seven of the rabbit C. perfringens strains (36.8% and in nine isolates from necropsies (21.9%. The cpa gene was found in 63.2% of rabbit strains and 76.9% of strains from other animal species. In rabbits, the pathological lesions associated with C. perfringens detection were predominantly forms of non-inflammatory enteropathies. In other species, C. perfringens was mainly associated with congestive-haemorrhagic enteropathy, but also with fatal traumatic lesions, degenerative diseases and organs with post-mortem autolysis. No clear correlation was observed between detection of b2 toxin gene and species-specific pathological features.

Virulence Factors and Antibiotic Susceptibility of Staphylococcus aureus Isolates in Ready-to-Eat Foods: Detection of S. aureus Contamination and a High Prevalence of Virulence Genes

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Suat Moi Puah

2016-02-01

Full Text Available Staphylococcus aureus is one of the leading causes of food poisoning. Its pathogenicity results from the possession of virulence genes that produce different toxins which result in self-limiting to severe illness often requiring hospitalization. In this study of 200 sushi and sashimi samples, S. aureus contamination was confirmed in 26% of the food samples. The S. aureus isolates were further characterized for virulence genes and antibiotic susceptibility. A high incidence of virulence genes was identified in 96.2% of the isolates and 20 different virulence gene profiles were confirmed. DNA amplification showed that 30.8% (16/52 of the S. aureus carried at least one SE gene which causes staphylococcal food poisoning. The most common enterotoxin gene was seg (11.5% and the egc cluster was detected in 5.8% of the isolates. A combination of hla and hld was the most prevalent coexistence virulence genes and accounted for 59.6% of all isolates. Antibiotic resistance studies showed tetracycline resistance to be the most common at 28.8% while multi-drug resistance was found to be low at 3.8%. In conclusion, the high rate of S. aureus in the sampled sushi and sashimi indicates the need for food safety guidelines.

Lipid-derived free radical production in superantigen-induced interstitial pneumonia

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Miyakawa, Hisako; Mason, Ronald P.; Jiang, JinJie; Kadiiska, Maria B.

2009-01-01

We studied the free radical generation involved in the development of interstitial pneumonia (IP) in an animal model of autoimmune disease. We observed an electron spin resonance (ESR) spectrum of α-(4-pyridyl-1-oxide)-N-tert-butylnitrone (POBN) radical adducts detected in the lipid extract of lungs in autoimmune-prone mice after intratracheal instillation of staphylococcal enterotoxin B. The POBN adducts detected by ESR were paralleled by infiltration of macrophages and neutrophils in the bronchoalveolar lavage fluid. To further investigate the mechanism of free radical generation, mice were pretreated with the macrophage toxicant gadolinium chloride, which significantly suppressed the radical generation. Free radical generation was also decreased by pretreatment with the xanthine oxidase (XO) inhibitor allopurinol, the iron chelator Desferal, and the inducible nitric oxide synthase (iNOS) inhibitor 1400W. Histopathologically, these drugs significantly reduced both the cell infiltration to alveolar septal walls and the synthesis of pulmonary collagen fibers. Experiments with NADPH oxidase knockout mice showed that NADPH oxidase did not contribute to lipid radical generation. These results suggest that lipid-derived carbon-centered free radical production is important in the manifestation of IP and that a macrophage toxicant, an XO inhibitor, an iron chelator, and an iNOS inhibitor protect against both radical generation and the manifestation of IP. PMID:19376221

Characterisation of virulence genes in methicillin susceptible and resistant Staphylococcus aureus isolates from a paediatric population in a university hospital of Medellín, Colombia

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Judy Natalia Jiménez

2011-12-01

Full Text Available Virulence and antibiotic resistance are significant determinants of the types of infections caused by Staphylococcus aureus and paediatric groups remain among the most commonly affected populations. The goal of this study was to characterise virulence genes of methicillin-susceptible S. aureus (MSSA and methicillin-resistant S. aureus (MRSA strains isolated from a paediatric population of a Colombian University Hospital during 2009. Sixty MSSA and MRSA isolates were obtained from paediatric patients between zero-14 years. We identified the genes encoding virulence factors, which included Panton-Valentine leucocidine (PVL, staphylococcal enterotoxins A-E, exfoliative toxins A and B and toxic shock syndrome toxin 1. Typing of the staphylococcal chromosome cassette mec (SCCmec was performed in MRSA strains. The virulence genes were more diverse and frequent in MSSA than in MRSA isolates (83% vs. 73%. MRSA strains harboured SCCmec types IVc (60%, I (30%, IVa (7% and V (3%. SCCmec type IVc isolates frequently carried the PVL encoding genes and harboured virulence determinants resembling susceptible strains while SCCmec type I isolates were often negative. PVL was not exclusive to skin and soft tissue infections. As previously suggested, these differences in the distribution of virulence factor genes may be due to the fitness cost associated with methicillin resistance.

Eugenol reduces the expression of virulence-related exoproteins in Staphylococcus aureus.

Science.gov (United States)

Qiu, Jiazhang; Feng, Haihua; Lu, Jing; Xiang, Hua; Wang, Dacheng; Dong, Jing; Wang, Jianfeng; Wang, Xiaoliang; Liu, Juxiong; Deng, Xuming

2010-09-01

Eugenol, an essential oil component in plants, has been demonstrated to possess activity against both gram-positive and gram-negative bacteria. This study examined the influence that subinhibitory concentrations of eugenol may have on the expression of the major exotoxins produced by Staphylococcus aureus. The results from a tumor necrosis factor (TNF) release assay and a hemolysin assay indicated that S. aureus cultured with graded subinhibitory concentrations of eugenol (16 to 128 microg/ml) dose dependently decreased the TNF-inducing and hemolytic activities of culture supernatants. Western blot analysis showed that eugenol significantly reduced the production of staphylococcal enterotoxin A (SEA), SEB, and toxic shock syndrome toxin 1 (the key exotoxins to induce TNF release), as well as the expression of alpha-hemolysin (the major hemolysin to cause hemolysis). In addition, this suppression was also evaluated at the transcriptional level via real-time reverse transcription (RT)-PCR analysis. The transcriptional analysis indicated that 128 microg/ml of eugenol remarkably repressed the transcription of the S. aureus sea, seb, tst, and hla genes. According to these results, eugenol has the potential to be rationally applied on food products as a novel food antimicrobial agent both to inhibit the growth of bacteria and to suppress the production of exotoxins by S. aureus.

Characterisation of virulence genes in methicillin susceptible and resistant Staphylococcus aureus isolates from a paediatric population in a university hospital of Medellín, Colombia.

Science.gov (United States)

Jiménez, Judy Natalia; Ocampo, Ana María; Vanegas, Johanna Marcela; Rodríguez, Erika Andrea; Garcés, Carlos Guillermo; Patiño, Luz Adriana; Ospina, Sigifredo; Correa, Margarita María

2011-12-01

Virulence and antibiotic resistance are significant determinants of the types of infections caused by Staphylococcus aureus and paediatric groups remain among the most commonly affected populations. The goal of this study was to characterise virulence genes of methicillin-susceptible S. aureus (MSSA) and methicillin-resistant S. aureus (MRSA) strains isolated from a paediatric population of a Colombian University Hospital during 2009. Sixty MSSA and MRSA isolates were obtained from paediatric patients between zero-14 years. We identified the genes encoding virulence factors, which included Panton-Valentine leucocidine (PVL), staphylococcal enterotoxins A-E, exfoliative toxins A and B and toxic shock syndrome toxin 1. Typing of the staphylococcal chromosome cassette mec (SCCmec) was performed in MRSA strains. The virulence genes were more diverse and frequent in MSSA than in MRSA isolates (83% vs. 73%). MRSA strains harboured SCCmec types IVc (60%), I (30%), IVa (7%) and V (3%). SCCmec type IVc isolates frequently carried the PVL encoding genes and harboured virulence determinants resembling susceptible strains while SCCmec type I isolates were often negative. PVL was not exclusive to skin and soft tissue infections. As previously suggested, these differences in the distribution of virulence factor genes may be due to the fitness cost associated with methicillin resistance.

Characteristics of Staphylococcus aureus isolated from acute, sub-acute and sub-clinical staphylococcosis in rabbits

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Rajeshkumar J. Tirpude

2012-12-01

Full Text Available Staphylococcus aureus bacteria isolated from different clinical presentations of staphylococcosis in rabbits were examined for the production of various virulence factors using biochemical and immunological tests. In the total of 106 S. aureus isolates; toxic shock syndrome toxin-1, staphylococcal enterotoxin-C, DNase, α-haemolysin, β-haemolysin, δ-haemolysin, protein A and clumping factor were observed with a frequency of 33.2, 16.98, 83.96, 69.81, 36.79, 100, 78.30 and 54.72 percent, respectively. No SE-A, SE-B and SE-D producing isolates were recovered in this study. All the S. aureus isolates from acute staphylococcosis produced TSST-1, SE-C and protein A. While δ–haemolysin and clumping factor were not detected in any acute isolates, these factors were observed at a relatively higher frequency in isolates from sub-acute and sub-clinical staphylococcosis. Coagulase type III was observed more predominantly with a frequency of 45.28%, while coagulase types V and VII were not observed in any isolate. Most of the virulence factors belonged to coagulase type III followed by type VI. TSST-1 and SE-C along with coagulase types III and VI could be correlated with the acute and sub-acute staphylococcal infections in rabbits in this study.

Mechanisms of Action and Cell Death Associated with Clostridium perfringens Toxins

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Mauricio A. Navarro

2018-05-01

Full Text Available Clostridium perfringens uses its large arsenal of protein toxins to produce histotoxic, neurologic and intestinal infections in humans and animals. The major toxins involved in diseases are alpha (CPA, beta (CPB, epsilon (ETX, iota (ITX, enterotoxin (CPE, and necrotic B-like (NetB toxins. CPA is the main virulence factor involved in gas gangrene in humans, whereas its role in animal diseases is limited and controversial. CPB is responsible for necrotizing enteritis and enterotoxemia, mostly in neonatal individuals of many animal species, including humans. ETX is the main toxin involved in enterotoxemia of sheep and goats. ITX has been implicated in cases of enteritis in rabbits and other animal species; however, its specific role in causing disease has not been proved. CPE is responsible for human food-poisoning and non-foodborne C. perfringens-mediated diarrhea. NetB is the cause of necrotic enteritis in chickens. In most cases, host–toxin interaction starts on the plasma membrane of target cells via specific receptors, resulting in the activation of intracellular pathways with a variety of effects, commonly including cell death. In general, the molecular mechanisms of cell death associated with C. perfringens toxins involve features of apoptosis, necrosis and/or necroptosis.

Viability of Staphylococcus xylosus isolated from artisanal sausages for application as starter cultures in meat products Viabilidade de Staphylococcus xylosus isoladas de embutidos artesanais para aplicação como cultivos iniciadores em produtos cárneos

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Ângela Maria Fiorentini

2009-03-01

Full Text Available Viability of Staphylococcus xylosus isolated from artisanal sausages for application as starter cultures in meat products Viability of Staphylococcus xylosus strains AD1 and U5isolated from natural fermented sausages was investigated as starter cultures in fermented sausages produced in the South Region of Brazil. The study demonstrated that the Staphylococcus xylosus strains AD1 and U5showed significant growth during fermentation, stability over freeze-dried process, negative reaction for staphylococcal enterotoxins and viability for using as a single-strain culture or associated with lactic acid bacteria for production of fermented sausages.Investigamos a viabilidade de cepas de Staphylococcus xylosus (AD1 e U5 isoladas de embutidos com fermentação natural, para aplicação como cultivos iniciadores em embutidos fermentados produzidos na Região Sul do Brasil. O estudo demonstrou que cepas de Staphylococcus xylosus (AD1 e U5 apresentaram crescimento significativo durante a fermentação, estabilidade no processo de liofilização e conservação, ausência de produção de enterotoxinas e viabilidade para aplicação como cultivo iniciador simples ou associado com bactérias lácticas na elaboração de embutidos fermentados.

Bacterial Toxins for Oncoleaking Suicidal Cancer Gene Therapy.

Science.gov (United States)

Pahle, Jessica; Walther, Wolfgang

For suicide gene therapy, initially prodrug-converting enzymes (gene-directed enzyme-producing therapy, GDEPT) were employed to intracellularly metabolize non-toxic prodrugs into toxic compounds, leading to the effective suicidal killing of the transfected tumor cells. In this regard, the suicide gene therapy has demonstrated its potential for efficient tumor eradication. Numerous suicide genes of viral or bacterial origin were isolated, characterized, and extensively tested in vitro and in vivo, demonstrating their therapeutic potential even in clinical trials to treat cancers of different entities. Apart from this, growing efforts are made to generate more targeted and more effective suicide gene systems for cancer gene therapy. In this regard, bacterial toxins are an alternative to the classical GDEPT strategy, which add to the broad spectrum of different suicide approaches. In this context, lytic bacterial toxins, such as streptolysin O (SLO) or the claudin-targeted Clostridium perfringens enterotoxin (CPE) represent attractive new types of suicide oncoleaking genes. They permit as pore-forming proteins rapid and also selective toxicity toward a broad range of cancers. In this chapter, we describe the generation and use of SLO as well as of CPE-based gene therapies for the effective tumor cell eradication as promising, novel suicide gene approach particularly for treatment of therapy refractory tumors.

Bystander protein protects potential vaccine-targeting ligands against intestinal proteolysis.

Science.gov (United States)

Reuter, Fabian; Bade, Steffen; Hirst, Timothy R; Frey, Andreas

2009-07-20

Endowing mucosal vaccines with ligands that target antigen to mucosal lymphoid tissues may improve immunization efficacy provided that the ligands withstand the proteolytic environment of the gastro-intestinal tract until they reach their destination. Our aim was to investigate whether and how three renowned ligands - Ulex europaeus agglutinin I and the B subunits of cholera toxin and E. coli heat-labile enterotoxin - master this challenge. We assessed the digestive power of natural murine intestinal fluid (natIF) using assays for trypsin, chymotrypsin and pancreatic elastase along with a test for nonspecific proteolysis. The natIF was compared with simulated murine intestinal fluid (simIF) that resembled the trypsin, chymotrypsin and elastase activities of its natural counterpart but lacked or contained albumins as additional protease substrates. The ligands were exposed to the digestive fluids and degradation was determined. The studies revealed that (i) the three pancreatic endoproteases constitute only one third of the total protease activity of natIF and (ii) the ligands resist proteolysis in natIF and protein-enriched simIF over 3 h but (iii) are partially destroyed in simIF that lacks additional protease substrate. We assume that the proteins of natIF are preferred substrates for the intestinal proteases and thus can protect vaccine-targeting ligands from destruction.

Screening Antibacterial Agent from Crude Extract of Marine-Derived Fungi Associated with Soft Corals against MDR-Staphylococcus haemolyticus

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Sabdaningsih, A.; Cristianawati, O.; Sibero, M. T.; Nuryadi, H.; Radjasa, O. K.; Sabdono, A.; Trianto, A.

2017-02-01

Multidrug resistant Staphylococcus haemolyticus is a Gram-positive bacteria and member of coagulase negative staphylococci (CoNS) which has the highest level of antimicrobial resistance. This nosocomial pathogen due to skin or soft tissue infections, bacteremia, septicemia, peritonitis, otitis media, meningitis and urinary tract infections. The ability to produce enterotoxins, hemolysins, biofilm, and cytotoxins could be an important characteristic for the successful of infection. Marine-derived fungi have potency as a continuous supply of bioactive compound. The aim of this research was screening antibacterial agent from crude extracts of marine-derived fungi associated with soft corals against MDR-S. haemolyticus. Among 23 isolates of marine-derived fungi isolated from 7 soft corals, there were 4 isolates active against MDR-S. haemolyticus. The screening was conducted by using agar plug diffusion method. Isolate PPSC-27-A4 had the highest antibacterial activity with diameter 23±9,6 mm. The crude extract from this isolate had been confirmed to antibacterial susceptibility test and it had the highest antibacterial activity in 12.2 mm with concentration of 300μg/ml from mycelia extract. PPSC-27-A4 had been characterized in molecular, based on the sequence analysis of 18S rRNA, PPSC-27-A4 isolate was identified as Trichoderma longibrachiatum.

Review article: anti-inflammatory mechanisms of action of Saccharomyces boulardii.

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Pothoulakis, C

2009-10-15

Saccharomyces boulardii, a well-studied probiotic, can be effective in inflammatory gastrointestinal diseases with diverse pathophysiology, such as inflammatory bowel disease (IBD), and bacterially mediated or enterotoxin-mediated diarrhoea and inflammation. To discuss the mechanisms of action involved in the intestinal anti-inflammatory action of S. boulardii. Review of the literature related to the anti-inflammatory effects of this probiotic. Several mechanisms of action have been identified directed against the host and pathogenic microorganisms. S. boulardii and S. boulardii secreted-protein(s) inhibit production of proinflammatory cytokines by interfering with the global mediator of inflammation nuclear factor kappaB, and modulating the activity of the mitogen-activated protein kinases ERK1/2 and p38. S. boulardii activates expression of peroxisome proliferator-activated receptor-gamma (PPAR-gamma) that protects from gut inflammation and IBD. S. boulardii also suppresses 'bacteria overgrowth' and host cell adherence, releases a protease that cleaves C. difficile toxin A and its intestinal receptor and stimulates antibody production against toxin A. Recent results indicate that S. boulardii may interfere with IBD pathogenesis by trapping T cells in mesenteric lymph nodes. The multiple anti-inflammatory mechanisms exerted by S. boulardii provide molecular explanations supporting its effectiveness in intestinal inflammatory states.

Review article: Anti-inflammatory mechanisms of action of Saccharomyces boulardii

Science.gov (United States)

Pothoulakis, C.

2009-01-01

SUMMARY Background Saccharomyces boulardii (S. boulardii), a well-studied probiotic, can be effective in inflammatory gastrointestinal diseases with diverse pathophysiology, such as Inflammatory Bowel Disease (IBD), and bacterially – or enterotoxin-mediated diarrhea and inflammation. Aim Discuss the mechanisms of action involved in the intestinal anti-inflammatory action of S. boulardii Methods Review of the literature related to the anti-inflammatory effects of this probiotic. Results Several mechanisms of action have been identified directed against the host and pathogenic microorganisms. S. boulardii and S. boulardii secreted protein(s) inhibit production of proinflammatory cytokines by interfering with the global mediator of inflammation nuclear factor κB, and modulating the activity of the mitogen-activated protein kinases ERK1/2 and p38. S. boulardii activates expression of peroxisome proliferator-activated receptor-gamma (PPAR-γ) that protects from gut inflammation and IBD. S. boulardii also suppresses “bacteria overgrowth” and host cell adherence, releases a protease that cleaves C. difficile toxin A and its intestinal receptor, and stimulates antibody production against toxin A. Recent results indicate that S. boulardii may interfere with IBD pathogenesis by trapping T cells in mesenteric lymph nodes. Conclusions The multiple anti-inflammatory mechanisms exerted by S. boulardii provide molecular explanations supporting its effectiveness in intestinal inflammatory states. PMID:19706150

Development of an ELISA microarray assay for the sensitive and simultaneous detection of ten biodefense toxins.

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Jenko, Kathryn; Zhang, Yanfeng; Kostenko, Yulia; Fan, Yongfeng; Garcia-Rodriguez, Consuelo; Lou, Jianlong; Marks, James D.; Varnum, Susan M.

2014-10-21

Plant and microbial toxins are considered bioterrorism threat agents because of their extreme toxicity and/or ease of availability. Additionally, some of these toxins are increasingly responsible for accidental food poisonings. The current study utilized an ELISA-based protein antibody microarray for the multiplexed detection of ten biothreat toxins, botulinum neurotoxins (BoNT) A, B, C, D, E, F, ricin, shiga toxins 1 and 2 (Stx), and staphylococcus enterotoxin B (SEB), in buffer and complex biological matrices. The multiplexed assay displayed a sensitivity of 1.3 pg/mL (BoNT/A, BoNT/B, SEB, Stx-1 and Stx-2), 3.3 pg/mL (BoNT/C, BoNT/E, BoNT/F) and 8.2 pg/mL (BoNT/D, ricin). All assays demonstrated high accuracy (75-120 percent recovery) and reproducibility (most coefficients of variation < 20%). Quantification curves for the ten toxins were also evaluated in clinical samples (serum, plasma, nasal fluid, saliva, stool, and urine) and environmental samples (apple juice, milk and baby food) with overall minimal matrix effects. The multiplex assays were highly specific, with little crossreactivity observed between the selected toxin antibodies. The results demonstrate a multiplex microarray that improves current immunoassay sensitivity for biological warfare agents in buffer, clinical, and environmental samples.

Handling Practices During Distribution of Kinnow-Mandarins (Citrus nobilis x Citrus deliciosa Used for Preparation of Fresh-Squeezed Juices and their Effects on Microbiological Safety

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M. Ghosh

2005-01-01

Full Text Available The objective of this study was to assess the microbiological safety of Kinnow-mandarins (Citrus nobilis X Citrus deliciosa used for preparation of fresh squeezed juices by street vendors. One hundred and fifty Kinnow mandarin samples were collected from different points in the distribution chain from the same lot, washed and aliquots of the wash samples were analyzed for total aerobes, Staphylococcus, total and fecal coliforms, Salmonella and Shigella. Although, there was no notable increase in the total aerobic, total or fecal coliform counts, total staphylococcal counts increased significantly (P<0.05 during distribution to street vendors; seventy-two samples from the street vendors showed the presence of coagulase positive Staphylococcus aureus, twenty-three of these produced enterotoxins B and C. Salmonella and Shigella were not detected in any of the samples. Sources of high numbers of enterotoxigenic S. aureus were traced to unhygienic manual handling by middle level buyers and by street vendors. The results of our study demonstrate the poor microbiological quality of Kinnow-mandarins, the possible entry points of contaminants in the distribution chain of these fruits and unhygienic vending practices. Appropriate intervention measures are needed to ensure safe fresh squeezed juices for consumers.

Toxinotyping and antimicrobial susceptibility of enterotoxigenic Clostridium perfringens isolates from mutton, beef and chicken meat.

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Khan, Madiha; Nazir, Jawad; Anjum, Aftab Ahmad; Ahmad, Mansur-Ud-Din; Nawaz, Muhammad; Shabbir, Muhammad Zubair

2015-08-01

A total of 300 meat samples comprising mutton, beef, and chicken meat (n = 100) collected from either local butcher shops or large meat outlets situated at various areas of Lahore City located in Punjab province of Pakistan were tested for the isolation of Clostridium perfringens. Prevalence of the organism was highest in the chicken (6 %) followed by mutton (5 %) and beef (1 %). Contamination level was high (10/150) in the samples collected from local butcher shops in comparison to the samples collected from large meat outlets (2/150). All of the raw meat samples were negative for the presence of alpha, beta and epsilon toxins of C. perfringens as detected through ELISA. Out of a total number of 12 isolates only half were capable of producing enterotoxins when cultured in trypticase glucose yeast (TGY) broth. Toxinotyping of the isolates showed that 3 were of type A while one each of the remaining three belonged to type B, C, and D. Antibiotic susceptibility testing of the toxin producing isolates revealed that C. perfringens were susceptible to chloramphenicol, ciprofloxacin, metronidazole, and ceftriaxone. All of the other drugs were relatively less effective with a least activity of amoxicillin against the isolates.

Surface and virulence properties of environmental Vibrio cholerae non-O1 from Albufera Lake (Valencia, Spain).

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Amaro, C; Toranzo, A E; González, E A; Blanco, J; Pujalte, M J; Aznar, R; Garay, E

1990-04-01

A total of 140 environmental Vibrio cholerae non-O1 isolates, together with several culture collection strains from both environmental and clinical sources, were studied in relation to hemagglutination, surface hydrophobicity, and the enzymatic, hemolytic, cytotoxic, and enterotoxic activities of their extracellular products. A total of 78 and 62% of the strains produced hemagglutinins and exohemagglutinins, respectively. Four different hemagglutinating and two exohemagglutinating activities were found by using eight sugars in the inhibition assays. Cell-bound mannose-sensitive hemagglutination was detected mainly in chicken blood, whereas fucose-sensitive hemagglutination was recorded only in human blood. Cell-bound hemagglutinin resistant to all sugars tested was the only one related to surface hydrophobicity. The surface properties varied along the growth curves. The non-O1 strains displayed strong enzymatic and hemolytic activities, except for esculin hydrolysis. Of 26 non-O1 isolates selected for cytotoxin and enterotoxin production, 23 showed a wide spectrum of cytotoxic effects on cell lines of poikilothermic and homoiothermic species, but they were weakly enterotoxigenic in the infant mouse test. All extracellular products of cytotoxic strains were proteolytic, lipolytic, and hemolytic, and a high percentage produced hemagglutination of chicken blood. The cytotoxic factors in the non-O1 strains analyzed were not R plasmid mediated.

Persistent production of TH2-type cytokines and polyclonal B cell activation after chronic administration of staphylococcal enterotoxin B in mice

NARCIS (Netherlands)

Florquin, S.; Amraoui, Z.; Goldman, M.

1996-01-01

In order to study the immunopathological consequences of repeated exposure to bacterial superantigens, we evaluated the production of cytokines, the profile of serum immunoglobulins and the tissue damage in BALB/c mice injected twice a week for 3 weeks with 50 micrograms of staphylococcal

X-ray crystal structure of the passenger domain of plasmid encoded toxin(Pet), an autotransporter enterotoxin from enteroaggregative Escherichia coli (EAEC)

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Domingo Meza-Aguilar, J. [Departamento de Salud Pública Facultad de Medicina UNAM, Ciudad Universitaria Coyoacán 04510, D.F. (Mexico); Laboratorio de Patogenicidad Bacteriana, Unidad de Hemato Oncología e Investigación, Hospital Infantil de México Federico Gómez 06720, D.F. (Mexico); Fromme, Petra [Department of Chemistry and Biochemistry, Arizona State University, Physical Sciences BLDG D-102, Tempe, AZ 85287 (United States); Torres-Larios, Alfredo [Instituto de Fisiología Celular UNAM, Ciudad Universitaria Coyoacán 04510, D.F. (Mexico); Mendoza-Hernández, Guillermo [Instituto de Química UNAM, Ciudad Universitaria Coyoacán 04510, D.F (Mexico); Hernandez-Chiñas, Ulises [Departamento de Salud Pública Facultad de Medicina UNAM, Ciudad Universitaria Coyoacán 04510, D.F. (Mexico); Laboratorio de Patogenicidad Bacteriana, Unidad de Hemato Oncología e Investigación, Hospital Infantil de México Federico Gómez 06720, D.F. (Mexico); Arreguin-Espinosa de los Monteros, Roberto A. [Instituto de Química UNAM, Ciudad Universitaria Coyoacán 04510, D.F (Mexico); and others

2014-03-07

Highlights: • X-ray crystal structure of the passenger domain of Plasmid encoded toxin at 2.3 Å. • Structural differences between Pet passenger domain and EspP protein are described. • High flexibility of the C-terminal beta helix is structurally assigned. - Abstract: Autotransporters (ATs) represent a superfamily of proteins produced by a variety of pathogenic bacteria, which include the pathogenic groups of Escherichia coli (E. coli) associated with gastrointestinal and urinary tract infections. We present the first X-ray structure of the passenger domain from the Plasmid-encoded toxin (Pet) a 100 kDa protein at 2.3 Å resolution which is a cause of acute diarrhea in both developing and industrialized countries. Pet is a cytoskeleton-altering toxin that induces loss of actin stress fibers. While Pet (pdb code: 4OM9) shows only a sequence identity of 50% compared to the closest related protein sequence, extracellular serine protease plasmid (EspP) the structural features of both proteins are conserved. A closer structural look reveals that Pet contains a β-pleaded sheet at the sequence region of residues 181–190, the corresponding structural domain in EspP consists of a coiled loop. Secondary, the Pet passenger domain features a more pronounced beta sheet between residues 135 and 143 compared to the structure of EspP.

An outbreak of food poisoning due to egg yolk reaction-negative Staphylococcus aureus.

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Miwa, N; Kawamura, A; Masuda, T; Akiyama, M

2001-03-20

An outbreak of staphylococcal food poisoning due to an egg yolk (EY) reaction-negative strain occurred in Japan. Twenty-one of 53 dam construction workers who ate boxed lunches prepared at their company cafeteria became ill, and eight required hospital treatment. The outbreak showed a typical incubation time (1.5-4 h with a median time of 2.7 h) and symptoms (vomiting and diarrhea) of staphylococcal food poisoning. Staphylococcus aureus, which produces staphylococcal enterotoxin (SE) A, was isolated from four fecal specimens of eight patients tested. Scrambled egg in the boxed lunches contained 20-40 ng/g of SEA, and 3.0 x 10(9)/g of viable S. aureus cells that produced this toxin. All isolates from patients and the food were EY reaction-negative, coagulase type II, and showed the same restriction fragment length polymorphism (RFLP) pattern. We concluded that the outbreak was caused by scrambled egg contaminated with EY reaction-negative S. aureus. In Japan, outbreaks of staphylococcal food poisoning are mainly caused by EY reaction-positive S. aureus, and EY reaction-negative colonies grown on agar plates containing EY are usually not analyzed further for detection of S. aureus. The present outbreak suggested that EY reaction-negative isolates should be subjected to further analysis to detect the causative agents of staphylococcal food poisoning.

Antagonistics against pathogenic Bacillus cereus in milk fermentation by Lactobacillus plantarum ZDY2013 and its anti-adhesion effect on Caco-2 cells against pathogens.

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Zhang, Zhihong; Tao, Xueying; Shah, Nagendra P; Wei, Hua

2016-04-01

Lactobacillus plantarum ZDY2013 is a potential probiotic isolated from fermented bean acid. In this study, we aimed to evaluate the in vitro antimicrobial activity of this organism against Bacillus cereus in milk fermentation, the antiadhesion ability on intestinal epithelial cells, as well as its ability to abrogate the cytotoxic effect and expression levels of genes. We found no antimicrobial activity produced by L. plantarum once the pH was adjusted to 6.0 and 7.0. The pH decreased continuously when L. plantarum and B. cereus were co-incubated during milk fermentation, which caused a decrease in the B. cereus counts. Antiadhesion assays showed that L. plantarum can significantly inhibit the adhesion of enterotoxin-producing B. cereus ATCC14579 and pathogenic B. cereus HN001 by inhibition, competition, and displacement. The supernatants of B. cereus, either alone or in conjunction with L. plantarum, caused damage to the membrane integrity of Caco-2 cells to release lactate dehydrogenase. In addition, L. plantarum tended to attenuate proinflammatory cytokine and oxidative stress gene expression on Caco-2 cells, inducing with B. cereus HN001 supernatants. This study provided systematic insights into the antagonistic effect of L. plantarum ZDY2013, and the information may be helpful to explore potential control measures for preventing food poisoning by lactic acid bacteria. Copyright © 2016 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Evidence of functional cell-mediated immune responses to nontypeable Haemophilus influenzae in otitis-prone children

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Seppanen, Elke; Tan, Dino; Corscadden, Karli J.; Currie, Andrew J.; Richmond, Peter C.; Thornton, Ruth B.

2018-01-01

Otitis media (OM) remains a common paediatric disease, despite advances in vaccinology. Susceptibility to recurrent acute OM (rAOM) has been postulated to involve defective cell-mediated immune responses to common otopathogenic bacteria. We compared the composition of peripheral blood mononuclear cells (PBMC) from 20 children with a history of rAOM (otitis-prone) and 20 healthy non-otitis-prone controls, and assessed innate and cell-mediated immune responses to the major otopathogen nontypeable Haemophilus influenzae (NTHi). NTHi was a potent stimulator of inflammatory cytokine secretion from PBMC within 4 hours, with no difference in cytokine levels produced between PBMC from cases or controls. In the absence of antigen stimulation, otitis-prone children had more circulating Natural Killer (NK) cells (potitis-prone and non-otitis-prone children (potitis-prone children are functional and respond to NTHi. CD8+ T cells and NK cells from both cases and controls produced IFNγ in response to polyclonal stimulus (Staphylococcal enterotoxin B; SEB), with more IFNγ+ CD8+ T cells present in cases than controls (pOtitis-prone children had more circulating IFNγ-producing NK cells (potitis-prone children mounted innate and T cell-mediated responses to NTHi challenge that were comparable to healthy children. These data provide evidence that otitis-prone children do not have impaired functional cell mediated immunity. PMID:29621281

Pathogen-Reactive T Helper Cell Analysis in the Pig

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Friederike Ebner

2017-05-01

Full Text Available There is growing interest in studying host–pathogen interactions in human-relevant large animal models such as the pig. Despite the progress in developing immunological reagents for porcine T cell research, there is an urgent need to directly assess pathogen-specific T cells—an extremely rare population of cells, but of upmost importance in orchestrating the host immune response to a given pathogen. Here, we established that the activation marker CD154 (CD40L, known from human and mouse studies, identifies also porcine antigen-reactive CD4+ T lymphocytes. CD154 expression was upregulated early after antigen encounter and CD4+CD154+ antigen-reactive T cells coexpressed cytokines. Antigen-induced expansion and autologous restimulation enabled a time- and dose-resolved analysis of CD154 regulation and a significantly increased resolution in phenotypic profiling of antigen-responsive cells. CD154 expression identified T cells responding to staphylococcal Enterotoxin B superantigen stimulation as well as T cells responding to the fungus Candida albicans and T cells specific for a highly prevalent intestinal parasite, the nematode Ascaris suum during acute and trickle infection. Antigen-reactive T cells were further detected after immunization of pigs with a single recombinant bacterial antigen of Streptococcus suis only. Thus, our study offers new ways to study antigen-specific T lymphocytes in the pig and their contribution to host–pathogen interactions.

Isolation of anti-toxin single domain antibodies from a semi-synthetic spiny dogfish shark display library

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Goldman Ellen R

2007-11-01

Full Text Available Abstract Background Shark heavy chain antibody, also called new antigen receptor (NAR, consists of one single Variable domain (VH, containing only two complementarity-determining regions (CDRs. The antigen binding affinity and specificity are mainly determined by these two CDRs. The good solubility, excellent thermal stability and complex sequence variation of small single domain antibodies (sdAbs make them attractive alternatives to conventional antibodies. In this report, we construct and characterize a diversity enhanced semi-synthetic NAR V display library based on naturally occurring NAR V sequences. Results A semi-synthetic shark sdAb display library with a complexity close to 1e9 was constructed. This was achieved by introducing size and sequence variations in CDR3 using randomized CDR3 primers of three different lengths. Binders against three toxins, staphylococcal enterotoxin B (SEB, ricin, and botulinum toxin A (BoNT/A complex toxoid, were isolated from panning the display library. Soluble sdAbs from selected binders were purified and evaluated using direct binding and thermal stability assays on the Luminex 100. In addition, sandwich assays using sdAb as the reporter element were developed to demonstrate their utility for future sensor applications. Conclusion We demonstrated the utility of a newly created hyper diversified shark NAR displayed library to serve as a source of thermal stable sdAbs against a variety of toxins.

Application of Probiotics in Poultry Production

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Martin Král

2012-05-01

Full Text Available In the last years probiotics have constantly increased in importance and aroused growing interest in animal nutrition. Probiotics are live microorganisms thought to be beneficial to the host organism. The bacteria of the probiotic attach to the intestinal mucosa, thereby forming a physical barrier that blocks the attachment of pathogenic bacteria. The mode of action of probiotics in poultry includes maintaining normal intestinal microflora by competitive exclusion and antagonism, altering metabolism by increasing digestive enzyme activity and decreasing bacterial enzyme activity and ammonia production, improving feed intake and digestion and neutralizing enterotoxins and stimulating the immune system. In experiment we research effect of probiotic on the performance of broiler chickens. A total number of 200 one day old broiler chickens were distributed to two dietary groups. Broiler chickens in control group were fed with standard feed mixture and experimental group with probiotics mixed with feed mixture. Body weight and GIT pH were recorded. Average body weight on the end of experiment in experimental group was 1493.6 g and 1689.6 g in control group. Average pH in experimental group was 2.79 in stomach, 6.28 in small intestine, 6.81 and 6.89 in caecum. In control group was average pH 3.54 in stomach, 6.41 in small intestine, 6.74 and 6.80 in caecum.

Identification of capsule, biofilm, lateral flagellum, and type IV pili in Vibrio mimicus strains.

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Tercero-Alburo, J J; González-Márquez, H; Bonilla-González, E; Quiñones-Ramírez, E I; Vázquez-Salinas, C

2014-11-01

Vibrio mimicus is a bacterium that causes gastroenteritis; it is closely related to Vibrio cholerae, and can cause acute diarrhea like cholera- or dysentery-type diarrhea. It is distributed worldwide. Factors associated with virulence (such as hemolysins, enterotoxins, proteases, phospholipases, aerobactin, and hemagglutinin) have been identified; however, its pathogenicity mechanism is still unknown. In pathogenic Vibrio species such as V. cholerae, Vibrio. parahaemolyticus and Vibrio vulnificus, capsule, biofilms, lateral flagellum, and type IV pili are structures described as essential for pathogenicity. These structures had not been described in V. mimicus until this work. We used 20 V. mimicus strains isolated from water (6), oyster (9), and fish (5) samples and we were able to identify the capsule, biofilm, lateral flagellum, and type IV pili through phenotypic tests, electron microscopy, PCR, and sequencing. In all tested strains, we observed and identified the presence of capsular exopolysaccharide, biofilm formation in an in vitro model, as well as swarming, multiple flagellation, and pili. In addition, we identified homologous genes to those described in other bacteria of the genus in which these structures have been found. Identification of these structures in V. mimicus is a contribution to the biology of this organism and can help to reveal its pathogenic behavior. Copyright © 2014 Elsevier Ltd. All rights reserved.

Stress tolerant virulent strains of Cronobacter sakazakii from food

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Md Fakruddin

2014-01-01

Full Text Available BACKGROUND: Cronobacter sakazakii is considered as an emerging foodborne pathogen. The aim of this study was to isolate and characterize virulent strains of Cronobacter sakazakii from food samples of Bangladesh. RESULT: Six (6 Cronobacter sakazakii was isolated and identified from 54 food samples on the basis of biochemical characteristics, sugar fermentation, SDS-PAGE of whole cell protein, plasmid profile and PCR of Cronobacter spp. specific genes (esak, gluA, zpx, ompA, ERIC, BOX-AIR and sequencing. These strains were found to have moderately high antibiotic resistance against common antibiotics and some are ESBL producer. Most of the C. sakazakii isolates were capable of producing biofilm (strong biofilm producer, extracellular protease and siderophores, curli expression, haemolysin, haemagglutinin, mannose resistant haemagglutinin, had high cell surface hydrophobicity, significant resistance to human serum, can tolerate high concentration of salt, bile and DNase production. Most of them produced enterotoxins of different molecular weight. The isolates pose significant serological cross-reactivity with other gram negative pathogens such as serotypes of Salmonella spp., Shigella boydii, Shigella sonnei, Shigella flexneri and Vibrio cholerae. They had significant tolerance to high temperature, low pH, dryness and osmotic stress. CONCLUSION: Special attention should be given in ensuring hygiene in production and post-processing to prevent contamination of food with such stress-tolerant virulent Cronobacter sakazakii.

Molecular Epidemiology of Methicillin-Resistant and Methicillin-Susceptible Staphylococcus aureus in the Ovine Dairy Chain and in Farm-Related Humans

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Guerrino Macori

2017-05-01

Full Text Available Staphylococcus aureus is a major cause of clinical infections in humans and its enterotoxins cause foodborne disease. In the present study, we tested a total of 51 isolates of S. aureus from small-ruminant dairy farms with artisan dairy facilities, all located in Latium, Italy. The farms have a known history of a high prevalence of methicillin-resistant S. aureus (MRSA. Most of the MRSA isolates (27 of 51 belonged to spa-type t127 (43.1%, followed by t2678 (3.9%, t044 (2%, t1166 (2%, and t1773 (2%. PFGE performed on mecA positive strains identified one cluster (≥ 80% of similarity, comprising 22 MRSA. Nine of twenty-two MRSA isolates were assigned human host origin, and 13 isolates did not belong to a specific host. During the characterization study, one strain isolated from bulk tank milk samples harbored the pvl gene; the strain was not enterotoxigenic with a non-specific host according to the biotyping scheme, highlighting the possible emerging risk of transmission of bacterial virulence factors by foods, the environment, and foodhandlers. These findings stress the importance of hygienic measures at all processing steps of the food production chain and underline that monitoring for the presence of MRSA throughout the food chain is essential for public health.

The Dysregulation of Polyamine Metabolism in Colorectal Cancer Is Associated with Overexpression of c-Myc and C/EBPβ rather than Enterotoxigenic Bacteroides fragilis Infection

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Anastasiya V. Snezhkina

2016-01-01

Full Text Available Colorectal cancer is one of the most common cancers in the world. It is well known that the chronic inflammation can promote the progression of colorectal cancer (CRC. Recently, a number of studies revealed a potential association between colorectal inflammation, cancer progression, and infection caused by enterotoxigenic Bacteroides fragilis (ETBF. Bacterial enterotoxin activates spermine oxidase (SMO, which produces spermidine and H2O2 as byproducts of polyamine catabolism, which, in turn, enhances inflammation and tissue injury. Using qPCR analysis, we estimated the expression of SMOX gene and ETBF colonization in CRC patients. We found no statistically significant associations between them. Then we selected genes involved in polyamine metabolism, metabolic reprogramming, and inflammation regulation and estimated their expression in CRC. We observed overexpression of SMOX, ODC1, SRM, SMS, MTAP, c-Myc, C/EBPβ (CREBP, and other genes. We found that two mediators of metabolic reprogramming, inflammation, and cell proliferation c-Myc and C/EBPβ may serve as regulators of polyamine metabolism genes (SMOX, AZIN1, MTAP, SRM, ODC1, AMD1, and AGMAT as they are overexpressed in tumors, have binding site according to ENCODE ChIP-Seq data, and demonstrate strong coexpression with their targets. Thus, increased polyamine metabolism in CRC could be driven by c-Myc and C/EBPβ rather than ETBF infection.

Molecular characterization of Staphylococcus aureus isolates from skin and soft tissue infections samples and healthy carriers in the Central Slovenia region.

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Svent-Kucina, Natasa; Pirs, Mateja; Kofol, Romina; Blagus, Rok; Smrke, Dragica Maja; Bilban, Marjan; Seme, Katja

2016-04-01

Staphylococcus aureus is among the most important human pathogens. It is associated with different infections and is a major cause of skin and soft tissue infections (SSTIs). The aim of our study was to compare S. aureus isolates associated with SSTIs with isolates obtained from healthy carriers in the Central Slovenia region in terms of antimicrobial susceptibility, genetic diversity by clonal complex (CC)/sequence type, spa type, and by toxin gene profiling. In total, 274 S. aureus isolates were collected prospectively by culturing wound samples from 461 SSTI patients and nasal samples from 451 healthy carriers. We have demonstrated high heterogeneity in terms of CCs and spa type in both groups of isolates. The main clone among SSTI strains was Panton-Valentine leukocidin gene (pvl) positive CC121, whereas the main clone among carrier strains was CC45 carrying a large range of toxin genes. The main spa type in both groups was t091. Pvl was more frequently present in SSTI strains (31.2% SSTI vs 3.6% carrier strains) and staphylococcal enterotoxin C was more frequently present in carrier strains (1.6% SSTI vs 17.0% carrier strains). We have also demonstrated that methicillin-resistant S. aureus was a rare cause (2.8%) of SSTIs in our region. © 2016 APMIS. Published by John Wiley & Sons Ltd.

Ciprofloxacin and Trimethoprim Cause Phage Induction and Virulence Modulation in Staphylococcus aureus

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Goerke, Christiane; Köller, Johanna; Wolz, Christiane

2006-01-01

In Staphylococcus aureus strains of human origin, phages which integrate into the chromosomal gene coding for β-hemolysin (hlb) are widely distributed. Most of them encode accessory virulence determinants such as staphylokinase (sak) or enterotoxins. Here, we analyzed the effects of ciprofloxacin and trimethoprim on phage induction and expression of phage-encoded virulence factors by using isolates from patients with cystic fibrosis for which the induction of hlb-converting phages was demonstrated in vivo (C. Goerke, S. Matias y Papenberg, S. Dasbach, K. Dietz, R. Ziebach, B. C. Kahl, and C. Wolz, J. Infect. Dis. 189:724-734, 2004) as well as a φ13 lysogen of phage-cured strain 8325-4. Treatment of lysogens with subinhibitory concentrations of either antibiotic resulted in (i) delysogenization of strains resembling the isolates picked up after chronic lung infection and (ii) replication of phages in the bacterial host in a dose-dependent manner. Ciprofloxacin treatment resulted in enhanced recA transcription, indicating involvement of the SOS response in phage mobilization. Induction of φ13 was linked to elevated expression of the phage-encoded virulence gene sak, chiefly due to the activation of latent phage promoters. In summary, we could show the induction of hlb-converting phages and a subsequent virulence modulation of the host bacterium by ciprofloxacin and trimethoprim. PMID:16377683

Detection of virulence genes and the phylogenetic groups of Escherichia coli isolated from dogs in Brazil

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Fernanda Morcatti Coura

2018-02-01

Full Text Available ABSTRACT: This study identified the virulence genes, pathovars, and phylogenetic groups of Escherichia coli strains obtained from the feces of dogs with and without diarrhea. Virulence genes and phylogenetic group identification were studied using polymerase chain reaction. Thirty-seven E. coli isolates were positive for at least one virulence factor gene. Twenty-one (57.8% of the positive isolates were isolated from diarrheal feces and sixteen (43.2% were from the feces of non-diarrheic dogs. Enteropathogenic E. coli (EPEC were the most frequently (62.2% detected pathovar in dog feces and were mainly from phylogroup B1 and E. Necrotoxigenic E. coli were detected in 16.2% of the virulence-positive isolates and these contained the cytotoxic necrotizing factor 1 (cnf1 gene and were classified into phylogroups B2 and D. All E. coli strains were negative for the presence of enterotoxigenic E. coli (ETEC enterotoxin genes, but four strains were positive for ETEC-related fimbriae 987P and F18. Two isolates were Shiga toxin-producing E. coli strains and contained the toxin genesStx2 or Stx2e, both from phylogroup B1. Our data showed that EPEC was the most frequent pathovar and B1 and E were the most common phylogroups detected in E. coli isolated from the feces of diarrheic and non-diarrheic dogs.

CfaE tip mutations in enterotoxigenic Escherichia coli CFA/I fimbriae define critical human intestinal binding sites.

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Baker, K K; Levine, M M; Morison, J; Phillips, A; Barry, E M

2009-05-01

Enterotoxigenic Escherichia coli (ETEC) use colonization factors to attach to the human intestinal mucosa, followed by enterotoxin expression that induces net secretion and diarrhoeal illness. ETEC strain H10407 expresses CFA/I fimbriae, which are composed of multiple CfaB structural subunits and a CfaE tip subunit. Currently, the contribution of these individual fimbrial subunits in intestinal binding remains incompletely defined. To identify the role of CfaE in attachment in the native ETEC background, an R181A single-amino-acid substitution was introduced by recombination into the H10407 genome. The substitution of R181A eliminated haemagglutination and binding of intestinal mucosa biopsies in in vitro organ culture assays, without loss of CFA/I fimbriae expression. Wild-type in trans plasmid-expressed cfaE restored the binding phenotype. In contrast, in trans expression of cfaE containing amino acid 181 substitutions with similar amino acids, lysine, methionine and glutamine did not restore the binding phenotype, indicating that the loss of the binding phenotype was due to localized areas of epitope disruption. R181 appears to have an irreplaceable role in the formation of a receptor-binding feature on CFA/I fimbriae. The results specifically indicate that the CfaE tip protein is a required binding factor in CFA/I-mediated ETEC colonization, making it a potentially important vaccine antigen. © 2009 Blackwell Publishing Ltd.

Immune responses of a chimaeric protein vaccine containing Mycoplasma hyopneumoniae antigens and LTB against experimental M. hyopneumoniae infection in pigs.

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Marchioro, Silvana B; Sácristan, Rubén Del Pozo; Michiels, Annelies; Haesebrouck, Freddy; Conceição, Fabricio R; Dellagostin, Odir A; Maes, Dominiek

2014-08-06

A recombinant chimaeric protein containing three Mycoplasma hyopneumoniae antigens (C-terminal portion of P97, heat shock protein P42, and NrdF) fused to an adjuvant, the B subunit of heat-labile enterotoxin of Escherichia coli (LTB), was used to immunize pigs against enzootic pneumonia. The systemic and local immune responses, as well as the efficacy of the chimaeric protein in inducing protection against experimental M. hyopneumoniae infection were evaluated. In total, 60 male piglets, purchased from a M. hyopneumoniae-free herd, at 4 weeks of age were randomly allocated to six different experimental groups of 10 animals each: recombinant chimaeric protein by intramuscular (IM) (1) or intranasal (IN) (2) administration, commercial bacterin by IM administration (3), and the adjuvant LTB by IM (4, control group A) or IN (5, control group B) administration. All groups were immunized at 24 and 38 days of age and challenged at 52 days of age. The sixth group that was not challenged was used as the negative control (IN [n=5] or IM [n=5] administration of the LTB adjuvant). Compared with the non-challenged group, administration of the chimaeric protein induced significant (Phyopneumoniae infection in pigs. This lack of effectiveness points towards the need for further studies to improve the efficacy of this subunit-based vaccine approach. Copyright © 2014 Elsevier Ltd. All rights reserved.

Therapeutic Inhibition of Pro-Inflammatory Signaling and Toxicity to Staphylococcal Enterotoxin B by a Synthetic Dimeric BB-Loop Mimetic of MyD88

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2012-07-27

56 stock solution and 75 ml of the 16 dilution buffer was mixed with 25 ml of the supernatant, incubated for 30 min at 65 oC to inactivate endogenous ...shock by staphylococcal pyrogenic exotoxin type C. Infect Immun 36: 123–128. Meier survival analysis and log-rank tests were performed to compare

Detection of cell surface hydrophobicity, biofilm and fimbirae genes in salmonella isolated from tunisian clinical and poultry meat.

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Ben Abdallah, Fethi; Lagha, Rihab; Said, Khaled; Kallel, Héla; Gharbi, Jawhar

2014-04-01

The aim of this study was to evaluate the ability of 15 serotypes of Salmonella to form biofilm on polystyrene, polyvinyl chloride (PVC) and glass surfaces. . Initially slime production was assessed on CRA agar and hydrophobicity of 20 Salmonella strains isolated from poultry and human and two Salmonella enterica serovar Typhimurium references strains was achieved by microbial adhesion to n-hexadecane. In addition, biofilm formation on polystyrene, PVC and glass surfaces was also investigated by using MTT and XTT colorimetric assay. Further, distribution of Salmonella enterotoxin (stn), Salmonella Enteritidis fimbrial (sef) and plasmid encoded fimbrial (pef) genes among tested strains was achieved by PCR. Salmonella strains developed red and white colonies on CRA and they are considered as hydrophilic with affinity values to n-hexadecane ranged between 0.29% and 29.55%. Quantitative biofilm assays showed that bacteria are able to form biofilm on polystyrene with different degrees and 54.54% of strains produce a strong biofilm on glass. In addition, all the strains form only a moderate (54.54%) and weak (40.91%) biofilm on PVC. PCR detection showed that only S. Enteritidis harbour Sef gene, whereas Pef and stn genes were detected in S. Kentucky, S. Amsterdam, S. Hadar, S. Enteritidis and S. Typhimurium. Salmonella serotypes are able to form biofilm on hydrophobic and hydrophilic industrial surfaces. Biofilm formation of Salmonella on these surfaces has an increased potential to compromise food safety and potentiate public health risk.

Effect of age and latent CMV infection on CD8+ CD56+ T cells (NKT-like) frequency and functionality.

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Hassouneh, Fakhri; Campos, Carmen; López-Sejas, Nelson; Alonso, Corona; Tarazona, Raquel; Solana, Rafael; Pera, Alejandra

2016-09-01

Changes in the T cell pool caused by CMV infection have been proposed to contribute to immunosenescence, but it has been postulated that CMV can also have some beneficial effects in young individuals improving the immune response to other pathogens. T cells expressing CD56 (NKT-like cells) are cytotoxic effector cells with a significant role in the immune response against cancer. We have studied how age and latent CMV infection affect the frequency of NKT-like cells (CD8+ CD56+ T cells) and their response to Staphylococcal Enterotoxin B (SEB) in the context of CMV and ageing. NKT-like cell percentage increases with the combination of both CMV and age. The response to SEB and the polyfunctional index of NKT-like cells also increase with age in CMV-seropositive individuals. In young individuals, CMV infection induces a shift on the polyfunctional profile of CD8+ CD56- T cells not observed on the NKT-like cells response. NKT-like cells expressing CD57 are expanded in CMV-seropositive individuals and are more polyfunctional than their CD57-  counterpart. In addition CD57- NKT-like cells are more polyfunctional than CD8+ CD56- CD57- T cells. The results support that the expansion of polyfunctional NKT-cells may have a beneficial effect on the immune response against pathogens. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Reduced Memory CD4+ T-Cell Generation in the Circulation of Young Children May Contribute to the Otitis-Prone Condition

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Sharma, Sharad K.; Casey, Janet R.

2011-01-01

Background. An explanation for the immunologic dysfunction that causes children to be prone to repeated episodes of acute otitis media (AOM) has long been sought. Poor antibody response has been associated with the otitis-prone condition; however, there is no precise mechanistic explanation for this condition. Methods. Non–otitis-prone and otitis-prone children with AOM or nasopharyngeal (NP) colonization caused by either Streptococcus pneumoniae or Haemophilus influenzae were compared for pathogen-specific CD4+ T-helper memory responses by stimulating peripheral blood mononuclear cells using 6 vaccine candidate S. pneumoniae and 3 H. influenzae protein antigens. Samples were analyzed by multi-parameter flow cytometry. Results. Significantly reduced percentages of functional CD45RALow memory CD4+ T cells producing specific cytokines (interferon γ, interleukin [IL]–2, IL-4 and IL-17a) were observed in otitis-prone children following AOM and NP colonization with either S. pneumoniae or H. influenzae. Immunoglobulin (Ig) G responses to the studied protein antigens were reduced, which suggests that antigen-specific B-cell function may be compromised as a result of poor T-cell help. Staphylococcal enterotoxin B stimulated similar cytokine patterns in memory CD4+T cells in both groups of children. Conclusions. Otitis-prone children have suboptimal circulating functional T-helper memory and reduced IgG responses to S. pneumoniae or H. influenzae after colonization and after AOM; this immune dysfunction causes susceptibility to recurrent AOM infections. PMID:21791667

Detection of Genes for Superantigen Toxins in Methicillin-Resistant Staphylococcus aureus Clinical Isolates in Karachi

International Nuclear Information System (INIS)

Taj, Y.; Fatima, I.; Ali, S. W.; Kazmi, S. U.

2014-01-01

Objective: To detect genes for enterotoxins, exfoliative and toxic shock syndrome toxins in Staphylococcus aureus (S. aureus) strains isolated from clinical specimens. Study Design: Cross-sectional observational study. Place and Duration of Study: Department of Molecular Genetics, Dr. Ziauddin Hospital, Karachi, from January to December 2010. Methodology: Two hundred and ninety eight S. aureus clinical isolates were obtained from various clinical samples received at Dr. Ziauddin Hospital, Karachi. Out of these, 115 were detected as methicillin resistant (MRSA) by cefoxitin disk diffusion test showing a prevalence rate of 38.6%. Detection of individual toxin genes was performed by Polymerase Chain Reaction (PCR) by using only one primer pair for each tube. Uniplex primers were preferred as multiplex primers are longer in base pairs and have the potential for cross reaction due to non-specific binding and increase in optimization time. Results: The possession of a single gene or more than a single gene in MRSA isolates was found in 61.73% of clinical samples; the highest number was found in pus swab, followed by sputum, blood, urethral swab, and urine. The prevalence of toxin genes was higher in MRSA as compared to methicillin sensitive (MSSA) isolates (19.12%). Conclusion: PCR detects strains possessing toxin genes independent of their expression. The possession of genes for super-antigens seems to be a frequent and habitual trait of S. aureus more so in MRSA. (author)

The Proteome of Biologically Active Membrane Vesicles from Piscirickettsia salmonis LF-89 Type Strain Identifies Plasmid-Encoded Putative Toxins

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Cristian Oliver

2017-09-01

Full Text Available Piscirickettsia salmonis is the predominant bacterial pathogen affecting the Chilean salmonid industry. This bacterium is the etiological agent of piscirickettsiosis, a significant fish disease. Membrane vesicles (MVs released by P. salmonis deliver several virulence factors to host cells. To improve on existing knowledge for the pathogenicity-associated functions of P. salmonis MVs, we studied the proteome of purified MVs from the P. salmonis LF-89 type strain using multidimensional protein identification technology. Initially, the cytotoxicity of different MV concentration purified from P. salmonis LF-89 was confirmed in an in vivo adult zebrafish infection model. The cumulative mortality of zebrafish injected with MVs showed a dose-dependent pattern. Analyses identified 452 proteins of different subcellular origins; most of them were associated with the cytoplasmic compartment and were mainly related to key functions for pathogen survival. Interestingly, previously unidentified putative virulence-related proteins were identified in P. salmonis MVs, such as outer membrane porin F and hemolysin. Additionally, five amino acid sequences corresponding to the Bordetella pertussis toxin subunit 1 and two amino acid sequences corresponding to the heat-labile enterotoxin alpha chain of Escherichia coli were located in the P. salmonis MV proteome. Curiously, these putative toxins were located in a plasmid region of P. salmonis LF-89. Based on the identified proteins, we propose that the protein composition of P. salmonis LF-89 MVs could reflect total protein characteristics of this P. salmonis type strain.

Molecular Typing of Staphylococcus aureus Isolated from Patients with Autosomal Dominant Hyper IgE Syndrome

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Inka Sastalla

2017-06-01

Full Text Available Autosomal dominant hyper IgE syndrome (AD-HIES is a primary immunodeficiency caused by a loss-of-function mutation in the Signal Transducer and Activator of Transcription 3 (STAT3. This immune disorder is clinically characterized by increased susceptibility to cutaneous and sinopulmonary infections, in particular with Candida and Staphylococcus aureus. It has recently been recognized that the skin microbiome of patients with AD-HIES is altered with an overrepresentation of certain Gram-negative bacteria and Gram-positive staphylococci. However, these alterations have not been characterized at the species- and strain-level. Since S. aureus infections are influenced by strain-specific expression of virulence factors, information on colonizing strain characteristics may provide insights into host-pathogen interactions and help guide management strategies for treatment and prophylaxis. The aim of this study was to determine whether the immunodeficiency of AD-HIES selects for unique strains of colonizing S. aureus. Using multi-locus sequence typing (MLST, protein A (spa typing, and PCR-based detection of toxin genes, we performed a detailed analysis of the S. aureus isolates (n = 13 found on the skin of twenty-one patients with AD-HIES. We found a low diversity of sequence types, and an abundance of strains that expressed methicillin resistance, Panton-Valentine leukocidin (PVL, and staphylococcal enterotoxins K and Q (SEK, SEQ. Our results indicate that patients with AD-HIES may often carry antibiotic-resistant strains that harbor key virulence factors.

Principles for new optical techniques in medical diagnostics for mHealth applications

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Balsam, Joshua Michael

Medical diagnostics is a critical element of effective medical treatment. However, many modern and emerging diagnostic technologies are not affordable or compatible with the needs and conditions found in low-income and middle-income countries and regions. Resource-poor areas require low-cost, robust, easy-to-use, and portable diagnostics devices compatible with telemedicine (i.e. mHealth) that can be adapted to meet diverse medical needs. Many suitable devices will need to be based on optical technologies, which are used for many types of biological analyses. This dissertation describes the fabrication and detection principles for several low-cost optical technologies for mHealth applications including: (1) a webcam based multi-wavelength fluorescence plate reader, (2) a lens-free optical detector used for the detection of Botulinum A neurotoxin activity, (3) a low cost micro-array reader that allows the performance of typical fluorescence based assays demonstrated for the detection of the toxin staphylococcal enterotoxin (SEB), and (4) a wide-field flow cytometer for high throughput detection of fluorescently labeled rare cells. This dissertation discusses how these technologies can be harnessed using readily available consumer electronics components such as webcams, cell phones, CCD cameras, LEDs, and laser diodes. There are challenges in developing devices with sufficient sensitivity and specificity, and approaches are presented to overcoming these challenges to create optical detectors that can serve as low cost medical diagnostics in resource-poor settings for mHealth.

Expression of virulence factors by Staphylococcus aureus grown in serum.

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Oogai, Yuichi; Matsuo, Miki; Hashimoto, Masahito; Kato, Fuminori; Sugai, Motoyuki; Komatsuzawa, Hitoshi

2011-11-01

Staphylococcus aureus produces many virulence factors, including toxins, immune-modulatory factors, and exoenzymes. Previous studies involving the analysis of virulence expression were mainly performed by in vitro experiments using bacterial medium. However, when S. aureus infects a host, the bacterial growth conditions are quite different from those in a medium, which may be related to the different expression of virulence factors in the host. In this study, we investigated the expression of virulence factors in S. aureus grown in calf serum. The expression of many virulence factors, including hemolysins, enterotoxins, proteases, and iron acquisition factors, was significantly increased compared with that in bacterial medium. In addition, the expression of RNA III, a global regulon for virulence expression, was significantly increased. This effect was partially restored by the addition of 300 μM FeCl₃ into serum, suggesting that iron depletion is associated with the increased expression of virulence factors in serum. In chemically defined medium without iron, a similar effect was observed. In a mutant with agr inactivated grown in serum, the expression of RNA III, psm, and sec4 was not increased, while other factors were still induced in the mutant, suggesting that another regulatory factor(s) is involved. In addition, we found that serum albumin is a major factor for the capture of free iron to prevent the supply of iron to bacteria grown in serum. These results indicate that S. aureus expresses virulence factors in adaptation to the host environment.

Evaluation of genetic and phenotypic consistency of Bacillus coagulans MTCC 5856: a commercial probiotic strain.

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Majeed, Muhammed; Nagabhushanam, Kalyanam; Natarajan, Sankaran; Sivakumar, Arumugam; Eshuis-de Ruiter, Talitha; Booij-Veurink, Janine; de Vries, Ynte P; Ali, Furqan

2016-04-01

Commercial probiotics preparation containing Bacillus coagulans have been sold in the market for several decades. Due to its high intra-species genomic diversity, it is very likely that B. coagulans strain may alter in different ways over multiple years of production. Therefore, the present study focuses to evaluate the genetic consistency and probiotic potential of B. coagulans MTCC 5856. Phenotypic and genotypic techniques including biochemical profiling, 16S rRNA sequencing, GTG 5″, BOX PCR fingerprinting, and Multi-Locus-Sequence typing (MLST) were carried out to evaluate the identity and consistency of the B. coagulans MTCC 5856. Further, in vitro probiotic potential, safety and stability at ambient temperature conditions of B. coagulans MTCC 5856 were evaluated. All the samples were identified as B. coagulans by biochemical profiling and 16S rRNA sequencing. GTG 5″, BOX PCR fingerprints and MLST studies revealed that the same strain was present over 3 years of commercial production. B. coagulans MTCC 5856 showed resistance to gastric acid, bile salt and exhibited antimicrobial activity in in-vitro studies. Additionally, B. coagulans MTCC 5856 was found to be non-mutagenic, non-cytotoxic, negative for enterotoxin genes and stable at ambient temperature (25 ± 2 °C) for 36 months. The data of the study verified that the same strain of B. coagulans MTCC 5856 was present in commercial preparation over multiple years of production.

Novel Clostridium difficile Anti-Toxin (TcdA and TcdB Humanized Monoclonal Antibodies Demonstrate In Vitro Neutralization across a Broad Spectrum of Clinical Strains and In Vivo Potency in a Hamster Spore Challenge Model.

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Hongyu Qiu

Full Text Available Clostridium difficile (C. difficile infection (CDI is the main cause of nosocomial antibiotic-associated colitis and increased incidence of community-associated diarrhea in industrialized countries. At present, the primary treatment of CDI is antibiotic administration, which is effective but often associated with recurrence, especially in the elderly. Pathogenic strains produce enterotoxin, toxin A (TcdA, and cytotoxin, toxin B (TcdB, which are necessary for C. difficile induced diarrhea and gut pathological changes. Administration of anti-toxin antibodies provides an alternative approach to treat CDI, and has shown promising results in preclinical and clinical studies. In the current study, several humanized anti-TcdA and anti-TcdB monoclonal antibodies were generated and their protective potency was characterized in a hamster infection model. The humanized anti-TcdA (CANmAbA4 and anti-TcdB (CANmAbB4 and CANmAbB1 antibodies showed broad spectrum in vitro neutralization of toxins from clinical strains and neutralization in a mouse toxin challenge model. Moreover, co-administration of humanized antibodies (CANmAbA4 and CANmAbB4 cocktail provided a high level of protection in a dose dependent manner (85% versus 57% survival at day 22 for 50 mg/kg and 20 mg/kg doses, respectively in a hamster gastrointestinal infection (GI model. This study describes the protective effects conferred by novel neutralizing anti-toxin monoclonal antibodies against C. difficile toxins and their potential as therapeutic agents in treating CDI.

Bacillus cereus and related species.

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Drobniewski, F A

1993-10-01

Bacillus cereus is a gram-positive aerobic or facultatively anaerobic spore-forming rod. It is a cause of food poisoning, which is frequently associated with the consumption of rice-based dishes. The organism produces an emetic or diarrheal syndrome induced by an emetic toxin and enterotoxin, respectively. Other toxins are produced during growth, including phospholipases, proteases, and hemolysins, one of which, cereolysin, is a thiol-activated hemolysin. These toxins may contribute to the pathogenicity of B. cereus in nongastrointestinal disease. B. cereus isolated from clinical material other than feces or vomitus was commonly dismissed as a contaminant, but increasingly it is being recognized as a species with pathogenic potential. It is now recognized as an infrequent cause of serious nongastrointestinal infection, particularly in drug addicts, the immunosuppressed, neonates, and postsurgical patients, especially when prosthetic implants such as ventricular shunts are inserted. Ocular infections are the commonest types of severe infection, including endophthalmitis, panophthalmitis, and keratitis, usually with the characteristic formation of corneal ring abscesses. Even with prompt surgical and antimicrobial agent treatment, enucleation of the eye and blindness are common sequelae. Septicemia, meningitis, endocarditis, osteomyelitis, and surgical and traumatic wound infections are other manifestations of severe disease. B. cereus produces beta-lactamases, unlike Bacillus anthracis, and so is resistant to beta-lactam antibiotics; it is usually susceptible to treatment with clindamycin, vancomycin, gentamicin, chloramphenicol, and erythromycin. Simultaneous therapy via multiple routes may be required.

Hybrids of Shigatoxigenic and Enterotoxigenic Escherichia coli (STEC/ETEC) Among Human and Animal Isolates in Finland.

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Nyholm, O; Heinikainen, S; Pelkonen, S; Hallanvuo, S; Haukka, K; Siitonen, A

2015-11-01

Diarrhoeagenic Escherichia coli (DEC) cause serious foodborne infections in humans. Total of 450 Shigatoxigenic E. coli (STEC) strains isolated from humans, animals and environment in Finland were examined by multiplex PCR targeting the virulence genes of various DEC pathogroups simultaneously. One per cent (3/291) of the human STEC and 14% (22/159) of the animal and environmental STEC had genes typically present in enterotoxigenic E. coli (ETEC). The strains possessed genes encoding both Shiga toxin 1 and/or 2 (stx1 and/or stx2 ) and ETEC-specific heat-stable (ST) enterotoxin Ia (estIa). The identified stx subtypes were stx1a, stx1c, stx2a, stx2d and stx2g. The three human STEC/ETEC strains were isolated from the patients with haemolytic uraemic syndrome and diarrhoea and from an asymptomatic carrier. The animal STEC/ETEC strains were isolated from cattle and moose. The human and animal STEC/ETEC strains belonged to 11 serotypes, of which O2:H27, O15:H16, O101:H-, O128:H8 and O141:H8 have previously been described to be associated with human disease. Identification of multiple virulence genes offers further information for assessing the virulence potential of STEC and other DEC. The emergence of novel hybrid pathogens should be taken into account in the patient care and epidemiological surveillance. © 2015 Blackwell Verlag GmbH.

Molecular characterization of Shigella spp. from patients in Gabon 2011-2013.

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Schaumburg, Frieder; Alabi, Abraham S; Kaba, Harry; Lell, Bertrand; Becker, Karsten; Grobusch, Martin P; Kremsner, Peter G; Mellmann, Alexander

2015-04-01

Shigella spp. dysentery is widespread in developing countries; the incidence is particularly high in children between 1-2 years of age. In sub-Saharan Africa, there is a paucity of epidemiological data on Shigella spp., with possible negative consequences for recognition and correct treatment choice for this life-threatening bacterial infection. We therefore characterized Shigella spp. isolates from Gabon. The antimicrobial resistance, virulence factors, genotypes and mobile genetic elements of Shigella isolates (29 S. flexneri; 5 S. boydii; 3 S. sonnei) from a retrospective strain collection were analyzed. High resistance rates were found for gentamicin and tetracycline (100%, 37/37), cotrimoxazole (92%, 34/37) and ampicillin (84%, 31/37). All isolate harbored ial and ipaH; no isolate produced Shiga toxins (stx1/2); enterotoxins (set1A/B) were only found in S. flexneri (n=19). Multilocus sequence types (MLST) clustered with global clones. A high prevalence of atypical class 1 integrons harboring blaOXA30 and aadA1 were detected in S. flexneri, while all S. sonnei carried class 2 integrons. There is a strong link of Gabonese Shigella spp. isolates with pandemic lineages as they cluster with major global clones and frequently carry atypical class 1 integrons which are frequently reported in Shigella spp. from Asia. © The Author 2014. Published by Oxford University Press on behalf of Royal Society of Tropical Medicine and Hygiene. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Detection of toxin genes and RAPD analysis of bacillus cereus isolates from different soil types

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Savic Dejana

2015-01-01

Full Text Available The aim of this study was to detect genes for enterotoxins (hbla, entFM and bceT and for emetic toxin (cer, to determine antibiotic resistance, and to estimate intraspecies diversity in B. cereus isolates by RAPD analysis. B. cereus was identified in 12 out of 117 indigenous Bacillus spp. using the classical microbiological methods and PCR. All isolates were resistant to penicillin and ampicillin, two to tetracyclin and four to trimethoprim-sulphamethoxazole. Also, all isolates produced inducible penicillinases and β-lactamase. Toxin genes were detected with PCR. EntFM and cer genes were present in all isolates, hbla in all, but two, and bceT in none. RAPD analysis was performed with four different primers, two of them designed for this study. The intraspecies diversity revealed 10 different patterns at the 90% similarity level. Two separate clusters were formed regardless of a soil type or utilization. The detection of genes encoding toxins in all B. cereus isolates indicated these bacteria as potentially pathogenic and seriously for human health. Regardless of a soil type or utilization, the RAPD analysis showed high intraspecies heterogeneity in B. cereus isolates. To the best of our knowledge, this is the first study to analyse the presence of entero- and emetic toxin genes and genetic heterogeneity in B. cereus isolates from different soil types and different soil utilization in Serbia. [Projekat Ministarstva nauke Republike Srbije, br. TR37006

Toxigenic profile of methicillin-sensitive and resistant Staphylococcus aureus isolated from special groups.

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de Souza, Camila Sena Martins; Fortaleza, Carlos Magno Castelo Branco; Witzel, Claudia Lima; Silveira, Mônica; Bonesso, Mariana Fávero; Marques, Silvio Alencar; Cunha, Maria de Lourdes Ribeiro de Souza da

2016-02-16

Staphylococcus aureus is characterized by its pathogenicity and high prevalence, causing disease in both healthy and immunocompromised individuals due to its easy dissemination. This fact is aggravated by the widespread dissemination of S. aureus carrying toxigenic genes. The objective of this study was to determine the toxigenic profile of methicillin-sensitive S. aureus (MSSA) and methicillin-resistant S. aureus (MRSA) in patients with purulent skin and/or soft tissue infections seen at the Dermatology Department of the University Hospital of the Botucatu Medical School, asymptomatic adults older than 60 years living in nursing homes, and prison inmates of the Avaré Detention Center. PCR was used for the detection of the mecA gene, enterotoxin genes (sea, seb, and sec), exfoliative toxins A and B (eta and etb), toxic shock syndrome toxin 1 (tst), panton-valentine leukocidin (lukS-PV and lukF-PV), and alpha- and delta-hemolysins or cytotoxins (hla and hld). The results showed a significant prevalence of toxigenic genes among S. aureus isolates from asymptomatic individuals, with the observation of a higher prevalence of cytotoxin genes. However, the panton-valentine leukocidin gene was only detected in MSSA isolated from patients with skin infections and the tst gene was exclusively found in MSSA isolated from prison inmates. The present study demonstrated a significant prevalence of toxigenic genes in MSSA and MRSA strains isolated from asymptomatic S. aureus carriers. There was a higher prevalence of cytotoxin genes.

The Proteome of Biologically Active Membrane Vesicles from Piscirickettsia salmonis LF-89 Type Strain Identifies Plasmid-Encoded Putative Toxins.

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Oliver, Cristian; Hernández, Mauricio A; Tandberg, Julia I; Valenzuela, Karla N; Lagos, Leidy X; Haro, Ronie E; Sánchez, Patricio; Ruiz, Pamela A; Sanhueza-Oyarzún, Constanza; Cortés, Marcos A; Villar, María T; Artigues, Antonio; Winther-Larsen, Hanne C; Avendaño-Herrera, Ruben; Yáñez, Alejandro J

2017-01-01

Piscirickettsia salmonis is the predominant bacterial pathogen affecting the Chilean salmonid industry. This bacterium is the etiological agent of piscirickettsiosis, a significant fish disease. Membrane vesicles (MVs) released by P. salmonis deliver several virulence factors to host cells. To improve on existing knowledge for the pathogenicity-associated functions of P. salmonis MVs, we studied the proteome of purified MVs from the P. salmonis LF-89 type strain using multidimensional protein identification technology. Initially, the cytotoxicity of different MV concentration purified from P. salmonis LF-89 was confirmed in an in vivo adult zebrafish infection model. The cumulative mortality of zebrafish injected with MVs showed a dose-dependent pattern. Analyses identified 452 proteins of different subcellular origins; most of them were associated with the cytoplasmic compartment and were mainly related to key functions for pathogen survival. Interestingly, previously unidentified putative virulence-related proteins were identified in P. salmonis MVs, such as outer membrane porin F and hemolysin. Additionally, five amino acid sequences corresponding to the Bordetella pertussis toxin subunit 1 and two amino acid sequences corresponding to the heat-labile enterotoxin alpha chain of Escherichia coli were located in the P. salmonis MV proteome. Curiously, these putative toxins were located in a plasmid region of P. salmonis LF-89. Based on the identified proteins, we propose that the protein composition of P. salmonis LF-89 MVs could reflect total protein characteristics of this P. salmonis type strain.

Cloning and Expression of Fusion Genes of Domain A-1 Protective Antigen of Bacillus Anthracis and Shigella Enterotoxin B Subunit (Stxb In E. Coil

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AH ahmadi

2015-02-01

Conclusion: The findings of the current study revealed that this antigen can be raised as an anti-cancer and recombinant vaccine candidate against types of Shigella, Escherichia coli and Bacillus anthracis which can be due to such factors as identification of antigen(PA by antibody PA20, its apoptosis induction properties, property of immunogenicity, adjuvant and delivery of STxB protein and high expression levels of Gb3 in human cancer cells.

Structure-Based Design and Synthesis of a Small Molecule that Exhibits Anti-inflammatory Activity by Inhibition of MyD88-mediated Signaling to Bacterial Toxin Exposure.

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Alam, Shahabuddin; Javor, Sacha; Degardin, Melissa; Ajami, Dariush; Rebek, Mitra; Kissner, Teri L; Waag, David M; Rebek, Julius; Saikh, Kamal U

2015-08-01

Both Gram-positive and Gram-negative pathogens or pathogen-derived components, such as staphylococcal enterotoxins (SEs) and endotoxin (LPS) exposure, activate MyD88-mediated pro-inflammatory cellular immunity for host defense. However, dysregulated MyD88-mediated signaling triggers exaggerated immune response that often leads to toxic shock and death. Previously, we reported a small molecule compound 1 mimicking BB-loop structure of MyD88 was capable of inhibiting pro-inflammatory response to SEB exposure in mice. In this study, we designed a dimeric structure compound 4210 covalently linked with compound 1 by a non-polar cyclohexane linker which strongly inhibited the production of pro-inflammatory cytokines in human primary cells to SEB (IC50 1-50 μm) or LPS extracted from Francisella tularensis, Escherichia coli, or Burkholderia mallei (IC50 10-200 μm). Consistent with cytokine inhibition, in a ligand-induced cell-based reporter assay, compound 4210 inhibited Burkholderia mallei or LPS-induced MyD88-mediated NF-kB-dependent expression of reporter activity (IC50 10-30 μm). Furthermore, results from a newly expressed MyD88 revealed that 4210 inhibited MyD88 dimer formation which is critical for pro-inflammatory signaling. Importantly, a single administration of compound 4210 in mice showed complete protection from lethal toxin challenge. Collectively, these results demonstrated that compound 4210 inhibits toxin-induced inflated pro-inflammatory immune signaling, thus displays a potential bacterial toxin therapeutic. © 2014 John Wiley & Sons A/S.