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Sample records for isolation molecular identification

  1. Molecular identification of clinical Nocardia isolates from India.

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    Rudramurthy, Shivaprakash M; Honnavar, Prasanna; Kaur, Harsimran; Samanta, Palash; Ray, Pallab; Ghosh, Anup; Chakrabarti, Arunaloke

    2015-10-01

    The epidemiology of nocardiosis is evolving with increasing number of Nocardia spp. causing human infection. In recent years, molecular techniques have been used to identify Nocardia spp. There are limited data available on the spectrum of Nocardia spp. isolated from clinical samples in India. Here, a molecular study was carried on 30 clinical isolates maintained in our National Culture Collection to evaluate the techniques used for identifying the agents. The isolates were identified by sequencing two promising genes: the 16S rRNA gene and hsp65. Both hsp65 and the 16S rRNA gene could reliably identify 90 % of Nocardia isolates, i.e. N. farcinica, N. cyriacigeorgica, N. brasiliensis, N. otitidiscaviarum, N. amamiensis and N. pneumoniae. The mean percentage dissimilarity of sequence identification was higher using the hsp65 gene (4 %, range 0-7.9 %) compared with the 16S rRNA gene (2.3 %, range 0-8.9 %). Two isolates that showed ambiguous results in both the short segment of the 16S rRNA gene and hsp65 sequences could be resolved by sequencing a larger fragment (∼1000 bp) of the 16S rRNA gene. Both of these isolates were identified as N. beijingensis with similarities of 99.8 and 100 % compared with the standard strain. Genotyping of N. cyriacigeorgica strains was performed using hsp65 gene sequences and compared with previously described genotypes. Our N. cyriacigeorgica isolates belonged to genotype 1 (n = 4) and genotype 2 (n = 2). The present study highlights a wide spectrum of Nocardia spp. in India and emphasizes the need for molecular techniques for identification to the species level.

  2. Isolation, identification and molecular characterisation of an isolate ...

    African Journals Online (AJOL)

    Nucleotide sequence analysis of the partial coat protein gene of the ZYMV isolate from KZN revealed 95−98% sequence identity with isolates occurring in central Europe and the Indian subcontinent, and 90−93% identity with isolates from Singapore and Taiwan. These high levels of sequence identity indicate that the KZN ...

  3. Pythium insidiosum: morphological and molecular identification of Brazilian isolates

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    Maria Isabel de Azevedo

    2012-07-01

    Full Text Available Pythium insidiosum is an oomycete belonging to the kingdom Stramenipila and it is the etiologic agent of pythiosis. Pythiosis is a life-threatening infectious disease characterized by the development of chronic lesions on cutaneous and subcutaneous, intestinal, and bone tissues in humans and many species of animals. The identification of P. insidiosum is important in order to implement a rapid and definitive diagnosis and an effective treatment. This study reports the identification of 54 isolates of P. insidiosum of horses, dogs and sheep that presented suspicious clinical lesions of pythiosis from different regions in Brazil, by using morphological and molecular assays. Throughout the PCR it was possible to confirm the identity of all Brazilian isolates as being P. insidiosum.

  4. The isolated Leptospira Spp. Identification by molecular biological techniques

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    Duangjai Suwancharoen

    2017-01-01

    Full Text Available Leptospirosis is a zoonotic disease caused by the bacteria of Leptospira spp. Identification of this bacterium relies on serotyping and genotyping. Data base for animal causative serovars in Thailand is limited. As the unknown serovars are found in the laboratory, they need to be sent overseas for referent identification. To reduce the cost, this research intended to develop a leptospiral identification method which is user–friendly and able to classify efficiently. Ten Leptospira isolations were cultured from urine samples. They were identified by three molecular biological techniques, including Pulsed-Field Gel Electrophoresis (PFGE, Variable Number Tandem Repeat (VNTR and Multilocus Sequence Typing (MLST. These methods were developed and compared to find the most suitable one for leptospiral identification. VNTR was found to be inappropriate since it could not identify the agents and it did not show the PCR product. PFGE and MLST gave the same results of the unknown 1 and 2 which were L.weilii sv Samin st Samin. Unknown 4 showed different results by each technique. Unknown 5 to 10 were likely to be L.meyeri sv Ranarum st ICF and Leptonema illini sv Illini st 3055 by PFGE but MLST could not identify the serovar. However, molecular biological technique for Leptospira identification should be done by several methods in order to confirm the result of each other.

  5. Molecular identification of Sporothrix clinical isolates in China*

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    Liu, Ting-ting; Zhang, Ke; Zhou, Xun

    2014-01-01

    In this study, we investigated the molecular phylogeny of 64 clinical isolates which were identified as Sporothrix schenckii sensu lato by morphological identification. All of the strains were isolates from patients from several provinces in China. The phylogeny was inferred by DNA sequence analyses based on datasets of the ribosomal internal transcribed spacer (ITS) and a combined ITS and partial β-tubulin region. Reference sequences were retrieved from GenBank. Results showed that all of the isolates were clustered in a distinct clade with a type of Sporothrix globosa. Our analysis showed that S. globosa is the causal agent of the tested sporotrichosis in China, rather than S. schenckii that was generally believed to be the case. The existence of S. schenckii in China remains to be confirmed. This study improved our understanding of the distribution of the species in S. schenckii complex. PMID:24390750

  6. Molecular identification of Sporothrix clinical isolates in China.

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    Liu, Ting-ting; Zhang, Ke; Zhou, Xun

    2014-01-01

    In this study, we investigated the molecular phylogeny of 64 clinical isolates which were identified as Sporothrix schenckii sensu lato by morphological identification. All of the strains were isolates from patients from several provinces in China. The phylogeny was inferred by DNA sequence analyses based on datasets of the ribosomal internal transcribed spacer (ITS) and a combined ITS and partial β-tubulin region. Reference sequences were retrieved from GenBank. Results showed that all of the isolates were clustered in a distinct clade with a type of Sporothrix globosa. Our analysis showed that S. globosa is the causal agent of the tested sporotrichosis in China, rather than S. schenckii that was generally believed to be the case. The existence of S. schenckii in China remains to be confirmed. This study improved our understanding of the distribution of the species in S. schenckii complex.

  7. Isolation and molecular identification of yeast strains from “Rabilé” a ...

    African Journals Online (AJOL)

    Isolation and molecular identification of yeast strains from “Rabilé” a starter of local fermented drink. Ibrahim Keita, Marius K Somda, Aly Savadogo, Iliassou Mogmenga, Ousmane Koita, Alfred S Traore ...

  8. Molecular Identification of Giardia duodenalis Isolates from Fars Province, Iran.

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    Mohammad Rayani

    2014-03-01

    Full Text Available Giardia duodenalis is one of the most common human intestinal protozoan parasites worldwide and is endemic throughout the world with a vast range of mammalian hosts. The present study aimed to identify the prevalence of G. duodenalis isolates and determine the most common of its assemblages in the patients referring to health centers and hospitals in Fars province, Iran that will be subjected to further molecular investigation.We collected 1000 human fecal samples from health centers and hospitals in Shiraz, Iran in a one year period from September 2009 to August 2010. Microscopic examination for the presence of G. duodenalis cysts and trophozoites was performed by direct wet mount before and after the concentration techniques. Extraction of DNA was performed by Phenol-Chloroform-Isoamylalcohol (PCI. G. duodenalis-positive specimens were analyzed by PCR. A fragment of the SSU-rDNA (292 bp gene was amplified by PCR using the forward primer RH11 and the reverse primer RH4. Genotyping was performed using sequence analysis of G. duodenalis glutamate dehydrogenase gene using primers GDHeF, GDHiF, and GDHiR.The prevalence of Giardia infection was 10.7% (107/1000 examined based on microscopic examination. PCR identified 80% (40/50 of the samples as positive for G. duodenalis based on SSU-rDNA amplification on sucrose gradient samples. Besides, genotyping results indicated 32 isolates (80% as assemblage AII and 8 isolates (20% as assemblage BIII and BIV based on the DNA sequence analysis of the glutamate dehydrogenase locus of G. duodenalis.The findings of this study emphasize that Iran (Fars Province is a favorable area for giardiasis with an anthroponotic infection route.

  9. Phenotypic and molecular identification of Fonsecaea pedrosoi strains isolated from chromoblastomycosis patients in Mexico and Venezuela.

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    Carolina Rojas, O; León-Cachón, Rafael B R; Pérez-Maya, Antonio Alí; Aguirre-Garza, Marcelino; Moreno-Treviño, María G; González, Gloria M

    2015-05-01

    Chromoblastomycosis is a chronic granulomatous disease caused frequently by fungi of the Fonsecaea genus. The objective of this study was the phenotypic and molecular identification of F. pedrosoi strains isolated from chromoblastomycosis patients in Mexico and Venezuela. Ten strains were included in this study. For phenotypic identification, we used macroscopic and microscopic morphologies, carbohydrate assimilation test, urea hydrolysis, cixcloheximide tolerance, proteolitic activity and the thermotolerance test. The antifungal activity of five drugs was evaluated against the isolates. Molecular identification was performed by sequencing the internal transcribed spacer (ITS) ribosomal DNA regions of the isolated strains. The physiological analysis and morphological features were variable and the precise identification was not possible. All isolates were susceptible to itraconazole, terbinafine, voriconazole and posaconazole. Amphotericin B was the least effective drug. The alignment of the 559-nucleotide ITS sequences from our strains compared with sequences of GenBank revealed high homology with F. pedrosoi (EU285266.1). In this study, all patients were from rural areas, six from Mexico and four from Venezuela. Ten isolates were identified by phenotypic and molecular analysis, using ITS sequence and demonstrated that nine isolates from Mexico and Venezuela were 100% homologous and one isolate showed a small genetic distance. © 2015 Blackwell Verlag GmbH.

  10. Isolation and molecular identification of Fusobacterium nucleatum from Nigerian patients with oro-facial infections.

    Science.gov (United States)

    Nwaokorie, F O; Coker, A O; Ogunsola, F T; Avika-Campos, M J; Gaetti-Jardim, E; Ayanbadejo, P O; Umeizudike, K A; Abdurrazaq, O T

    2011-01-01

    Fusobacterium nucleatum is one of the most common anaerobic bacteria present in the oral cavity and is often isolated from infections involving other body sites. To characterise F. nucleatum strains from patients attending a teaching hospital in Nigeria in order to provide information on the methods for accurate identification of anaerobes in clinical specimen. Fusobacterium nucleatum specie from 50 patients presenting with oro-facial infections were studied by culture on Fusobacterium selective agar and fastidious anaerobe agar. The isolates were characterised based on colonial morphology, microscopy, lipase production, susceptibility to kanamycin and colistin and resistance to vancomycin. Biochemical tests were performed using a commercial test kit. The identity of the isolates was confirmed based on molecular characterization performed using polymerase chain reaction (PCR) analysis. Forty-eight (96%) F. nucleatum isolates were obtained from the 50 patients by culture and all the isolates were identified by colonial appearance and microscopy based on their unique spindle shape with tapered ends. Only 26 (54.2%) of the 48 isolates were identified by commercial API 20A test kit while PCR confirmed the identity of all the isolates. Anaerobes are involved in human infections and their study is quite cumbersome due to tedious nature and high cost of the techniques involved. Cultural method is reliable in the isolation and identification of F. nucleatum species. PCR is a rapid and simple method that can complement the phenotypic identification of anaerobes and would assist in their full identification.

  11. Molecular Identification and Conventional Susceptibility Testing of Iranian Clinical Mycobacterium fortuitum Isolates

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    Parvin Heidarieh

    2010-01-01

    Full Text Available Rapidly growing mycobacteria (RGM are capable of producing diseases in humans. Since mycobacteria vary in their susceptibility, precise identification is critical for adoption of correct drug therapy. The main aim of this study was molecular identification and evaluation of antimicrobial susceptibility pattern of Iranian clinically isolated Myocbacterium fortuitum.Materials and MethodsA total of 72 presumptively identified isolates of clinical atypical mycobacteria collected by Isfahan Research Center for Infectious Diseases & Tropical Medicine during 2006-2008 were included in the current study. A combination of conventional and molecular tests was applied to identify the isolates. Molecular methods including genus and group specific PCR and PCR-Restriction Algorithm (PRA based on hsp65 gene were applied to achieve exact identification of mycobacterial strains. Antimicrobial susceptibility testing on M. fortuitum isolates was performed by in-house prepared broth microdilution test..ResultsOut of 72 collected atypical mycobacteria isolates, we identified 25 strains of M. fortuitum. All strains had the specific molecular markers of mycobacterial identity and similar species specific PRA pattern of the international type strain of M. fortuitum. Drug susceptibility testing showed that the M. fortuitum isolates are sensitive to amikacin, sulfamethoxazole and ciprofloxacin (100%, imipenem (92%, clarithromycin (76%, cefoxitin (56% and doxycycline (16%.ConclusionMolecular identification of atypical mycobacteria based on PRA is a reliable and rapid approach which can identify mycobacterial strains to the species level. Our study showed that M. fortuitum plays a significant role in pulmonary and extrapulmonary infection in patients and should be given proper considerations when clinical samples are processed.

  12. Molecular identification of Echinococcus granulosus isolates from ruminants in Greece.

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    Roinioti, Erifylli; Papathanassopoulou, Aegli; Theodoropoulou, Ioanna; Simsek, Sami; Theodoropoulos, Georgios

    2016-08-15

    Cystic echinococcosis is a parasitic disease caused by Echinococcus granulosus, a cestode with worldwide distribution. Data on the circulating Echinococcus granulosus genotypes in Greek livestock is scant. The aim of the present study was to conduct a genetic analysis of 82 Echinococcus granulosus isolates from ruminants in Greece, including areas which until today have not been the subject of studies. The analysis relied on a PCR assay targeting cytochrome c oxidase, subunit 1 gene (CO1), followed by bidirectional sequence analysis of the amplification product. Eighty (n=80) of the 82 (97.6%) isolates were allocated to Echinococcus granulosus sensu stricto (G1-G3) and were classified in 13 distinct haplotypes (9 common and 4 novel) with 12 polymorphic sites. The presence of the dominant haplotype EG1 as was documented in the European populations, was indicated in the country. Almost all regions shared the same common haplotype. In comparison to this predominant haplotype, the number of the nucleotide changes in all the other haplotypes ranged from 1 to 5. All nucleotide changes proved to be transitions (A↔G or C↔T). Two fertile hydatid cysts of sheep origin in different areas (Arkadia, Ilia) of the Peloponnese were identified as Echinococcus canadensis (G7 genotype). Copyright © 2016 Elsevier B.V. All rights reserved.

  13. Phenotypic and molecular identification of Sporothrix isolates from an epidemic area of sporotrichosis in Brazil.

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    Oliveira, Manoel Marques Evangelista; Almeida-Paes, Rodrigo; Muniz, Mauro Medeiros; Gutierrez-Galhardo, Maria Clara; Zancope-Oliveira, Rosely Maria

    2011-10-01

    Sporotrichosis has significantly increased in Brazil in the last decade, particularly in the state of Rio de Janeiro, with the occurrence of an epidemic related to zoonotic transmission from cats to humans. Recently, four new phylogenetic species were incorporated into the Sporothrix species complex based on the phenotypic and molecular characteristics, and a new species name (Sporothrix brasiliensis) was proposed for some of the Sporothrix isolates from this epidemic. This study describes the characterization of 246 isolates obtained from patients attending the Laboratory of Infectious Dermatology, IPEC-FIOCRUZ, between 1998 and 2008, together with one environmental sample. Two hundred and six of the isolates (83.4%) were characterized as S. brasiliensis, 15 (6.0%) as S. schenckii, and one (0.5%) as S. mexicana. Twenty-five isolates (10.1%) could not be identified according to their phenotype and were classified as Sporothrix spp. The calmodulin gene was sequenced to confirm the identity of these isolates. The molecular analysis demonstrated that 24 of the isolates were S. brasiliensis, with the remainder being a S. globosa isolate. The isolate characterized phenotypically as S. mexicana was clustered on the S. schenckii clade. The correlation between molecular data and phenotypic characteristics described in this study is fundamental to the identification of the Sporothrix complex.

  14. Species identification and molecular typing of human Brucella isolates from Kuwait.

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    Mustafa, Abu S; Habibi, Nazima; Osman, Amr; Shaheed, Faraz; Khan, Mohd W

    2017-01-01

    Brucellosis is a zoonotic disease of major concern in Kuwait and the Middle East. Human brucellosis can be caused by several Brucella species with varying degree of pathogenesis, and relapses are common after apparently successful therapy. The classical biochemical methods for identification of Brucella are time-consuming, cumbersome, and provide information limited to the species level only. In contrast, molecular methods are rapid and provide differentiation at intra-species level. In this study, four molecular methods [16S rRNA gene sequencing, real-time PCR, enterobacterial repetitive intergenic consensus (ERIC)-PCR and multilocus variable-number tandem-repeat analysis (MLVA)-8, MLVA-11 and MLVA-16 were evaluated for the identification and typing of 75 strains of Brucella isolated in Kuwait. 16S rRNA gene sequencing of all isolates showed 90-99% sequence identity with B. melitensis and real-time PCR with genus- and species- specific primers identified all isolates as B. melitensis. The results of ERIC-PCR suggested the existence of 75 ERIC genotypes of B. melitensis with a discriminatory index of 0.997. Cluster classification of these genotypes divided them into two clusters, A and B, diverging at ~25%. The maximum number of genotypes (n = 51) were found in cluster B5. MLVA-8 analysis identified all isolates as B. melitensis, and MLVA-8, MLVA-11 and MLVA-16 typing divided the isolates into 10, 32 and 71 MLVA types, respectively. Furthermore, the combined minimum spanning tree analysis demonstrated that, compared to MLVA types discovered all over the world, the Kuwaiti isolates were a distinct group of MLVA-11 and MLVA-16 types in the East Mediterranean Region.

  15. Isolation, enumeration, molecular identification and probiotic potential evaluation of lactic acid bacteria isolated from sheep milk

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    L.B. Acurcio

    2014-06-01

    Full Text Available Lactic acid bacteria species were molecularly identified in milk from Lacaune, Santa Inês and crossbred sheep breeds and their in vitro probiotic potential was evaluated. The species identified were Enterococcus faecium (56.25%, E. durans (31.25% and E. casseliflavus (12.5%. No other lactic acid bacteria species, such as lactobacilli, was identified. Most of the isolated enterococci were resistant to gastric pH (2.0 and to 0.3% oxgall. All tested enterococci were resistant to ceftazidime, oxacillin and streptomycin and sensible to clindamycin, erythromycin and penicillin. The resistance to ciprofloxacin, gentamicin, tetracycline and vancomycin varied among tested species. All tested enterococci strongly inhibited (P<0.05 Escherichia coli and Listeria monocytogenes, moderately inhibited E. faecalis and Staphylococcus aureus and did not inhibit Pseudomonas aeruginosa, Salmonella enterica var. Typhimurium and also one E. durans sample isolated from sheep milk. Four samples of E. faecium, one of E. durans and one of E. casseliflavus presented the best probiotic potential.

  16. Molecular identification of Mycobacterium tuberculosis complex isolates from Kermanshah Province, Iran.

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    Moghaddam, Roghieh; Mosavari, Nader; Mahalati, Ardeshir Hesampoor

    2016-12-01

    Tuberculosis is one of the most important zoonotic diseases in the world. Rapid diagnosis of the disease and identification of species is extremely important for proper treatment of the disease as some species of the complex are resistant to the first-line of tuberculosis drugs. The aim of present study was molecular identification of Mycobacterium tuberculosis (MTB) complex isolates from Kermanshah Province, Iran, which were submitted to the Tuberculosis Reference Laboratory at Razi Vaccine and Serum Research Institute (Tehran, Iran). To identify the genus Mycobacterium, all isolates were subjected to 16S rRNA polymerase chain reaction (PCR), and PCR-IS6110 was subsequently used to confirm that the isolates belonged to MTB complex. Finally, region of difference (RD) typing was used to identify the species in the complex. The results of 16S rRNA and IS6110 PCR analysis showed the presence of 543-bp and 245-bp bands, respectively. Furthermore, 146bp, 172bp, 235bp, and 369bp at RD1, RD4, RD9, and RD12, respectively, were observed during RD typing. Thus, based on the results, all isolates were identified as MTB. It is worth mentioning that most tuberculosis cases are identified on the basis of acid-fast bacilli detection, and antibiotic therapy is immediately initiated subsequently. Moreover, it should be noted that some of these acid-fast positive cases might not be of genus Mycobacterium, and thus, the antibiotics prescribed might threaten the health of the patients. Additionally, if the identified bacilli are not within MTB complex, the drug therapy would differ. However, Mycobacterium bovis, which is a member of MTB complex and is resistant to pyrazinamide, requires exact strain identification. Based on the findings, individual isolates should be identified by RD typing methods, which could clearly discriminate the species from each other. Copyright © 2016.

  17. Molecular identification of Mycobacterium tuberculosis complex isolates from Kermanshah Province, Iran

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    Roghieh Moghaddam

    2016-01-01

    Full Text Available Tuberculosis is one of the most important zoonotic diseases in the world. Rapid diagnosis of the disease and identification of species is extremely important for proper treatment of the disease as some species of the complex are resistant to the first-line of tuberculosis drugs. The aim of present study was molecular identification of Mycobacterium tuberculosis (MTB complex isolates from Kermanshah Province, Iran, which were submitted to the Tuberculosis Reference Laboratory at Razi Vaccine and Serum Research Institute (Tehran, Iran. To identify the genus Mycobacterium, all isolates were subjected to 16S rRNA polymerase chain reaction (PCR, and PCR-IS6110 was subsequently used to confirm that the isolates belonged to MTB complex. Finally, region of difference (RD typing was used to identify the species in the complex. The results of 16S rRNA and IS6110 PCR analysis showed the presence of 543-bp and 245-bp bands, respectively. Furthermore, 146bp, 172bp, 235bp, and 369bp at RD1, RD4, RD9, and RD12, respectively, were observed during RD typing. Thus, based on the results, all isolates were identified as MTB. It is worth mentioning that most tuberculosis cases are identified on the basis of acid-fast bacilli detection, and antibiotic therapy is immediately initiated subsequently. Moreover, it should be noted that some of these acid-fast positive cases might not be of genus Mycobacterium, and thus, the antibiotics prescribed might threaten the health of the patients. Additionally, if the identified bacilli are not within MTB complex, the drug therapy would differ. However, Mycobacterium bovis, which is a member of MTB complex and is resistant to pyrazinamide, requires exact strain identification. Based on the findings, individual isolates should be identified by RD typing methods, which could clearly discriminate the species from each other.

  18. Comparison of biochemical and molecular methods for the identification of bacterial isolates associated with failed loggerhead sea turtle eggs.

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    Awong-Taylor, J; Craven, K S; Griffiths, L; Bass, C; Muscarella, M

    2008-05-01

    Comparison of biochemical vs molecular methods for identification of microbial populations associated with failed loggerhead turtle eggs. Two biochemical (API and Microgen) and one molecular methods (16s rRNA analysis) were compared in the areas of cost, identification, corroboration of data with other methods, ease of use, resources and software. The molecular method was costly and identified only 66% of the isolates tested compared with 74% for API. A 74% discrepancy in identifications occurred between API and 16s rRNA analysis. The two biochemical methods were comparable in cost, but Microgen was easier to use and yielded the lowest discrepancy among identifications (29%) when compared with both API 20 enteric (API 20E) and API 20 nonenteric (API 20NE) combined. A comparison of API 20E and API 20NE indicated an 83% discrepancy between the two methods. The Microgen identification system appears to be better suited than API or 16s rRNA analysis for identification of environmental isolates associated with failed loggerhead eggs. Most identification methods are not intended for use with environmental isolates. A comparison of identification systems would provide better options for identifying environmental bacteria for ecological studies.

  19. Molecular identification of Giardia duodenalis isolates from domestic dogs and cats in Wroclaw, Poland.

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    Piekarska, Jolanta; Bajzert, Joanna; Gorczykowski, Michał; Kantyka, Magdalena; Podkowik, Magdalena

    2016-09-01

    Giardia duodenalis (G. intestinalis) is a common protozoan causing gastrointestinal disorders in many species of mammals. The genus of Giardia has high molecular diversity. Dogs and cats, in addition to their typical infection with assemblages C, D and F, may be a reservoir of zoonotic assemblages (A and B). The aim of this study was a genetic characteristic of Giardia isolates of dogs and cats from the area of Wroclaw (Poland). A total of 128 and 33 faecal samples from dogs and cats, respectively, were analyzed by routine coprological methods. The animals were diagnosed on the presence of G. duodenalis antigens in faeces soluble with the use of SNAP Giardia (IDEXX Laboratories) immunosorbent assay. 27 DNA isolates of Giardia were subjected to molecular identification (PCR-RFLP). The prevalence of G. duodenalis was 21.1% (27/128) in dogs and 15.1% (5/33) in cats. In dogs, C assemblage was present in 18 (81%) positive stool samples, D assemblage in 2 (9%) samples, B assemblage present in one (4.5%), and mixed assemblages (C and D) occurred in one (4.5%) sample. F assemblage was found in 4 (80%) cats' positive stool samples and A assemblage occurred in one case (20%). Confirmation of the presence of A and B zoonotic assemblages suggests that infected pets can be a threat to human health. This study describes for the first time the presence of mixed infections within host-specific C and D assemblages in dogs in Poland.

  20. Isolation and Molecular Identification of Some Blue-Green Algae (Cyanobacteria from Freshwater Sites in Tokat Province of Turkey

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    Tunay Karan

    2017-11-01

    Full Text Available Collected blue-green algae (cyanobacteria from freshwater sites throughout Tokat province and its outlying areas were isolated in laboratory environment and their morphological systematics were determined and also their species identifications were studied by molecular methods. Seven different species of blue-green algae collected from seven different sites were isolated by purifying in cultures in laboratory environment. DNA extractions were made from isolated cells and extracted DNAs were amplified by using PCR. Cyanobacteria specific primers were used to amplify 16S rRNA and phycocyanine gene regions using PCR. Phylogenetic identification of species were conducted by evaluation of obtained sequence analysis data by using computer software. According to species identification by sequence analysis, it was seen that molecular data supports morphological systematics.

  1. Isolation and molecular identification of endophytic diazotrophs from seeds and stems of three cereal crops.

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    Huawei Liu

    Full Text Available Ten strains of endophytic diazotroph were isolated and identified from the plants collected from three different agricultural crop species, wheat, rice and maize, using the nitrogen-free selective isolation conditions. The nitrogen-fixing ability of endophytic diazotroph was verified by the nifH-PCR assay that showed positive nitrogen fixation ability. These identified strains were classified by 879F-RAPD and 16S rRNA sequence analysis. RAPD analyses revealed that the 10 strains were clustered into seven 879F-RAPD groups, suggesting a clonal origin. 16S rRNA sequencing analyses allowed the assignment of the 10 strains to known groups of nitrogen-fixing bacteria, including organisms from the genera Paenibacillus, Enterobacter, Klebsiella and Pantoea. These representative genus are not endophytic diazotrophs in the conventional sense. They may have obtained nitrogen fixation ability through lateral gene transfer, however, the evolutionary forces of lateral gene transfer are not well known. Molecular identification results from 16S rRNA analyses were also confirmed by morphological and biochemical data. The test strains SH6A and MZB showed positive effect on the growth of plants.

  2. The molecular identification of Streptococcus equi subsp. equi strains isolated within New Zealand.

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    Patty, O A; Cursons, R T M

    2014-03-01

    To identify Streptococcus equi subsp. equi (S. equi) by PCR analysis and obtain isolates by culture, in order to investigate the strains of S. equi infecting horses within New Zealand. A diagnostic PCR, based on the amplification of the seeI gene for S. equi, was used on 168 samples submitted from horses with and without clinical signs of strangles. Samples were also processed and cultured on selective media for the isolation of β-haemolytic colonies. In addition, the hypervariable region of the seM gene of S. equi was amplified and then sequenced for strain typing purposes. Of the 168 samples, 35 tested positive for S. equi using PCR. Thirty-two confirmed samples were from horses with a clinical diagnosis of strangles and three were from horses where clinical information was unavailable. Only 22/35 (63%) confirmed S. equi samples were successfully isolated following culture. Strain typing demonstrated that two novel seM alleles of S. equi were found in New Zealand with SeM-99 strains being restricted to the North Island while SeM-100 strains were found in both North and South Islands. The application of PCR for the laboratory confirmation of strangles allowed for a rapid and sensitive identification of S. equi. Moreover, seM typing revealed that within the samples examined two strains of S. equi co-circulated within the North Island of New Zealand but only one strain in the South Island. PCR reduces the time required to obtain laboratory confirmation of strangles compared with culture methods. It also has greater sensitivity in detecting S. equi infections, which is of particular importance in the detection of carrier animals which normally shed low numbers of bacteria. Additionally, seM molecular typing can differentiate between bacterial strains, assisting in the monitoring of local strains of S. equi subsp. equi causing disease.

  3. Molecular and Phenotypic Identification and Speciation of Malassezia Yeasts Isolated from Egyptian Patients with Pityriasis Versicolor

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    Elshabrawy, Walaa Othman; Sallam, Manar

    2017-01-01

    Introduction Pityriasis Versicolor (PV) is a common health problem caused by genus Malassezia, a lipophilic fungi found as a part of the normal flora of skin. Although PV is common in Egypt, there is little information regarding the Malassezia species distribution in PV patients to date. Aim To spot a light on the distribution and clinico-epidemiological features of the Malassezia species in PV patients and healthy individuals that were established by conventional phenotypic and molecular techniques. Materials and Methods A cross-sectional study including 167 individuals; 137 clinically suspected PV patients attending Mansoura University Hospitals, Egypt and 30 healthy control individuals, was carried out. Characterization of Malassezia species was performed phenotypically by conventional, culture-based methods and biochemical tests. Genomic DNA was extracted from isolated colonies for PCR amplification of the highly conserved 26S rDNA region with further species level identification by Restriction Fragment Length Polymorphism (RFLP) using Hha1 and BstC1 enzymes. The association of Malassezia species with epidemiological profile and clinical characteristics was studied. Results A 94.2% of PV samples and 13.3% of control samples were positive by Potassium Hydroxide (KOH) while 71.5% of PV samples and 16.7% of control samples yielded growth in culture with high statistically significant differences (p=0.0001, for both methods). By phenotypic methods, only 75.5% of isolates from patients were identified as: M. furfur (51.4%), M. globosa, (29.7%), M. restricta (13.5%) and M. pachydermatis (5.4%) while by RFLP technique, six species were revealed: M. furfur (44.9%), M. globosa (24.5%), M. sympodialis (12.2 %), M. restricta (10.2%), M. obtusa (4.1%) and M. pachydermatis (4.1%). Most species were isolated from hypopigmented lesions of PV patients aged between 20-29 years. Neck and back were the most common affected sites. Only M. furfur (10%) and M. globosa (6.7%) were

  4. Molecular and Phenotypic Identification and Speciation of Malassezia Yeasts Isolated from Egyptian Patients with Pityriasis Versicolor.

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    Elshabrawy, Walaa Othman; Saudy, Niveen; Sallam, Manar

    2017-08-01

    Pityriasis Versicolor (PV) is a common health problem caused by genus Malassezia , a lipophilic fungi found as a part of the normal flora of skin. Although PV is common in Egypt, there is little information regarding the Malassezia species distribution in PV patients to date. To spot a light on the distribution and clinico-epidemiological features of the Malassezia species in PV patients and healthy individuals that were established by conventional phenotypic and molecular techniques. A cross-sectional study including 167 individuals; 137 clinically suspected PV patients attending Mansoura University Hospitals, Egypt and 30 healthy control individuals, was carried out. Characterization of Malassezia species was performed phenotypically by conventional, culture-based methods and biochemical tests. Genomic DNA was extracted from isolated colonies for PCR amplification of the highly conserved 26S rDNA region with further species level identification by Restriction Fragment Length Polymorphism (RFLP) using Hha1 and BstC1 enzymes. The association of Malassezia species with epidemiological profile and clinical characteristics was studied. A 94.2% of PV samples and 13.3% of control samples were positive by Potassium Hydroxide (KOH) while 71.5% of PV samples and 16.7% of control samples yielded growth in culture with high statistically significant differences (p=0.0001, for both methods). By phenotypic methods, only 75.5% of isolates from patients were identified as: M. furfur (51.4%), M. globosa , (29.7%), M. restricta (13.5%) and M. pachydermatis (5.4%) while by RFLP technique, six species were revealed: M. furfur (44.9%), M. globosa (24.5%), M. sympodialis (12.2 %), M. restricta (10.2%), M. obtusa (4.1%) and M. pachydermatis (4.1%). Most species were isolated from hypopigmented lesions of PV patients aged between 20-29 years. Neck and back were the most common affected sites. Only M. furfur (10%) and M. globosa (6.7%) were identified in healthy controls. M. furfur and M

  5. Evaluation of molecular markers for Phytophthora ramorum detection and identification using a standardized library of isolates

    Science.gov (United States)

    F.N. Martin; M. Coffey; R. Hamelin; P. Tooley; M. Garbelotto; K. Hughes; T. Kubisiak

    2008-01-01

    A number of molecular diagnostic procedures for detection of Phytophthora ramorum have been reported in the literature. In an effort to evaluate the specificity of 10 of these techniques a standardized DNA library for 317 isolates was assembled that included 60 described species as well as 22 taxonomically unclassified isolates. These were sent blind...

  6. The application of molecular methods in the identification of isolated strains of parainfluenza 3 virus of cattle

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    Veljović Lj.

    2014-01-01

    Full Text Available Bovine parainfluenza 3 virus (PI3 causes respiratory infections in cattle and sheep with great economic losses in livestock. The aim of this investigation was to determine the significance of molecular methods in the identification of isolated strains of PI3 virus. Twenty cattle nasal swabs were analyzed for the presence of PI3 using the standard virology method of virus isolation in MBDK cell line and virus neutralization test. The identification of isolated strains was confirmed by RT-PCR and method of direct sequencing with primers for PI3 fusion (F protein gene. PI3 virus was isolated and identified in four nasal swabs using the standard virology method and RT-PCR. The analysis of nucleotide sequences of isolated PI3 strains showed high similarity with sequences isolated from cattle in Asia. Our results showed that molecular methods are very useful in the diagnosis of PI3 infections as well as for the identification and characterization of PI3 strains in Serbia. [Projekat Ministarstva nauke Republike Srbije, br. 31008 i br. 175073

  7. Isolation and Molecular Identification of Streptomyces spp. with Antibacterial Activity from Northwest of Iran

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    Hadi Maleki

    2013-05-01

    Full Text Available Introduction: Streptomyces are a group of prokaryotes that are usually found in all types of ecosystems including water and soil. This group of bacteria is noteworthy as antibiotic producers; so the isolation and characterization of new species seemed to be crucial in introduction of markedly favorable antibiotics. Therefore, in this study we aim to isolate and characterize novel strains of Streptomyces with high antibiotic production capability. Methods: To achieve this goal, from 140 isolates collected throughout northwest of Iran, 12 selected Streptomyces isolates which exhibited high antibacterial activity against pathogenic bacteria were subjected to PCR reaction for identification via 16S rDNA gene and random amplified polymorphic DNA (RAPD pattern analysis. Results: Analysis of morphological and biochemical characteristics and the 16S rDNA gene sequence indicated that all 12 selected isolates belonged to the genus Streptomyces. Moreover, screening of the isolates with regard to their antimicrobial activity against indicator bacteria as well as their classification using RAPD analysis revealed that G614C1 and K36C5 isolates have considerable antimicrobial activity and high similarity to Streptomyces coelicolor and Sreptomyces albogriseolus, respectively. Conclusion: Since many isolates in this study showed inhibitory effects against pathogenic bacteria, soil of northwest of Iran could be used as a rich source to be explored for novel Streptomyces strains with high potency of antibiotic production.

  8. Molecular identification of endophytic fungi isolated from needle leaves of conifers in bohyeon mountain, Korea.

    Science.gov (United States)

    Yoo, Jae-Joon; Eom, Ahn-Heum

    2012-12-01

    Fungal endophytes are microfungi that live in plants without causing apparent symptoms of infection. This study was conducted to identify endophytic fungi isolated from leaves of coniferous trees in Bohyeon Mountain of Korea. We collected leaves of two species of coniferous trees, Pinus densiflora and Pinus koraiensis, from 11 sites in the study area. A total 58 isolates were obtained and identified using molecular and morphological characteristics. Four species of endophytic fungi were isolated from P. densiflora: Lophodermium conigenum, Leotiomycetes sp., Septoria pini-thunbergii, and Polyporales sp., while two fungal species were isolated from P. koraiensis: Eurotiomycetes sp. and Rhytismataceae sp. The most frequently isolated species were L. conigenum and S. pini-thunbergii.

  9. Molecular Identification and Echinocandin Susceptibility of Candida parapsilosis Complex Bloodstream Isolates in Italy, 2007-2014.

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    Grazia Lovero

    Full Text Available The Candida parapsilosis group encompasses three species: C. parapsilosis, C. orthopsilosis, and C. metapsilosis. Here, we describe the incidence and echinocandin susceptibility profile of bloodstream isolates of these three species collected from patients admitted to an Italian university hospital from 2007 to 2014. Molecular identification of cryptic species of the C. parapsilosis complex was performed using polymerase chain reaction amplification of the gene encoding secondary alcohol dehydrogenase, followed by digestion with the restriction enzyme BanI. Minimum inhibitory concentrations were determined using the broth microdilution method according to European Committee for Antimicrobial Susceptibility Testing (EUCAST EDef 7.2 and Clinical Laboratory Standards Institute (CLSI M27-A3 guidelines, and the results were compared with those obtained using the E-test and Sensititre methods. Of the 163 C. parapsilosis complex isolates, 136 (83.4% were identified as C. parapsilosis, and 27 (16.6% as C. orthopsilosis. The species-specific incidences were 2.9/10,000 admissions for C. parapsilosis and 0.6/10,000 admissions for C. orthopsilosis. No resistance to echinocandins was detected with any of the methods. The percent essential agreement (EA between the EUCAST and E-test/Sensititre methods for anidulafungin, caspofungin, and micafungin susceptibility was, respectively, as follows: C. parapsilosis, 95.6/97.8, 98.5/88.2, and 93.4/96.3; C. orthopsilosis, 92.6/92.6, 96.3/77.8, and 63.0/66.7. The EA between the CLSI and E-test/Sensititre methods was, respectively, as follows: C. parapsilosis, 99.3/100, 98.5/89.0, and 96.3/98.5; C. orthopsilosis, 96.3/92.6, 100/81.5, and 92.6/88.9. Only minor discrepancies, ranging from 16.9% (C. parapsilosis to 11.1% (C. orthopsilosis, were observed between the CLSI and E-test/Sensititre methods. In conclusion, this epidemiologic study shows a typical C. parapsilosis complex species distribution, no echinocandin

  10. Molecular identification and genotyping of Pseudomonas aeruginosa isolated from cystic fibrosis and non-cystic fibrosis patients with bronchiectasis.

    Science.gov (United States)

    Eusebio, Nadia; Amorim, Adelina A; Gamboa, Fernanda; Araujo, Ricardo

    2015-03-01

    There is no standard methodology for the molecular identification and genotyping of Pseudomonas aeruginosa which are frequently isolated in bronchiectasis patients. Hence, the main goal of this work was to propose a methodology capable to simultaneously identify and genotype, in less than 6 h, clinical P. aeruginosa collected from cystic fibrosis (CF) and non-CF patients with bronchiectasis. Molecular analyses were conducted in clinical isolates by testing the newly colony-PCR strategy and SNaPaer assay. A total of 207 isolates of P. aeruginosa were collected from clinical samples. To assess the assay specificity, other Gram-negative non-aeruginosa bacteria, namely Pseudomonas and Burkholderia, were tested. The complete group of 23 markers included in the SNaPaer panel was observed exclusively in P. aeruginosa; more than 18 markers failed in other bacteria. A total of 43 SnaP profiles were obtained for clinical P. aeruginosa, being the profiles highly patient-specific. Six CF patients were colonized with P. aeruginosa isolates with very distinct SnaP profiles, particularly following adjustments on antibiotic therapy, thus suggesting changes on the dynamics and dominance of these bacteria. SnaPaer proved to be a good and reliable tool for identification and genotyping of clinical P. aeruginosa in a single-tube multiplex PCR. Combined with the proposed colony-PCR strategy, SnaPaer assay facilitates the molecular analysis of P. aeruginosa. © FEMS 2014. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  11. Microbiological and molecular identification of bacterial species isolated from nasal and oropharyngeal mucosa of fuel workers in Riyadh, Saudi Arabia.

    Science.gov (United States)

    AlWakeel, Suaad S

    2017-09-01

    This study aimed to determine the bacterial species colonizing the nasal and oropharyngeal mucosa of fuel workers in Central Riyadh, Saudi Arabia on a microbiological and molecular level. Throat and nasal swab samples were obtained from 29 fuel station attendants in the period of time extending from March to May 2014 in Riyadh, Saudi Arabia. Microbiological identification techniques were utilized to identify the bacterial species isolated. Antibiotic sensitivity was assessed for each of the bacterial isolates. Molecular identification techniques based on PCR analysis of specific genomic sequences was conducted and was the basis on which phylogeny representation was done for 10 randomly selected samples of the isolates. Blood was drawn and a complete blood count was conducted to note the hematological indices for each of the study participants. Nineteen bacterial species were isolated from both the nasal cavity and the oropharynx including Streptococcus thoraltensis , alpha-hemolytic streptococci, Staphylococcus hominis , coagulase-negative staphylococci, Leuconostoc mesenteroides , Erysipelothrix rhusiopathiae and several others. We found 100% sensitivity of the isolates to ciprofloxacin, cefuroxime and gentamicin. Whereas cefotaxime and azithromycin posted sensitivities of 85.7% and 91.4%, respectively. Low sensitivities (fuel products may be a contributing factor to bacterial colonization of the respiratory tract in fuel workers.

  12. Molecular identification and pathogenicity of Citrobacter and Serratia species isolated from cultured Oreochromis niloticus

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    Manal I. El-Barbary

    2017-09-01

    Full Text Available This study aimed to isolate and characterize some pathogenic bacterial strains belonging to the family Enterobacteriaceae. They had been isolated from gills, liver, kidney and skin of naturally infected Oreochromis niloticus and had been identified by biochemical test and 16S rRNA gene using four universal primers. Additionally, the isolates were tested for antimicrobial susceptibility, histopathological alterations of liver, kidney and gills and the pathogenicity of the identified isolates for O. niloticus. The results of phylogenetic analysis placed the isolates in the family Enterobacteriaceae (genera Serratia and Citrobacter based on 99% homology. The primer pair (17F and 1390R is the most appropriate pair of universal primers employed for the identification of 16S rRNA gene as it covers as much as possible of the variable regions (Vs. V1 and V2 regions of 16S rRNA gene presented weak evidence of the diversity of the genera Serratia. The mortality rate was 40–60% after challenging O. niloticus by identified isolates, which revealed its sensitivity to ciprofloxacin and norfloxacin. Histological changes showed dilation in sinusoids with severe vacuolar degeneration in the liver, tubular degeneration and hemorrhage between renal tubules with pyknotic nuclei in the kidney, epithelial hyperplasia, aneurism and evident epithelium interstitial edema in gills of O. niloticus. This study concluded that these isolates should be considered as an opportunistic pathogen of O. niloticus. The study also states that the sequencing of 16S rRNA is an important tool for the identification of unknown bacterial species of fish pathogen. Keywords: Citrobacter sp., Serratia sp., Phylogenetic analysis, Histology, Antibiotic sensitivity, Oreochromis niloticus

  13. Identification, characterization and molecular epidemiology of Escherichia coli isolated from lamb and goat kids with diarrhoea

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    Süheyla Türkyılmaz

    2013-01-01

    Full Text Available Neonatal diarrhoea is a serious health problem on commercial farms. Enterovirulent Escherichia coli is a significant aetiological agent of neonatal diarrhoea. In this work, identification and classification of E. coli isolates obtained from lambs and goat kids with diarrhoea were studied along with antibiotic resistance and clonal relationships of enterovirulent strains. A total of 107 E. coli strains isolated from animals on 43 farms were investigated. Specific virulence genes were determined by multiplex and uniplex polymerase chain reaction. Testing of antibiotic susceptibility was carried out by the Vitek II compact system. The relationship of E. coli isolates was determined by enterobacterial repetitive intergenic consensus polymerase chain reaction. A total of 39 (36.4% enterovirulent E. coli strains were identified and of this 19 (48.7% were shiga toxigenic, 12 (30.8% enterotoxigenic and 8 (20.5% enteropathogenic. Three isolates (7.7% were found to be positive for extended spectrum beta lactamase; 10 (25.6% isolates showed multi-drug resistance to antimicrobials. A total of 28 types were detected by enterobacterial repetitive intergenic consensus polymerase chain reaction. Twenty strains had distinct types while 5 types were common for 2 strains and 3 types were common for 3 strains. This is the first current determination of types, clonality and antibiotic resistance of enterovirulent E. coli isolated from small ruminants with diarrhoea. The results of this study showed that the rates of shiga toxigenic, enterotoxigenic and enteropathogenic isolates of E. coli are high in the western part of Turkey. Although these isolates were not clonal, presence of multidrug resistant isolates may cause public health problems.

  14. Identification of benthic diatoms isolated from the eastern tidal flats of the Yellow Sea: Comparison between morphological and molecular approaches.

    Science.gov (United States)

    An, Sung Min; Choi, Dong Han; Lee, Jung Ho; Lee, Howon; Noh, Jae Hoon

    2017-01-01

    Benthic diatoms isolated from tidal flats in the west coast of Korea were identified through both traditional morphological method and molecular phylogenetic method for methodological comparison. For the molecular phylogenetic analyses, we sequenced the 18S rRNA and the ribulose bisphosphate carboxylase large subunit coding gene, rbcL. Further, the comparative analysis allowed for the assessment of the suitability as a genetic marker for identification of closely related benthic diatom species and as potential barcode gene. Based on the traditional morphological identification system, the 61 isolated strains were classified into 52 previously known taxa from 13 genera. However, 17 strains could not be classified as known species by morphological analyses, suggesting a hidden diversity of benthic diatoms. The Blast search on NCBI's Genebank indicated that the reference sequences for most of the species were absent for the benthic diatoms. Of the two genetic markers, the rbcL genes were more divergent than the 18S rRNA genes. Furthermore, a long branch attraction artefact was found in the 18S rRNA phylogeny. These results suggest that the rbcL gene is a more appropriate genetic marker for identification and classification of benthic diatoms. Considering their high diversity and simple shapes, and thus the difficulty associated with morphological classification of benthic diatoms, a molecular approach could provide a relatively easy and reliable classification system. However, this study suggests that more effort should be made to construct a reliable database containing polyphasic taxonomic data for diatom classification.

  15. Isolation and molecular identification chitinase-producing Streptomyces strains and examination of their in-vitro antagonistic effects

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    Alireza Dehnad

    2015-12-01

    Full Text Available Introduction: The chemical fungicides are used widely in the world. To reduce the application of synthetic fungicides in treating plant diseases, biological methods are considered as an alternative way to control plant diseases. Many actinomycetes, particularly Streptomyces species are biological agents against a broad spectrum of fungal plant pathogens. The purpose of this study was using the kitinolitik actinomycetes isolated from soil of Eastern Azerbaijan province In order to produce biological pesticides. Materials and methods: Soil samples were taken from different areas of Eastern Azerbaijan province. According to Streptomyces morphological features, single colonies were isolated. To identify the bacteria by molecular characteristic, the genomic DNA was extracted and then the sequences of 16S rDNA were replicated. By using specific primers the bacterial isolates containing chitinase gene were screened. The isolates consisted Chitinase enzyme and were antagonistically cultured with Alternaria genus which is a fungal plant pathogen. Results: Out of 60 soil collected samples, 31 Streptomyces bacterial isolates were separated. Four isolates showed positive results to selectivity action of the chitinase enzyme. Treatment of 3 bacterial isolates with 2 pathogenic fungi showed that AE09 is the most effective anti-fungal isolates. Discussion and conclusion: Soils in Eastern Azerbaijan province are rich of Streptomyces bacteria which generate antifungal compounds. Obtaining the Streptomyces bacteria which have chitinase gene, can lead to identification of very effective strains as anti-fungal.

  16. Molecular identification of fungi isolated from bean tissues with anthracnose symptoms

    International Nuclear Information System (INIS)

    Vanegas Berrouet, Katherin M; Gutierrez Sanchez, Pablo A; Marin Montoya, Mauricio A

    2014-01-01

    In this work, endophytic fungi from leaves and pods of bean presenting anthracnose symptoms were isolated from plants collected at different municipalities in the province of Antioquia (Colombia). Isolates were identified by sequencing the RDNA its regions together with the examination of reproductive structures during sporulation in culture media. Colletotrichum lindemuthianum, the causal agent of anthracnose was isolated in all samples showing symptoms of this disease. These results were confirmed by duplex PCR using the specific primers CD1/CD2 and CY1/CY2. Additionally, 17 endophytic fungi were obtained. Fourteen isolates did not sporulate in culture media (Mycelia sterilia) but were identified by phylogenetic analysis of the regions as the Ascomycetes: Leptosphaerulina (3), Diaporthe (3), Gibberella (1), Plectosphaerella (1) and Biscogniauxia (1) and the Mitosporic genera phoma (2), Alternaria (2) and Stemphylium (1) Three isolates were identified combining morphological and molecular analysis as Fusarium (2) and Curvularia lunata (1). This work increases our knowledge of the mycobiota of legume plants and will serve as support of future studies aimed at determining the effect of these fungi on the development of anthracnose as well as other problems affecting the bean crop.

  17. Identification and molecular characterization of Chryseobacterium vrystaatense ST1 isolated from oligomineral water of southeast Serbia

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    Kojić M.

    2012-01-01

    Full Text Available The isolation and molecular characterization of bacterial strains isolated from water sources in the Vlasina Mountain in southeast Serbia, confirmed the presence of a new species Chryseobacterium vrystaatense ST1. This Gram- negative species showed an extremely low level of biochemical reactivity in biochemical tests. The gene for 16S rRNA was amplified by PCR using universal primers and sequenced. Comparison of 16S rRNA gene sequence and phenotypic features indicated that the isolate ST belonged to Chryseobacterium vrystaatense. A BLAST search of sequenced 1088 nucleotides of the 16S rRNA gene with all sequences deposited in the NCBI collection showed the highest similarity (98% with the strain Chryseobacterium vrystaatense sp. nov., designated as strain R-23533. The very high homology of these two strains allowed classification of our strain at the species level, but some differences indicate, and indirectly confirm, that the isolate ST is an authentic representative. On the basis of these results, we could conclude that Chryseobacterium vrystaatense ST was for first time isolated in Serbia, which is particularly important when one bears in mind that there are only three sequences of this species deposited in the NCBI collection.

  18. Microbiological and molecular identification of bacterial species isolated from nasal and oropharyngeal mucosa of fuel workers in Riyadh,

    Directory of Open Access Journals (Sweden)

    Suaad S. AlWakeel

    2017-09-01

    Full Text Available This study aimed to determine the bacterial species colonizing the nasal and oropharyngeal mucosa of fuel workers in Central Riyadh, Saudi Arabia on a microbiological and molecular level. Throat and nasal swab samples were obtained from 29 fuel station attendants in the period of time extending from March to May 2014 in Riyadh, Saudi Arabia. Microbiological identification techniques were utilized to identify the bacterial species isolated. Antibiotic sensitivity was assessed for each of the bacterial isolates. Molecular identification techniques based on PCR analysis of specific genomic sequences was conducted and was the basis on which phylogeny representation was done for 10 randomly selected samples of the isolates. Blood was drawn and a complete blood count was conducted to note the hematological indices for each of the study participants. Nineteen bacterial species were isolated from both the nasal cavity and the oropharynx including Streptococcus thoraltensis, alpha-hemolytic streptococci, Staphylococcus hominis, coagulase-negative staphylococci, Leuconostoc mesenteroides, Erysipelothrix rhusiopathiae and several others. We found 100% sensitivity of the isolates to ciprofloxacin, cefuroxime and gentamicin. Whereas cefotaxime and azithromycin posted sensitivities of 85.7% and 91.4%, respectively. Low sensitivities (<60% sensitivity to the antibiotics ampicillin, erythromycin, clarithromycin and norfloxacin were observed. Ninety-seven percent similarity to the microbial bank species was noted when the isolates were compared to it. Most hematological indices recorded were within the normal range. In conclusion, exposure to toxic fumes and compounds within fuel products may be a contributing factor to bacterial colonization of the respiratory tract in fuel workers.

  19. Microbial quality and molecular identification of cultivable microorganisms isolated from an urban drinking water distribution system (Limassol, Cyprus).

    Science.gov (United States)

    Botsaris, George; Kanetis, Loukas; Slaný, Michal; Parpouna, Christiana; Makris, Konstantinos C

    2015-12-01

    Microorganisms can survive and multiply in aged urban drinking water distribution systems, leading to potential health risks. The objective of this work was to investigate the microbial quality of tap water and molecularly identify its predominant cultivable microorganisms. Tap water samples collected from 24 different households scattered in the urban area of Limassol, Cyprus, were microbiologically tested following standard protocols for coliforms, E. coli, Pseudomonas spp., Enterococcus spp., and total viable count at 22 and 37 °C. Molecular identification was performed on isolated predominant single colonies using 16SrRNA sequencing. Approximately 85% of the household water samples were contaminated with one or more microorganisms belonging to the genera of Pseudomonas, Corynebacterium, Agrobacterium, Staphylococcus, Bacillus, Delftia, Acinetobacter, Enterococcus, Enterobacter, and Aeromonas. However, all samples tested were free from E. coli. This is the first report in Cyprus molecularly confirming specific genera of relevant microbial communities in tap water.

  20. Molecular identification of fungal isolates and hatching success of green turtle (Chelonia mydas) nests.

    Science.gov (United States)

    Candan, Esra Deniz

    2018-02-26

    The aim of this study is to investigate the fungal diversity of green turtle nests and to examine phylogenetic relationships among these isolates. During the nesting season, samples of intra-nest sand and failed eggs were collected from 25% of the surviving nests in Sugözü Beaches, which are amongst the most important nesting beaches for endangered green turtles in the Mediterranean. Twenty-three fungi were identified by molecular techniques. Fungal isolates belonged to genera Aspergillus, Emericella, Rhizopus, Actinomucor and Apophysomyces with two undescribed species. Aspergillus variecolor, Aspergillus quadrilinieatus, Aspergillus tubingensis, Rhizopus oryzae, Actinomucor elegans and Apophysomyces variabilis were firstly detected in all sea turtle nests within this study. Our results demonstrate that 36.4% of the nests had fungal contamination. Also hatching success of the nests contaminated by fungi were significantly lower than the uncontaminated nests (P = 0.029). Also, this may represent a threat to marine turtles and a risk for the health of conservation workers. This study is the first molecular phylogenetic study associated with sea turtle nests in the eastern Mediterranean coast and contributes to the wider body of literature on fungal invasion of sea turtle nests with firstly isolated species. These findings are important for improving potential conservation measures for the nest sites.

  1. Isolation and molecular identification of Naegleria fowleri from Nile river, Egypt.

    Science.gov (United States)

    Al-Herrawy, Ahmad Z; Gad, Mahmoud A

    2015-12-01

    Members of the genus Naegleria are free-living amoebae distributed in various aquatic environments. Naegleria fowleri is the only species that can cause fatal primary amoebic meningoencephalitis in humans. A total of 48 Nile water samples were collected from the water stream passing though Cairo. The samples were processed for the detection of Naegleria spp. using non-nutrient agar at 45°C. The isolates of Naegleria spp. were identified based on the morphologic criteria of trophozoite, flagellated and cyst stages. Molecular characterization of the isolates was performed using PCR. The obtained results showed that Naegleria spp. were found in 45.8% of Nile water samples by means of microscopic examination. Seasonally, the highest prevalence of Naegleria spp. was recorded in summer (66.7%). Moreover, the highest prevalence of N. fowleri was recorded in summer (25%). The occurrence of heat-tolerant Naegleria spp., especially N. fowleri, in Nile water should be considered as a potential health threat.

  2. Isolation, molecular identification and quinolone-susceptibility testing of Arcobacter spp. isolated from fresh vegetables in Spain.

    Science.gov (United States)

    González, Ana; Bayas Morejón, Isidro Favián; Ferrús, María Antonia

    2017-08-01

    Some species of the Arcobacter genus are considered emerging foodborne and waterborne enteropathogens. However, the presence of Arcobacter spp. in vegetables very little is known, because most studies have focused on foods of animal origin. On the other hand, quinolones are considered as first-line drugs for the treatment of infection by campylobacteria in human patients, but few data are currently available about the resistance levels to these antibiotics among Arcobacter species. Therefore, the aim of this study was to investigate the presence and diversity of arcobacters isolated from fresh vegetables such as lettuces, spinaches, chards and cabbages. Resistance to quinolones of the isolates was also investigated. One hundred fresh vegetables samples purchased from seven local retail markets in Valencia (Spain) during eight months were analysed. The study included 41 lettuces, 21 spinaches, 34 chards and 4 cabbages. Samples were analysed by culture and by molecular methods before and after enrichment. By culture, 17 out of 100 analysed samples were Arcobacter positive and twenty-five isolates were obtained from them. Direct detection by PCR was low, with only 4% Arcobacter spp. positive samples. This percentage increased considerably, up 20%, after 48 h enrichment. By polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP), 17 out of the 25 isolates were identified as A. butzleri and 8 as A. cryaerophilus. Only two A. butzleri isolates showed resistance to levofloxacin and ciprofloxacin. The sequencing of a fragment of the QRDR region of the gyrA gene from the quinolones-resistant isolates revealed the presence of a mutation in position 254 of this gene (C-T transition). This study is the first report about the presence of pathogenic species of Arcobacter spp. in chards and cabbages and confirms that fresh vegetables can act as transmission vehicle to humans. Moreover, the presence of A. butzleri quinolone resistant in vegetables could

  3. Identification of benthic diatoms isolated from the eastern tidal flats of the Yellow Sea: Comparison between morphological and molecular approaches

    Science.gov (United States)

    Choi, Dong Han; Lee, Jung Ho; Lee, Howon; Noh, Jae Hoon

    2017-01-01

    Benthic diatoms isolated from tidal flats in the west coast of Korea were identified through both traditional morphological method and molecular phylogenetic method for methodological comparison. For the molecular phylogenetic analyses, we sequenced the 18S rRNA and the ribulose bisphosphate carboxylase large subunit coding gene, rbcL. Further, the comparative analysis allowed for the assessment of the suitability as a genetic marker for identification of closely related benthic diatom species and as potential barcode gene. Based on the traditional morphological identification system, the 61 isolated strains were classified into 52 previously known taxa from 13 genera. However, 17 strains could not be classified as known species by morphological analyses, suggesting a hidden diversity of benthic diatoms. The Blast search on NCBI’s Genebank indicated that the reference sequences for most of the species were absent for the benthic diatoms. Of the two genetic markers, the rbcL genes were more divergent than the 18S rRNA genes. Furthermore, a long branch attraction artefact was found in the 18S rRNA phylogeny. These results suggest that the rbcL gene is a more appropriate genetic marker for identification and classification of benthic diatoms. Considering their high diversity and simple shapes, and thus the difficulty associated with morphological classification of benthic diatoms, a molecular approach could provide a relatively easy and reliable classification system. However, this study suggests that more effort should be made to construct a reliable database containing polyphasic taxonomic data for diatom classification. PMID:28622375

  4. Molecular identification and amphotericin B susceptibility testing of clinical isolates of Aspergillus from 11 hospitals in Korea.

    Science.gov (United States)

    Heo, Min Seok; Shin, Jong Hee; Choi, Min Ji; Park, Yeon Joon; Lee, Hye Soo; Koo, Sun Hoe; Lee, Won Gil; Kim, Soo Hyun; Shin, Myung Geun; Suh, Soon Pal; Ryang, Dong Wook

    2015-11-01

    We investigated the species distribution and amphotericin B (AMB) susceptibility of Korean clinical Aspergillus isolates by using two Etests and the CLSI broth microdilution method. A total of 136 Aspergillus isolates obtained from 11 university hospitals were identified by sequencing the internal transcribed spacer (ITS) and β-tubulin genomic regions. Minimal inhibitory concentrations (MICs) of AMB were determined in Etests using Mueller-Hinton agar (Etest-MH) and RPMI agar (Etest-RPG), and categorical agreement with the CLSI method was assessed by using epidemiological cutoff values. ITS sequencing identified the following six Aspergillus species complexes: Aspergillus fumigatus (42.6% of the isolates), A. niger (23.5%), A. flavus (17.6%), A. terreus (11.0%), A. versicolor (4.4%), and A. ustus (0.7%). Cryptic species identifiable by β-tubulin sequencing accounted for 25.7% (35/136) of the isolates. Of all 136 isolates, 36 (26.5%) had AMB MICs of ≥2 μg/mL by the CLSI method. The categorical agreement of Etest-RPG with the CLSI method was 98% for the A. fumigatus, A. niger, and A. versicolor complexes, 87% for the A. terreus complex, and 37.5% for the A. flavus complex. That of Etest-MH was ≤75% for the A. niger, A. flavus, A. terreus, and A. versicolor complexes but was higher for the A. fumigatus complex (98.3%). Aspergillus species other than A. fumigatus constitute about 60% of clinical Aspergillus isolates, and reduced AMB susceptibility is common among clinical isolates of Aspergillus in Korea. Molecular identification and AMB susceptibility testing by Etest-RPG may be useful for characterizing Aspergillus isolates of clinical relevance.

  5. Molecular Identification of Unusual Pathogenic Yeast Isolates by Large Ribosomal Subunit Gene Sequencing: 2 Years of Experience at the United Kingdom Mycology Reference Laboratory▿

    Science.gov (United States)

    Linton, Christopher J.; Borman, Andrew M.; Cheung, Grace; Holmes, Ann D.; Szekely, Adrien; Palmer, Michael D.; Bridge, Paul D.; Campbell, Colin K.; Johnson, Elizabeth M.

    2007-01-01

    Rapid identification of yeast isolates from clinical samples is particularly important given their innately variable antifungal susceptibility profiles. We present here an analysis of the utility of PCR amplification and sequence analysis of the hypervariable D1/D2 region of the 26S rRNA gene for the identification of yeast species submitted to the United Kingdom Mycology Reference Laboratory over a 2-year period. A total of 3,033 clinical isolates were received from 2004 to 2006 encompassing 50 different yeast species. While more than 90% of the isolates, corresponding to the most common Candida species, could be identified by using the AUXACOLOR2 yeast identification kit, 153 isolates (5%), comprised of 47 species, could not be identified by using this system and were subjected to molecular identification via 26S rRNA gene sequencing. These isolates included some common species that exhibited atypical biochemical and phenotypic profiles and also many rarer yeast species that are infrequently encountered in the clinical setting. All 47 species requiring molecular identification were unambiguously identified on the basis of D1/D2 sequences, and the molecular identities correlated well with the observed biochemical profiles of the various organisms. Together, our data underscore the utility of molecular techniques as a reference adjunct to conventional methods of yeast identification. Further, we show that PCR amplification and sequencing of the D1/D2 region reliably identifies more than 45 species of clinically significant yeasts and can also potentially identify new pathogenic yeast species. PMID:17251397

  6. Bovine Herpesvirus 4 in Parana State, Brazil: case report, viral isolation, and molecular identification

    Directory of Open Access Journals (Sweden)

    Ernesto Renato Kruger

    2015-03-01

    Full Text Available Bovine Herpesvirus 4 (BoHV-4 is a member of Gammaherpesvirinaesub-family and belongs to genus Rhadinovirus. This virus has been associated with different clinical manifestations and research activity has put forward a strong correlation among virus infection, postpartum metritis, and abortion. The goal of this work was to characterize a virus strain isolate from a cow’s uterine outflow. From swabs drawn of uterine secretion, a virus strain was isolated and characterized by its cytopathology, morphology, and molecular biology approaches. In culture there was CPE development, characterized mainly by long strands with several small balloons along them, radiated from infected cells. Electron microscopy analysis revealed virus particles that had icosahedrical capsid symmetry surrounded by a loose envelope, typical of a herpesvirus. A 2,571 bp PCR product after HindIII digestion generated four fragments, whose base pair composition were 403, 420, 535, and 1,125 bp. Restriction enzymes HindIII and BamHI generated the expected diagnostic bands as well as a 2,350 bp hypermolar fragment as a result of BamHI treatment to demonstrate that agent was a bovine herpesvirus 4, appertaining to DN-599 group.

  7. Morphological and molecular identification of filamentous Aspergillus flavus and Aspergillus parasiticus isolated from compound feeds in South Africa.

    Science.gov (United States)

    Iheanacho, Henry E; Njobeh, Patrick B; Dutton, Francis M; Steenkamp, Paul A; Steenkamp, Lucia; Mthombeni, Julian Q; Daru, Barnabas H; Makun, Anthony H

    2014-12-01

    Isolation of filamentous species of two Aspergillum genera from compound feeds produced in South Africa, and subsequent extraction of their individual DNA in this study, presents a simple but rapid molecular procedure for high through-put analysis of the individual morphological forms. DNA was successfully isolated from the Aspergillus spp. from agar cultures by use of a commercial kit. Agarose gel electrophoresis fractionation of the fungi DNA, showed distinct bands. The DNA extracted by this procedure appears to be relatively pure with a ratio absorbance at 260 and 280 nm. However, the overall morphological and molecular data indicated that 67.5 and 51.1% of feed samples were found to be contaminated with Aspergillus flavus and Aspergillus parasiticus, respectively, with poultry feed having the highest contamination mean level of 5.7 × 105 CFU/g when compared to cattle (mean: 4.0 × 106 CFU/g), pig (mean: 2.7 × 104 CFU/g) and horse (1.0 × 102 CFU) feed. This technique presents a readily achievable, easy to use method in the extraction of filamentous fungal DNA and it's identification. Hence serves as an important tool towards molecular study of these organisms for routine analysis check in monitoring and improving compound feed quality against fungal contamination. Copyright © 2014 Elsevier Ltd. All rights reserved.

  8. Identification and molecular epidemiology of dermatophyte isolates by repetitive-sequence-PCR-based DNA fingerprinting using the DiversiLab system in Turkey.

    Science.gov (United States)

    Koc, A Nedret; Atalay, Mustafa A; Inci, Melek; Sariguzel, Fatma M; Sav, Hafize

    2017-05-01

    Dermatophyte species, isolation and identification in clinical samples are still difficult and take a long time. The identification and molecular epidemiology of dermatophytes commonly isolated in a clinical laboratory in Turkey by repetitive sequence-based PCR (rep-PCR) were assessed by comparing the results with those of reference identification. A total of 44 dermatophytes isolated from various clinical specimens of 20 patients with superficial mycoses in Kayseri and 24 patients in Hatay were studied. The identification of dermatophyte isolates was based on the reference identification and rep-PCR using the DiversiLab System (BioMerieux). The genotyping of dermatophyte isolates from different patients was determined by rep-PCR. In the identification of dermatophyte isolates, agreement between rep-PCR and conventional methods was 87.8 % ( 36 of 41). The dermatophyte strains belonged to four clones (A -D) which were determined by the use of rep-PCR. The dermatophyte strains in Clone B, D showed identical patterns with respect to the region. In conclusion, rep-PCR appears to be useful for evaluation of the identification and clonal relationships between Trichophyton rubrum species complex and Trichophyton mentagrophytes species complex isolates. The similarity and diversity of these isolates may be assessed according to different regions by rep-PCR. © 2017 Blackwell Verlag GmbH.

  9. Isolation, biochemical and molecular identification of Nocardia species among TB suspects in northeastern, Tanzania; a forgotten or neglected threat?

    Science.gov (United States)

    Hoza, Abubakar S; Mfinanga, Sayoki G S; Moser, Irmgard; König, Brigitte

    2017-06-08

    Pulmonary nocardiosis mimic pulmonary tuberculosis in most clinical and radiological manifestations. In Tanzania, where tuberculosis is one of the major public health threat clinical impact of nocardiosis as the cause of the human disease remains unknown. The objective of the present study was to isolate and identify Nocardia isolates recovered from TB suspects in Northeastern, Tanzania by using biochemical and molecular methods. The study involved 744 sputum samples collected from 372 TB suspects from four periphery diagnostic centers in Northeastern, Tanzania. Twenty patients were diagnosed as having presumptively Nocardia infections based on microscopic, cultural characteristics and biomèrieux ID 32C Yeast Identification system and confirmed using 16S rRNA and hsp65 gene specific primers for Nocardia species and sequencing. Biochemically, the majority of the isolates were N. asteroides (n = 8/20, 40%), N. brasiliensis (n = 4/20, 20%), N. farcinica (n = 3/20, 15%), N. nova (n = 1/20, 5%). Other aerobic actinomycetales included Streptomyces cyanescens (n = 2/20, 10%), Streptomyces griseus, Actinomadura madurae each (n = 1/20, 5%). Results of 16S rRNA and hsp65 sequencing were concordant in 15/17 (88. 2%) isolates and discordant in 2/17 (11.8%) isolates. Majority of the isolates belonged to N. cyriacigeorgica and N. farcinica, four (23.5%) each. Our findings suggest that Nocardia species may be an important cause of pulmonary nocardiosis that is underdiagnosed or ignored. This underscores needs to consider pulmonary nocardiosis as a differential diagnosis when there is a failure of anti-TB therapy and as a possible cause of human infections.

  10. Molecular detection and species identification of Enterocytozoon bieneusi isolated from immunocompetent Orang Asli in Malaysia.

    Science.gov (United States)

    Ashikin, Azah; Al-Mekhlafi, Hesham M; Moktar, Norhayati; Anuar, Tengku Shahrul

    2017-04-01

    Most studies of opportunistic infections focus on immunocompromised patients. However, there is a lack of information on microsporidiosis in healthy people (immunocompetent) worldwide. This study aimed to detect and identify microsporidia species in immunocompetent Orang Asli living in Pahang, Malaysia. Orang Asli is a collective term for a group of indigenous people that usually reside in the interior regions of Peninsular Malaysia. They comprise about 0.7% of the total population in Malaysia and 76% of them lived below the poverty line i.e., poor housing conditions with the lack of access to safe drinking water and adequate sanitation, contaminated environment, high illiteracy rate and unhygienic practices by these people. Stool samples were collected from 209 Orang Asli and analyzed for detecting the presence of Enterocytozoon bieneusi and Encephalitozoon intestinalis by polymerase chain reaction assay targeting small subunit ribosomal RNA gene. E. bieneusi was detected in 8 individuals (3.83%). This infection was commonly found in males than females (5.2% vs. 2.7%). All infected Orang Asli were adults, with a mean age of 44years. Diarrhea and other gastrointestinal symptoms were reported in one case (12.5%) among individuals infected with this species. These findings clearly show that exposure to E. bieneusi may actually be common than reported. The accurate detection and identification of microsporidian species by molecular technique will improve therapy, clinical manifestations and prognosis of this infection, as no antiparasitic therapy has been approved for E. bieneusi. It is hoped that these findings will allow the formulation of better health management and disease prevention advisories, and improvement in the standards of health in similar communities. Copyright © 2017 Elsevier B.V. All rights reserved.

  11. Molecular identification and antifungal susceptibility profiles of Candida parapsilosis complex species isolated from culture collection of clinical samples

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    Fábio Silvestre Ataides

    2015-08-01

    Full Text Available AbstractINTRODUCTION:Candida parapsilosis is a common yeast species found in cases of onychomycosis and candidemia associated with infected intravascular devices. In this study, we differentiated Candida parapsilosis sensu stricto, Candida orthopsilosis , and Candida metapsilosis from a culture collection containing blood and subungual scraping samples. Furthermore, we assessed the in vitro antifungal susceptibility of these species to fluconazole, itraconazole, voriconazole, posaconazole, amphotericin B, and caspofungin.METHODS:Differentiation of C. parapsilosis complex species was performed by amplification of the secondary alcohol dehydrogenase (SADH gene and digestion by the restriction enzyme Ban I. All isolates were evaluated for the determination of minimal inhibitory concentrations using Etest, a method for antifungal susceptibility testing.RESULTS:Among the 87 isolates, 78 (89.7% were identified as C. parapsilosis sensu stricto , five (5.7% were identified as C. orthopsilosis , and four (4.6% were identified as C. metapsilosis . Analysis of antifungal susceptibility showed that C. parapsilosis sensu strictoisolates were less susceptible to amphotericin B and itraconazole. One C. parapsilosis sensu stricto isolate was resistant to amphotericin B and itraconazole. Moreover, 10.2% of C. parapsilosis sensu stricto isolates were resistant to caspofungin. Two C. parapsilosis sensu strictoisolates and one C. metapsilosis isolate were susceptible to fluconazole in a dose-dependent manner.CONCLUSIONS:We reported the first molecular identification of C. parapsilosiscomplex species in State of Goiás, Brazil. Additionally, we showed that although the three species exhibited differences in antifungal susceptibility profiles, the primary susceptibility of this species was to caspofungin.

  12. Molecular identification of a Trichinella isolate from a naturally infected pig in Tibet, China.

    Science.gov (United States)

    Li, Ling Zhao; Wang, Zhong Quan; Jiang, Peng; Zhang, Xi; Ren, Hui Jun; Cui, Jing

    2011-12-01

    The first human case with trichinellosis was reported in 1964 in Tibet, China. However, up to the present, the etiological agent of trichinellosis has been unclear. The aim of this study was to identify a Tibet Trichinella isolate at a species level by PCR-based methods. Multiplex PCR revealed amplicon of the expected size (173 bp) for Trichinella spiralis in assays containing larval DNA from Tibet Trichinella isolate from a naturally infected pig. The Tibet Trichinella isolate was also identified by PCR amplification of the 5S ribosomal DNA intergenic spacer region (5S ISR) and mitochondrial large-subunit ribosomal RNA (mt-lsrDNA) gene sequences. The results showed that 2 DNA fragments (749 bp and 445 bp) of the Tibet Trichinella isolate were identical to that of the reference isolates of T. spiralis. The Tibet Trichinella isolate might be classifiable to T. spiralis. This is the first report on T. spiralis in southwestern China.

  13. Morphological and molecular identification of filamentous fungi isolated from cosmetic powders

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    Flavia Cristina Jastale Pinto

    2012-12-01

    Full Text Available Seven fungi were isolated from 50 samples of cosmetic powders. Morphological analyses and ribosomal DNA Internal Transcribed Spacers sequencing were performed which allowed the discrimination of the isolated fungi as Aspergillus fumigatus, Penicillium sp., and Cladosporium sp. which could have, among their species, potentially pathogenic microorganisms.

  14. Molecular identification of TEM-116 beta-lactamase gene in isolates ...

    African Journals Online (AJOL)

    Purpose: Purpose: To determine TEM-116 beta-lactamase gene prevalence in drug-resistant Pseudomonas aeruginosa isolates from Pakistan. Methods: Sequence analysis of TEM beta-lactamase isolates and their antibiotic susceptibility patterns were carried out. Quantitative bacteriostatic concentrations for commonly ...

  15. Molecular identification of TEM-116 beta-lactamase gene in isolates ...

    African Journals Online (AJOL)

    Purpose: Purpose: To determine TEM-116 beta-lactamase gene prevalence in drug-resistant. Pseudomonas aeruginosa isolates from Pakistan. Methods: Sequence analysis of TEM beta-lactamase isolates and their antibiotic susceptibility patterns were carried out. Quantitative bacteriostatic concentrations for commonly ...

  16. Phenotypic and molecular identification of Sporothrix isolates of clinical origin in Northeast China.

    Science.gov (United States)

    Yu, Xiaohong; Wan, Zhe; Zhang, Zhenying; Li, Fuqiu; Li, Ruoyu; Liu, Xiaoming

    2013-08-01

    Sporotrichosis is the most common deep mycosis in Northeast China which is an area of high epidemicity due to contact with reeds or cornstalks. In this study, we have characterized a total of 74 clinical isolates from fixed cutaneous, lymphocutaneous and disseminated clinical forms and from Heilongjiang, Jilin, and Liaoning provinces, respectively. All isolates (previously as Sporothrix schenckii) were identified as Sporothrix globosa according to their phenotypic characteristics and calmodulin gene sequences analysis. They were subdivided into two sub-clades (S. globosa I and S. globosa II). Most of our isolates (71/74) presented restricted growth at 37 °C, which differed from a previous report. Up to now, S. globosa is the only pathogenic species in Northeast China, no matter what kind of clinical form and which region it is isolated from. Most of our clinical isolates (68/74) were clustered with three Chinese environmental isolates reported in the literature. The new findings of S. globosa isolates on division and thermotolerance at 37 °C described in this study will help us gain a better understanding of S. globosa.

  17. Identification of lignionolytic fungi isolated in Mexico using its like molecular marker

    International Nuclear Information System (INIS)

    Rivera-Rios, J. M.; Cruz Ramirez, M. G.; Cruz Madrid, L. C.; Medina Moreno, S. A.; Tellez-Jurado, A.; Arana-Cuenca, A.; Maqueda Galvez, A. P.

    2009-01-01

    Huejutla de Reyes is a place with a warm-humid climate and counts on an annual average temperature of 30 degree centigrade. We collected fungi that growth in wood or trees with the purpose of isolation this ligninolytic fungi in two seasons (one is spring, before raining station and another one in autumn, during raining station). The experimental work was done in the laboratory of the university, where the fungi were isolated. (Author)

  18. Identification and molecular characterization of Roseomonas genomospecies 5 isolated from Umbilical Cord Blood Unit

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    J.M. Bello-López

    2017-01-01

    Conclusions: This is the first report on the isolation of Roseomonas genomospecies 5 in a UCBU for transplantation, an unusual bacteria isolated from umbilical cord blood, associated with a possible immunosuppression in the donor. Its presence in UCBU can be fatal in immunocompromised patients if it were used for transplantation of Hematopoietic Stem Cells (HSC, due to the potential virulence of the strains and the resistance to antimicrobials commonly used.

  19. Identification and molecular characterization of a novel duck Tembusu virus isolate from Southwest China.

    Science.gov (United States)

    Zhu, Kesen; Huang, Juan; Jia, Renyong; Zhang, Bin; Wang, Mingshu; Zhu, Dekang; Chen, Shun; Liu, Mafeng; Yin, Zhongqiong; Cheng, Anchun

    2015-11-01

    Tembusu virus (TMUV) has caused significant economic losses in the Chinese duck industry and may have been overlooked regarding its zoonotic transmission potential. A novel TMUV isolate (named CQW1) was separated from the liver tissue of a young duck in Southwest China. The CQW1 isolate proliferated in embryonated duck eggs and led to death within 3-4 days post-inoculation. Furthermore, CQW1 replicated in duck embryo fibroblast (DEF) cells and caused a cytopathic effect (CPE). The disease emerged on a duck farm in Southwest China and was reproduced by animal experiment. We found that CQW1 was detectable by RT-PCR in brain and liver tissues of dead ducklings within 5 days after inoculation. Most importantly, concentrated nuclei, neuronophagia and microglial nodules were observed in the brain tissue of the inoculated ducklings, and additionally, the liver tissue was affected, mainly by disordered lobular architecture, degeneration, necrosis and regenerated hepatocytes. Analysis of the complete genome sequence showed that CQW1 was 10,992 nt in length with two nucleotide insertions and shared 96.8% to 99.1% and 98.4% to 99.6% identity at nucleotide and amino acid level, respectively, with Chinese isolates. Phylogenetic analysis of the nucleotide sequences demonstrated that the CQW1 isolate was closely related to other members of the genus Flavivirus and formed a new clade together with the GX2013H isolate. Also, the CQW1 isolate demonstrated the highest average pairwise distance value among the Chinese isolates. In the present study, we obtained evidence that TMUV is present in Southwest China. Extensive pathological and epidemiological studies are urgently needed.

  20. Species identification and molecular characterization of Acanthamoeba isolated from contact lens paraphernalia.

    Science.gov (United States)

    Lee, S M; Choi, Y J; Ryu, H W; Kong, H H; Chung, D I

    1997-06-01

    We applied ribosomal RNA gene (rDNA) polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) to identify Acanthamoeba isolates from contact lens paraphernalia, and characterized these on the basis of mitochondrial DNA (mtDNA) RFLP and isoenzyme analysis. The 22 Acanthamoeba strains used as reference strains were obtained from the American Type Culture Collection. Twenty-eight isolates were classified into six ribogroups, as follows: Acanthamoeba ribogroup (AcRG) 1 consisted of 18 isolates; AcRG 2, of three, AcRG 3, of three; AcRG 4, of two; AcRG 5, of one, and AcRG 6, of one. AcRG 1, which was the most frequently isolated type, was identified as A. lugdunensis, and AcRG 2 as A. hatchetti. AcRG 4 was identified as A. triangularis, while AcRG 3 and AcRG 5 were closely related to A. triangularis. AcRG 6 was identified as A. castellanii. The mtDNA RFLP patterns and zymograms for five isoenzymes of the isolates belonging to a ribogroup were identical to one another. The mtDNA digestion phenotype and zymogram for acid phosphatase (AcP) of AcRG 1 were identical to those of A. lugdunensis L3a and KA/E2, the type strain and corneal isolates from a Korean keratitis patient, respectively. The mtDNA digestion phenotype and zymogram for AcP of AcRG 6 were identical to those of A. castellanii Castellani and KA/E3, the type strain and another corneal isolate found in Korea, respectively. The mtDNA RFLP and zymogram for AcP of AcRG 2 were very similar to those of A. hatchetti BH-2 and Chang, respectively the type strain and a pathogen. The mtDNA RFLP and zymogram for AcP of AcRG 4 were similar to those of A. triangularis SH621, the type strain. The mtDNA RFLP patterns of AcRG 3 and 5 were unique. These results showed that the riboprints, mtDNA RFLP and zymograms of 22 of 28 Acanthamoeba isolates were the same as or very similar to those of the clinical isolates, which can probably be regarded as keratopathogens. More attention should be paid to the prevention

  1. MOLECULAR IDENTIFICATION AND ANTIMICROBIAL RESISTANCE PATTERN OF SEVEN CLINICAL ISOLATES OF Nocardia spp. IN BRAZIL

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    Larissa Anuska Zeni CONDAS

    2015-06-01

    Full Text Available Nocardia is a ubiquitous microorganism related to pyogranulomatous infection, which is difficult to treat in humans and animals. The occurrence of the disease is on the rise in many countries due to an increase in immunosuppressive diseases and treatments. This report of cases from Brazil presents the genotypic characterization and the antimicrobial susceptibility pattern using the disk-diffusion method and inhibitory minimal concentration with E-test® strips. In summary, this report focuses on infections in young adult men, of which three cases were cutaneous, two pulmonary, one neurological and one systemic. The pulmonary, neurological and systemic cases were attributed to immunosuppressive diseases or treatments. Sequencing analysis of the 16S rRNA segments (1491 bp identified four isolates of Nocardia farcinica, two isolates of Nocardia nova and one isolate of Nocardia asiatica. N. farcinica was involved in two cutaneous, one systemic and other pulmonary cases; N. nova was involved in one neurological and one pulmonary case; and Nocardia asiatica in one cutaneous case. The disk-diffusion antimicrobial susceptibility test showed that the most effective antimicrobials were amikacin (100%, amoxicillin/clavulanate (100%, cephalexin (100% and ceftiofur (100%, while isolates had presented most resistance to gentamicin (43%, sulfamethoxazole/trimethoprim (43% and ampicillin (29%. However, on the inhibitory minimal concentration test (MIC test, only one of the four isolates of Nocardia farcinica was resistant to sulfamethoxazole/trimethoprim.

  2. Isolation and identification of burn wound superbugs by molecular technique and their susceptibility to silver nanoparticles

    Science.gov (United States)

    Mala, R.; Celsia, A. S. Ruby

    2018-02-01

    Burn wound is a global problem affecting millions of people. It is the major cause of mortality and morbidity. This study was aimed to isolate and identify the wound isolates by 16S rRNA and to assess their susceptibility to antibiotics and silver nanoparticles. Silver nanoparticles were synthesized using aqueous extract of A.indica. The silver nanoparticles were characterized by FESEM, XRD, FTIR and DSC. Antibacterial susceptibility of the isolates was assessed by well diffusion method. The wound isolates were identified as S.aureus and E.coli. Both isolates were resistant to β lactum antibiotics, aminoglycoside, quinolones and macrolides. The inhibition zone exhibited by all antibiotics against both organisms was less than 5 mm. The size of silver nanoparticles were recorded as 55 nm. XRD confirmed the crystalline nature of the nanoparticles. TGA and DSC of silver nanoparticles showed the loss of weight and the melting point of silver nanoparticles was recorded at 871.3°C. Silver nano particles inhibited S.aureus and E.coli with an inhibition zone of 27 mm and 32 mm respectively. Therefore the study demonstrated that only silver containing dressings can be used in burn wounds infected by multi drug resistant super bugs.

  3. Molecular identification and characterization of Colletotrichum sp. isolates from Tahiti lime, tamarillo, and mango

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    Sanabria Adriana

    2010-11-01

    mango were induced by the species Colletotrichum gloeosporioides, which was also fund in few

    citrus samples. RAMS data analysis indicated the existence of two distinct species groups, with a low similarity index (35%. RAM profiles also showed a clear host differentiation of isolates. The C. acutatum population originated from tamarillo exhibited a narrow and homogeneous genetic base, while the C. acutatum population from Tahiti lime was more heterogeneous and  genetically complex, as determined by the analysis of molecular variance (AMOVA and of Ni-Li coefficient. The C. gloeosporioides

  4. Molecular Identification of Nocardia Isolates from Clinical Samples and an Overview of Human Nocardiosis in Brazil

    OpenAIRE

    Baio, Paulo Victor Pereira; Ramos, Juliana Nunes; dos Santos, Louisy Sanches; Soriano, Morgana Fonseca; Ladeira, Elisa Martins; Souza, M?nica Cristina; Camello, Thereza Cristina Ferreira; Ribeiro, Marcio Garcia; Hirata Junior, Raphael; Vieira, Ver?nica Viana; Mattos-Guaraldi, Ana Lu?za

    2013-01-01

    BackgroundNocardia sp. causes a variety of clinical presentations. The incidence of nocardiosis varies geographically according to several factors, such as the prevalence of HIV infections, transplants, neoplastic and rheumatic diseases, as well as climate, socio-economic conditions and laboratory procedures for Nocardia detection and identification. In Brazil the paucity of clinical reports of Nocardia infections suggests that this genus may be underestimated as a cause of human diseases and...

  5. Optimization and molecular identification of novel cellulose degrading bacteria isolated from Egyptian environment

    Directory of Open Access Journals (Sweden)

    Azhar A. Hussain

    2017-06-01

    Full Text Available Cellulase producing bacteria were isolated from both soil and ward poultry, using CMC (carboxymethylcellulose agar medium and screened by iodine method. Cellulase activity of the isolated bacteria was determined by DNS (dinitrosalicylic acid method. The highly cellulolytic isolates (BTN7A, BTN7B, BMS4 and SA5 were identified on the basis of Gram staining, morphological cultural characteristics, and biochemical tests. They were also identified with 16S rDNA analysis. The phylogenetic analysis of their 16S rDNA sequence data showed that BTN7B has 99% similarity with Anoxybacillus flavithermus, BMS4 has 99% similarity with Bacillus megaterium, SA5 has 99% homology with Bacillus amyloliquefaciens and BTN7A was 99% similar with Bacillus subtilis. Cellulase production by these strains was optimized by controlling different environmental and nutritional factors such as pH, temperature, incubation period, different volumes of media, aeration rate and carbon source. The cellulase specific activity was calculated in each case. In conclusion four highly cellulolytic bacterial strains were isolated and identified and the optimum conditions for each one for cellulase production were determined. These strains could be used for converting plant waste to more useful compounds.

  6. Isolation, screening and molecular identification of novel bacterial strain removing methylene blue from water solutions

    Science.gov (United States)

    Kilany, Mona

    2017-11-01

    The potentially deleterious effects of methylene blue (MB) on human health drove the interest in its removal promptly. Bioremediation is an effective and eco friendly for removing MB. Soil bacteria were isolated and examined for their potential to remove MB. The most potent bacterial candidate was characterized and identified using 16S rRNA sequence technique. The evolutionary history of the isolate was conducted by maximum likelihood method. Some physiochemical parameters were optimized for maximum decolorization. Decolorization mechanism and microbial toxicity study of MB (100 mg/l) and by-products were investigated. Participation of heat killed bacteria in color adsorption have been investigated too. The bacterial isolate was identified as Stenotrophomonas maltophilia strain Kilany_MB 16S ribosomal RNA gene with 99% sequence similarity. The sequence was submitted to NCBI (Accession number = KU533726). Phylogeny depicted the phylogenetic relationships between 16S ribosomal RNA gene, partial sequence (1442 bp), of the isolated strain and other strains related to Stenotrophomonas maltophilia in the GenBank database. The optimal conditions were investigated to be pH 5 at 30 °C, after 24 h using 5 mg/l MB showing optimum decolorization percentage (61.3%). Microbial toxicity study demonstrated relative reduction in the toxicity of MB decolorized products on test bacteria. Mechanism of color removal was proved by both biosorption and biodegradation, where heat-killed and live cells showed 43 and 52% of decolorization, respectively, as a maximum value after 24-h incubation. It was demonstrated that the mechanism of color removal is by adsorption. Therefore, good performance of S maltophilia in MB color removal reinforces the exploitation of these bacteria in environmental clean-up and restoration of the ecosystem.

  7. Identification of carbapenemase-mediated resistance among Enterobacteriaceae bloodstream isolates: A molecular study from India

    Directory of Open Access Journals (Sweden)

    Srujana Mohanty

    2017-01-01

    Full Text Available Acquired resistance in carbapenem-resistant Enterobacteriaceae (CRE conferred by carbapenemases is a major concern worldwide. Consecutive, non-duplicate isolates of Escherichia coli (EC and Klebsiella pneumoniae from clinically diagnosed bloodstream infections were screened for the presence of carbapenem resistance by standard disk-diffusion method and minimum inhibitory concentration breakpoints using the Clinical and Laboratory Standards Institute guidelines. Carbapenemase-encoding genes were amplified by polymerase chain reaction. Of 387 isolates (214 K. pneumoniae, 173 EC tested, 93 (24.03% were found to be CRE. Of these, 71 (76.3% were positive for at least one tested carbapenemase gene. The frequency of carbapenemase genes was New Delhi metallo-β-lactamse-1 (65.6%, oxacillinase (OXA-48 (24.7%, OXA-181 (23.6%, Verona integron-encoded metallo-β-lactamase (6.4% and K. pneumoniae carbapenemase (2.1%. Our study identified presence of carbapenemases in a large proportion of CRE isolates. Delineation of resistance mechanisms is important in view of future therapeutics concerned with the treatment of CRE and for aiding control efforts by surveillance and infection control interventions.

  8. Identification of Taenia asiatica in China: molecular, morphological, and epidemiological analysis of a Luzhai isolate.

    Science.gov (United States)

    Eom, Keeseon S; Jeon, Hyung-Kyu; Kong, Yoon; Hwang, Ui Wook; Yang, Yichao; Li, Xueming; Xu, Longqi; Feng, Zheng; Pawlowski, Zbigniew S; Rim, Han-Jong

    2002-08-01

    Multiple analysis has characterized a recently described tapeworm of people, Taenia asiatica, in mainland China. Six adult tapeworms collected from people of the Zhuang minority residing in the southern part of China (Luzhai isolate) were comparatively analyzed with other tapeworms from people: T. asiatica (n = 2, South Korea), T. saginata (n = 1, Poland; n = 1, Korea), and T. solium (n = 1, People's Republic of China). Experimental infections with eggs from the Luzhai isolate in pigs and cattle produced cysticerci, each with a hookletless scolex and with wartlike formations on the external surface of the bladder wall. There were rostellar protrusions in the scolices of adult worms. Random amplified polymorphic DNA analysis using 3 arbitrary primers produced bands identical to those of the Korean T. asiatica. Conversely, T. saginata and T. solium exhibited different banding patterns. Phylogenetic relationships inferred from the complete nucleotide sequences of the internal transcribed spacer 2 placed the Chinese tapeworms consistently within the T. asiatica clade by 96% bootstrapping value in the maximum likelihood analysis, 96% in maximum parsimony, and 100% in neighbor joining. These collective data demonstrate that T. asiatica is sympatrically distributed with the other 2 species of Taenia in the human host in mainland China.

  9. EnterohemorrhagicEscherichia coliO157 in milk and dairy products from Libya: Isolation and molecular identification by partial sequencing of 16S rDNA.

    Science.gov (United States)

    Garbaj, Aboubaker M; Awad, Enas M; Azwai, Salah M; Abolghait, Said K; Naas, Hesham T; Moawad, Ashraf A; Gammoudi, Fatim T; Barbieri, Ilaria; Eldaghayes, Ibrahim M

    2016-11-01

    The aim of this work was to isolate and molecularly identify enterohemorrhagic Escherichia coli (EHEC) O157 in milk and dairy products in Libya, in addition; to clear the accuracy of cultural and biochemical identification as compared with molecular identification by partial sequencing of 16S rDNA for the existing isolates. A total of 108 samples of raw milk (cow, she-camel, and goat) and locally made dairy products (fermented cow's milk, Maasora, Ricotta and ice cream) were collected from some regions (Janzour, Tripoli, Kremiya, Tajoura and Tobruk) in Libya. Samples were subjected to microbiological analysis for isolation of E. coli that was detected by conventional cultural and molecular method using polymerase chain reaction and partial sequencing of 16S rDNA. Out of 108 samples, only 27 isolates were found to be EHEC O157 based on their cultural characteristics (Tellurite-Cefixime-Sorbitol MacConkey) that include 3 isolates from cow's milk (11%), 3 isolates from she-camel's milk (11%), two isolates from goat's milk (7.4%) and 7 isolates from fermented raw milk samples (26%), isolates from fresh locally made soft cheeses (Maasora and Ricotta) were 9 (33%) and 3 (11%), respectively, while none of the ice cream samples revealed any growth. However, out of these 27 isolates, only 11 were confirmed to be E. coli by partial sequencing of 16S rDNA and E. coli O157 Latex agglutination test. Phylogenetic analysis revealed that majority of local E. coli isolates were related to E. coli O157:H7 FRIK944 strain. These results can be used for further studies on EHEC O157 as an emerging foodborne pathogen and its role in human infection in Libya.

  10. Enterohemorrhagic Escherichia coli O157 in milk and dairy products from Libya: Isolation and molecular identification by partial sequencing of 16S rDNA

    Science.gov (United States)

    Garbaj, Aboubaker M.; Awad, Enas M.; Azwai, Salah M.; Abolghait, Said K.; Naas, Hesham T.; Moawad, Ashraf A.; Gammoudi, Fatim T.; Barbieri, Ilaria; Eldaghayes, Ibrahim M.

    2016-01-01

    Aim: The aim of this work was to isolate and molecularly identify enterohemorrhagic Escherichia coli (EHEC) O157 in milk and dairy products in Libya, in addition; to clear the accuracy of cultural and biochemical identification as compared with molecular identification by partial sequencing of 16S rDNA for the existing isolates. Materials and Methods: A total of 108 samples of raw milk (cow, she-camel, and goat) and locally made dairy products (fermented cow’s milk, Maasora, Ricotta and ice cream) were collected from some regions (Janzour, Tripoli, Kremiya, Tajoura and Tobruk) in Libya. Samples were subjected to microbiological analysis for isolation of E. coli that was detected by conventional cultural and molecular method using polymerase chain reaction and partial sequencing of 16S rDNA. Results: Out of 108 samples, only 27 isolates were found to be EHEC O157 based on their cultural characteristics (Tellurite-Cefixime-Sorbitol MacConkey) that include 3 isolates from cow’s milk (11%), 3 isolates from she-camel’s milk (11%), two isolates from goat’s milk (7.4%) and 7 isolates from fermented raw milk samples (26%), isolates from fresh locally made soft cheeses (Maasora and Ricotta) were 9 (33%) and 3 (11%), respectively, while none of the ice cream samples revealed any growth. However, out of these 27 isolates, only 11 were confirmed to be E. coli by partial sequencing of 16S rDNA and E. coli O157 Latex agglutination test. Phylogenetic analysis revealed that majority of local E. coli isolates were related to E. coli O157:H7 FRIK944 strain. Conclusion: These results can be used for further studies on EHEC O157 as an emerging foodborne pathogen and its role in human infection in Libya. PMID:27956766

  11. Enterohemorrhagic Escherichia coli O157 in milk and dairy products from Libya: Isolation and molecular identification by partial sequencing of 16S rDNA

    Directory of Open Access Journals (Sweden)

    Aboubaker M. Garbaj

    2016-11-01

    Full Text Available Aim: The aim of this work was to isolate and molecularly identify enterohemorrhagic Escherichia coli (EHEC O157 in milk and dairy products in Libya, in addition; to clear the accuracy of cultural and biochemical identification as compared with molecular identification by partial sequencing of 16S rDNA for the existing isolates. Materials and Methods: A total of 108 samples of raw milk (cow, she-camel, and goat and locally made dairy products (fermented cow’s milk, Maasora, Ricotta and ice cream were collected from some regions (Janzour, Tripoli, Kremiya, Tajoura and Tobruk in Libya. Samples were subjected to microbiological analysis for isolation of E. coli that was detected by conventional cultural and molecular method using polymerase chain reaction and partial sequencing of 16S rDNA. Results: Out of 108 samples, only 27 isolates were found to be EHEC O157 based on their cultural characteristics (Tellurite-Cefixime-Sorbitol MacConkey that include 3 isolates from cow’s milk (11%, 3 isolates from she-camel’s milk (11%, two isolates from goat’s milk (7.4% and 7 isolates from fermented raw milk samples (26%, isolates from fresh locally made soft cheeses (Maasora and Ricotta were 9 (33% and 3 (11%, respectively, while none of the ice cream samples revealed any growth. However, out of these 27 isolates, only 11 were confirmed to be E. coli by partial sequencing of 16S rDNA and E. coli O157 Latex agglutination test. Phylogenetic analysis revealed that majority of local E. coli isolates were related to E. coli O157:H7 FRIK944 strain. Conclusion: These results can be used for further studies on EHEC O157 as an emerging foodborne pathogen and its role in human infection in Libya.

  12. Prevalence and molecular identification of Cryptosporidium isolates from pet lizards and snakes in Italy.

    Science.gov (United States)

    Rinaldi, L; Capasso, M; Mihalca, A D; Cirillo, R; Cringoli, G; Cacciò, S

    2012-11-01

    In order to acquire prevalence and genetic data on Cryptosporidium infections in captive lizards and snakes kept as pets, a survey was conducted on 150 individual reptiles from southern Italy. Fecal samples were preserved in 5% formalin and analyzed using a commercial immunofluorescence assay (IFA) for the detection of Cryptosporidium oocysts and Giardia cysts. IFA revealed the presence of Cryptosporidium oocysts in nine of the 150 samples examined (6.0%), precisely in 6/125 snakes (4.8%) and in 3/25 lizards (12.0%); all fecal samples tested negative for the presence of Giardia cysts. Molecular characterization based on nested PCR amplification and sequencing of the SSU-rRNA gene, revealed the presence of Cryptosporidium serpentis in three samples from snakes (Boa constrictor constrictor, Elapheguttata guttata guttata and Python molurus).

  13. Prevalence and molecular identification of Cryptosporidium isolates from pet lizards and snakes in Italy

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    Rinaldi L.

    2012-11-01

    Full Text Available In order to acquire prevalence and genetic data on Cryptosporidium infections in captive lizards and snakes kept as pets, a survey was conducted on 150 individual reptiles from southern Italy. Fecal samples were preserved in 5% formalin and analyzed using a commercial immunofluorescence assay (IFA for the detection of Cryptosporidium oocysts and Giardia cysts. IFA revealed the presence of Cryptosporidium oocysts in nine of the 150 samples examined (6.0%, precisely in 6/125 snakes (4.8% and in 3/25 lizards (12.0%; all fecal samples tested negative for the presence of Giardia cysts. Molecular characterization based on nested PCR amplification and sequencing of the SSU-rRNA gene, revealed the presence of Cryptosporidium serpentis in three samples from snakes (Boa constrictor constrictor, Elapheguttata guttata guttata and Python molurus.

  14. Isolation and molecular identification of Acanthamoeba spp from oasis water in Tunisia.

    Science.gov (United States)

    Dendana, F; Trabelsi, H; Neiji, S; Sellami, H; Kammoun, S; Makni, F; Feki, J; Cheikhrouhou, F; Ayadi, A

    2018-04-01

    In the southern Tunisia Oasis, we conducted 211 water with drawals from various water traffic sites. This water is used for agriculture, swimming or various other human activities. Acanthamoeba genus was detected in 82% of collected samples. Sequencing of the amplification products with primers P892C/P892 has allowed us to detect genotypic variation with predominance of T4 genotype (51%) and presence of the genotypes T14, T5, T3, T16, T15, T10, T11, T9 and T7. They T4, T3, T5, T15, T11 and T10 genotypes have a high potential for pathogenicity and a very high degree of virulence due to their production of serine proteases and extracellular cysteine enzymes involved in tissue degradation of the host. T4 genotype was the most abundant in the environment as well as in infections caused by Acanthamoeba spp. T5 genotype was ranked second and T3 genotype was less abundant in the environment and its pathogenicity is discussed. Acanthamoeba strains with the genotypes T16, T9 and T7 were considered non pathogenic. In fact, they have been isolated only from the environment. However, for these strains, their role as a reservoir can be a real risk to human health. Copyright © 2018. Published by Elsevier Inc.

  15. Isolation, Identification and Molecular Typing of Cryptococcus neoformans from Pigeon Droppings and Other Environmental Sources in Tripoli, Libya.

    Science.gov (United States)

    Ellabib, Mohamed S; Aboshkiwa, Mohamed A; Husien, Walid M; D'Amicis, Roberta; Cogliati, Massimo

    2016-08-01

    Cryptococcus neoformans and C. gattii are the major cause of fungal meningitis, a potentially lethal mycosis. Since pigeon excreta and other environmental sources can be considered a significant environmental reservoir of this species in urban areas, 100 samples of pigeon excreta and 420 samples from Eucalyptus camaldulensis and Olea europaea (olive tree) around the city of Tripoli, Libya, were collected. C. neoformans was isolated and identified using standard biochemical assays from 46 samples: 34 from pigeon droppings, 3 from Eucalyptus trees and 9 from olive trees. Molecular typing revealed that all isolates from pigeon droppings belonged to molecular type VNI (C. neoformans var. grubii) and mating type αA, whereas those from trees included also the molecular type VNII and VNIII (AD hybrids). The present study reports, for the first time, information about the distribution of species, mating types and molecular types of C. neoformans/C. gattii species complex in Libya.

  16. Identification and molecular mapping of a wheat gene for resistance to an unadapted isolate of Colletotrichum cereale.

    Science.gov (United States)

    Inoue, Yoshihiro; Mori, Ryota; Takahashi, Yujiro; Kiguchi, So; Enomoto, Takashi; Chuma, Izumi; Tosa, Yukio

    2013-06-01

    To elucidate genetic mechanisms of host species specificity between graminicolous anthracnose fungi and gramineous plants, infection assays were performed with a Sorghum isolate (Colletotrichum sublineolum), a Digitaria isolate (C. hanaui), a Polypogon isolate (C. cereale), and an Avena isolate (C. cereale). They were specifically virulent on the plants from which they were isolated. When 72 wheat lines were inoculated with an unadapted isolate from Asia Minor bluegrass (Cgp29), however, some exceptional cultivars were recognized. Although most cultivars were resistant to Cgp29, 'Hope' was susceptible. In F2 populations derived from crosses between three resistant cultivars-'Norin 4' (N4), 'Chinese Spring' (CS), and 'Shin-chunaga' (Sch)-and the susceptible Hope, resistant and susceptible seedlings segregated in a 3:1 ratio, suggesting that a major gene is involved in the resistance of each cultivar to Cgp29. In F2 populations derived from crosses between the three resistant cultivars, all seedlings were resistant, suggesting that these three cultivars carry the same gene. This resistance gene was designated as "resistance to Colletotrichum cereale 1" (Rcc1). Analysis with the CS-Hope chromosome substitution lines and molecular mapping revealed that Rcc1 was located on the long arm of chromosome 5A. Cytologically, Rcc1 was mainly associated with hypersensitive reaction. These results suggest that major genes similar to those controlling cultivar specificity are involved in the resistance of wheat against the unadapted isolate of C. cereale.

  17. The Occurrence of Phellinus torulosus in Apulia and Basilicata (Southern Italy : IDentification of Isolates by Morphologic, Microscopic, and Molecular Means

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    G. Campanile

    2004-08-01

    Full Text Available Basidiomes of Phellinus torulosus were found in 9 oak woods in Apulia and Basilicata (Southern Italy; from these basidiomes 138 isolates of P. torulosus were obtained and identified by their morphologic, microscopic, and molecular characteristics. Based on the type of aerial mycelium (fluffy, cottony or powdery and its growth, 9 morphotypes were identified. The morphology of the cultures was not correlated with the microscopic character of the 9 morphotypes. Molecular analysis, such as intergenic transcribed spacers-restriction fragment length polymorphism (ITS-RFLP and sequencing of the ITS region, confirmed the results obtained with microscopy analysis.

  18. Molecular Identification of Isolated Fungi from Unopened Containers of Greek Yogurt by DNA Sequencing of Internal Transcribed Spacer Region

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    Irshad M. Sulaiman

    2014-06-01

    Full Text Available In our previous study, we described the development of an internal transcribed spacer (ITS1 sequencing method, and used this protocol in species-identification of isolated fungi collected from the manufacturing areas of a compounding company known to have caused the multistate fungal meningitis outbreak in the United States. In this follow-up study, we have analyzed the unopened vials of Greek yogurt from the recalled batch to determine the possible cause of microbial contamination in the product. A total of 15 unopened vials of Greek yogurt belonging to the recalled batch were examined for the detection of fungi in these samples known to cause foodborne illness following conventional microbiological protocols. Fungi were isolated from all of the 15 Greek yogurt samples analyzed. The isolated fungi were genetically typed by DNA sequencing of PCR-amplified ITS1 region of rRNA gene. Analysis of data confirmed all of the isolated fungal isolates from the Greek yogurt to be Rhizomucor variabilis. The generated ITS1 sequences matched 100% with the published sequences available in GenBank. In addition, these yogurt samples were also tested for the presence of five types of bacteria (Salmonella, Listeria, Staphylococcus, Bacillus and Escherichia coli causing foodborne disease in humans, and found negative for all of them.

  19. Molecular identification and in-vitro antifungal susceptibility testing of Candida species isolated from patients with onychomycosis

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    Keyvan Pakshir

    2015-03-01

    Full Text Available Background and Purpose: Candida species are the most opportunistic fungi affecting the nails and resulting in onychomycosis. In this study, we identified and evaluated in-vitro susceptibility of the recovered isolates against fluconazole (FLC, voriconazole (VRC, and clotrimazole (CLT using the Clinical and Laboratory Standards Institute (CLSI M27-A3 document. Materials and Methods: From patients with either clinically or mycologically proven onychomycosis, 97 isolates comprising of seven Candida species were isolated, which were identified by both conventional and molecular techniques such as polymerase chain reaction-restriction fragment length polymorphism. In addition, Candida dubliniensis was confirmed by restriction endonuclease analysis. Antifungal susceptibility of each isolate against the three azoles applied in this study was determined using the CLSI microdilution reference method M27-A3. Results: Candida parapsilosis (C. parapsilosis was the most frequently isolated species (n=44, followed by C. albicans (n=23, C. tropicalis (n=13, C. glabrata (n=7, C. krusei (n=6, C. guilliermondii (n=3, and C. dubliniensis (n=1. All the isolates were susceptible to CLT. VRC had lower minimum inhibitory concentration (MIC values for the isolates compared to FLC. Geometric mean MIC values of VRC, FLC, and CLT for C. parapsilosis isolates were 0.07 µg/ml, 0.8 µg/ml, and 0.35 µg/ml, respectively. Collectively, all species exhibited greater susceptibility to VRC in comparison to C. albicans (P≤0.001. Conclusion: This study showed that non-albicans Candida species were the most common etiologic agents of non-dermatophyte onychomycosis. The major antifungal agents used in clinics to empirically treat yeast onychomycosis are FLC and CLT. Our data suggested that CLT is a better choice for the treatment of Candida onychomycosis, especially in drug resistant cases.

  20. Intra- and inter-isolate variation of ribosomal and protein-coding genes in Pleurotus: implications for molecular identification and phylogeny on fungal groups.

    Science.gov (United States)

    He, Xiao-Lan; Li, Qian; Peng, Wei-Hong; Zhou, Jie; Cao, Xue-Lian; Wang, Di; Huang, Zhong-Qian; Tan, Wei; Li, Yu; Gan, Bing-Cheng

    2017-06-26

    The internal transcribed spacer (ITS), RNA polymerase II second largest subunit (RPB2), and elongation factor 1-alpha (EF1α) are often used in fungal taxonomy and phylogenetic analysis. As we know, an ideal molecular marker used in molecular identification and phylogenetic studies is homogeneous within species, and interspecific variation exceeds intraspecific variation. However, during our process of performing ITS, RPB2, and EF1α sequencing on the Pleurotus spp., we found that intra-isolate sequence polymorphism might be present in these genes because direct sequencing of PCR products failed in some isolates. Therefore, we detected intra- and inter-isolate variation of the three genes in Pleurotus by polymerase chain reaction amplification and cloning in this study. Results showed that intra-isolate variation of ITS was not uncommon but the polymorphic level in each isolate was relatively low in Pleurotus; intra-isolate variations of EF1α and RPB2 sequences were present in an unexpectedly high amount. The polymorphism level differed significantly between ITS, RPB2, and EF1α in the same individual, and the intra-isolate heterogeneity level of each gene varied between isolates within the same species. Intra-isolate and intraspecific variation of ITS in the tested isolates was less than interspecific variation, and intra-isolate and intraspecific variation of RPB2 was probably equal with interspecific divergence. Meanwhile, intra-isolate and intraspecific variation of EF1α could exceed interspecific divergence. These findings suggested that RPB2 and EF1α are not desirable barcoding candidates for Pleurotus. We also discussed the reason why rDNA and protein-coding genes showed variants within a single isolate in Pleurotus, but must be addressed in further research. Our study demonstrated that intra-isolate variation of ribosomal and protein-coding genes are likely widespread in fungi. This has implications for studies on fungal evolution, taxonomy

  1. Identification and determination of antibiotic susceptibilities of Brucella strains isolated from patients in van, Turkey by conventional and molecular methods.

    Science.gov (United States)

    Parlak, Mehmet; Güdücüoğlu, Hüseyin; Bayram, Yasemin; Çıkman, Aytekin; Aypak, Cenk; Kılıç, Selçuk; Berktaş, Mustafa

    2013-01-01

    Brucellosis is a worldwide zoonotic disease and still constitutes a major public health problem. In this study, we aimed to identify biovars of Brucella strains isolated from clinical specimens taken from brucellosis patients from the Eastern Anatolia region as well determine the susceptibility of these isolates to tigecycline and azithromycin, drugs that may serve as alternatives to the conventional drugs used in the therapy. Seventy-five Brucella spp. isolates were included in the study. All strains were identified by both conventional and molecular methods. Brucella Multiplex PCR kit (FC-Biotech, Code: 0301, Turkey) and B. melitensis biovar typing PCR kit (FC-Biotech, Code: 0302, Turkey) were used for molecular typing. Antimicrobial susceptibilities of all strains were determined by E-tests. By conventional biotyping, 73 strains were identified as B. melitensis biovar 3 and two strains as B. abortus biovar 3. Molecular typing results were compatible with conventional methods. The MIC50 and MIC90 values of doxycycline were 0.047 and 0.094; tigecycline 0.094 and 0.125; trimethoprim/sulfamethoxazole 0.064 and 0.19; ciprofloxacin 0.19 for both; streptomycin 0.75 and 1; rifampin 1 and 2 and azithromycin 4 and 8. According to the MIC values, doxycycline was found to be the most effective antibiotic, followed by tigecycline, trimethoprim-sulfamethoxazole and ciprofloxacin. Currently recommended antibiotics for the treatment of brucellosis such as doxycycline, rifampin, streptomycin, trimethoprim-sulfamethoxazole and ciprofloxacin were found to be still effective. While our results showed that tigecycline can be used an alternative agent in the treatment of brucellosis, azithromycin has not been confirmed as an appropriate agent for the treatment.

  2. Isolation and molecular identification of lactic acid bacteria from King grass and their application to improve the fermentation quality of sweet Sorghum.

    Science.gov (United States)

    Shah, Assar Ali; Xianjun, Yuan; Zhihao, Dong; Junfeng, Li; Shao, Tao

    2017-12-04

    The aim of the present study was isolation and molecular identification of lactic acid bacteria from King grass and their application to improve the fermentation quality of sweet Sorghum. Seventy-six strains of LAB were isolated; five strains were selected for Physiological and morphological tests and 16S rRNA sequencing. All five strains grew at different pH 3.5-8.0, different temperature 35, 40, 45, 50 °C and different NaCl concentrations 3, 6.5, 9.5%. Strains HDASK were identified Lactobacillus plantarum and SK3907, SK2A32, SK3A42 and ASKDD Pediococcus acidilactici. Three isolated strains and one commercial strain were added to sweet sorghum. Silage was prepared of four treatments and one control with three replicates as control (SKC, adding 2 ml/kg sterilizing water), L. plantarum commercial bacteria (SKP), L. plantarum (HDASK) isolated from King grass (SKA), P. acidilactici (SK3907) isolated from King grass (SKB) and P. acidilactici (ASKDD) isolated from King grass (SKD). All silage were prepared using polyethylene terephthalate bottles, and incubated at room temperature for different ensiling days. The level of pH, acetic acid, NH3-N, water soluble carbohydrate and butyric acid was significantly (P mold and LAB were significantly (P Sorghum silage.

  3. Isolation and molecular identification of a UV-resistant strain of Dietzia maris and antioxidant activity of pigment

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    Narges Zamanian

    2016-09-01

    Full Text Available Introduction: The ability of radioresistant bacteria to survive high levels of UV radiation has been linked to their strong DNA repair systems and ability to produce primary and secondary metabolic products. The biosynthesis of pigments provides an opportunity for bacteria to live in radiation-rich environment. Recent radiation-responsive pigments are used commercially as food colorants, anticancer drugs, as well as antibiotics and for cosmetic purposes. Materials and methods: Soil sample of Omidiyeh city was collected during the spring of 2014 and UV-resistant strain was isolated after primary and secondary screening. Then it was identified by molecular methods (16S rRNA gene sequencing. Antioxidant activity of pigment was evaluated by 2,2 -diphenyl-1-picryl hydrazyl (DPPH and the reducing power of pigments were analyzed by ferric chloride. Results: In this present study, new UV-resistant strain NM2 was isolated and by comparison of these 16S rRNA gene sequences to public database using the BLAST, the genus and species of the isolate was identified as Dietzia maris with 99% similarity. Extraction of pigment from isolated strain was carried out by methanol and acetone as solvents. The spectrum is characterized by maximum peak at 473 nm for pigment of NM2 strain. Antioxidant activity and the reducing ability of pigments increased by increasing their concentrations. NM2 strain pigment showed EC50 concentration of 3.30 mg/ml for DPPH free radical scavenging activity, and EC50 concentration of 28.46 µg/ml for reducing power. Discussion and conclusion: Isolation of natural resources of pigment is very important with high anti-oxidant activity. In the current study, pigment of UV-resistant bacteria demonstrated a strong antioxidant activity in vitro and pigment of these bacteria could play an important role in UV tolerance. Pigment of UV-resistant bacteria may be an appropriate source for antioxidative-related functional foods and the pharmaceutical

  4. Potential use of molecular-typing methods for the identification and characterization of salmonella enterica serotypes isolated in the poultry production chain

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    CM Baratto

    2012-09-01

    Full Text Available Salmonella is widespread in nature and can be found in all links of the poultry production chain. Due to its high impact on meat processing, techniques for the rapid detection and reproducible characterization of Salmonella serotypes in foods are needed. The present study investigated the potential of molecular profiling to identify and differentiate 15 Salmonella serotypes isolated from the poultry production chain, based on 5 primers by random amplified polymorphic DNA (RAPD, enterobacterial repetitive intergenic consensus (ERIC-PCR, amplification of rDNA internal spacer analysis (RISA, and amplified ribosomal DNA restriction analysis (ARDRA of 16S-23S rRNA internal spacer region (ISR cleaved with Alu I and Hha I restriction enzymes. Three isolates of each serotype were analyzed for the identification of similar and different profiles. Dendrograms were constructed from molecular profiles using the UPGMA method (unweighted pair-group method for the arithmetic averages and the software program WinBoot. The present study indicates the usefulness of RISA and ARDRA of the 16S-23S rRNA intergenic spacer region (ISR for systematic, epidemiological, and diagnostic purposes. Since these techniques can be used for the differentiation of serotypes, they are highly promising for the characterization of Salmonella serotypes and intra-serotypes. Data indicate that these techniques may be used to produce more consistent, reliable, and reproducible results in the identification and epidemiological study (traceability of Salmonella in the poultry industry.

  5. ISOLATION AND MOLECULAR IDENTIFICATION OF POTENTIALLY PATHOGENIC Escherichia coli AND Campylobacter jejuni IN FERAL PIGEONS FROM AN URBAN AREA IN THE CITY OF LIMA, PERU

    Science.gov (United States)

    CABALLERO, Moisés; RIVERA, Isabel; JARA, Luis M.; ULLOA-STANOJLOVIC, Francisco M.; SHIVA, Carlos

    2015-01-01

    SUMMARY Feral pigeons (Columbia livia) live in close contact with humans and other animals. They can transmit potentially pathogenic and zoonotic agents. The objective of this study was to isolate and detect strains of diarrheagenic Escherichia coli and Campylobacter jejuni of urban feral pigeons from an area of Lima, Peru. Fresh dropping samples from urban parks were collected for microbiological isolation of E. coli strains in selective agar, and Campylobacter by filtration method. Molecular identification of diarrheagenic pathotypes of E.coli and Campylobacter jejuni was performed by PCR. Twenty-two parks were sampled and 16 colonies of Campylobacter spp. were isolated. The 100% of isolates were identified as Campylobacter jejuni. Furthermore, 102 colonies of E. coliwere isolated and the 5.88% resulted as Enteropathogenic (EPEC) type and 0.98% as Shiga toxin-producing E. coli (STEC). The urban feral pigeons of Lima in Peru can act as a reservoir or carriers of zoonotic potentially pathogenic enteric agents. PMID:26603225

  6. Isolation and Molecular Identification of Vermamoeba vermiformis Strains from Soil Sources in El Hierro Island, Canary Islands, Spain.

    Science.gov (United States)

    Reyes-Batlle, María; Wagner, Carolina; Zamora-Herrera, Jonadab; Vargas-Mesa, Alejandro; Sifaoui, Ines; González, Ana C; López-Arencibia, Atteneri; Valladares, Basilio; Martínez-Carretero, Enrique; Piñero, José E; Lorenzo-Morales, Jacob

    2016-07-01

    Free-living amoebae (FLA) are widely distributed protozoa in the environment and have been isolated from many sources such as dust, soil and water. Furthermore, some genera/species of FLA such as Naegleria fowleri, Balamuthia mandrillaris and Acanthamoeba spp. are also able to cause opportunistic infections in humans and other animals. More recently, FLA have been reported to be environmental carriers of pathogenic bacteria, fungi and viruses, and thus have gained further importance from the public health point of view. Among them, Acanthamoeba spp. and Vermamoeba vermiformis have been described in many occasions as the most common carriers of pathogens of high medical relevance such as Legionella pneumophila and Mycobacterium spp. In this study, 24 soil samples were collected from the island of El Hierro, Canary Islands, Spain, in order to check for the presence of V. vermiformis strains in these samples. Soil samples were cultured on 2 % non-nutrient agar plates covered with a thin layer of heat-killed E. coli and checked daily for the presence of Vermamoeba. After a week, V. vermiformis amoebae were observed in 5 of the 24 processed samples (20.8 %) incubated at room temperature and 37 °C. Molecular characterization was carried out by amplifying the 18S rDNA gene and DNA sequencing, confirming that the isolated strains belonged to Vermamoeba vermiformis species. The high percentage of V. vermiformis in the studied soil sources should raise awareness in the region since these amoebae are potential environmental carriers of pathogens of high medical relevance.

  7. Molecular identification of waterborne free living amoebae (Acanthamoeba, Naegleria and Vermamoeba) isolated from municipal drinking water and environmental sources, Semnan province, north half of Iran.

    Science.gov (United States)

    Javanmard, Ehsan; Niyyati, Maryam; Lorenzo-Morales, Jacob; Lasjerdi, Zohreh; Behniafar, Hamed; Mirjalali, Hamed

    2017-12-01

    The present study tested 80 samples of municipal, geothermal and recreational water samples for the occurrence of waterborne free living amoebae (FLA) including Acanthamoeba, Balamuthia mandrillaris, Vahlkampfiids and Vermamoeba in Semnan province, North half of Iran. Four sets of primers including JDP1,2 primers, ITS1,2 primers (Vahlkampfiids), 16S rRNABal primers (Balamuthia mandrillaris) and NA1,2 primers (Vermamoeba) were used to confirm the morphological identification. From the 80 water samples tested in the present study, 16 (20%) were positive for the outgrowth of free living amoebae based on the morphological page key. Out of the 34 municipal water samples, 7 (20.6%) were positive for outgrowth of Free living amoeba, belonging to Vermamoeba, Naegleria and Acanthamoeba using molecular tools. Three out of the six investigated hot springs were also contaminated with Naegleria spp. Sequencing of the ITS1,2 region of the Vahlkampfiid isolates revealed the highest homology with N. gruberi (2 isolates), N. australiensis (1 isolate) and N. pagei (3 isolates). This is the first report of N. gruberi in the country. Using morphological and molecular analysis, Balamuthia mandrillaris was undetected in all the water samples. The present study further confirmed the occurrence of potentially pathogenic waterborne free living amoebae in habitats with high human activity. It is of utmost importance that more studies are conducted to evaluate the niches of B. mandrillaris and N. fowleri in Iran and worldwide. Such investigations regarding the relevance of FLA as a hazard to humans, should be brought to the notice of the health authorities. Copyright © 2017. Published by Elsevier Inc.

  8. Isolation and molecular identification of Mycobacterium from commercially available pasteurized milk and raw milk samples collected from two infected cattle farms in Alborz Province, Iran.

    Science.gov (United States)

    Eftekhari, Mohsen; Mosavari, Nader

    2016-12-01

    Mycobacterium avium subsp. paratuberculosis (MAP) is an etiological agent of Johne's disease in ruminant including cattle, sheep and goats. This disease is considered an economically important disease in cattle. Animals with paratuberculosis shed viable MAP, particularly in their milk and feces. MAP may be involved in the development of Crohn's disease in humans through the consumption of contaminated milk and dairy products. Common methods of pasteurization are not enough to kill all MAP present in the milk and the bacterium has been isolated from raw milk, pasteurized milk and cheese samples. The purpose of this study was to evaluate two different methods for detecting MAP in milk and milk products. We analyzed the commonly used methods such as culture and molecular biology for identification of MAP. For this study, 50 milk samples from cows with suspected Johne's disease located in two dairy farms and 10 commercially available pasteurized milk and cheese samples from the market in Karaj city, Iran were selected. Following Ziehl-Neelsen staining of milk samples, direct microscopic detection of MAP was performed. All milk samples were centrifuged, and the concentrated samples were decontaminated using hexadecyl pyridinium chloride. The decontaminated milk suspensions were washed three times by centrifuging, and the collected filtrates were cultivated on Herrold's egg yolk medium enriched by Mycobactin J. Finally, identification and confirmation of isolates to MAP was performed using IS900-nested polymerase chain reaction (PCR). According to the obtained results by culture and PCR methods, none of the pasteurized milk and cheese samples showed the presence of MAP. However, 10% of the tested raw milk samples collected from suspected cattle showed the presence of MAP by both culture and PCR methods. Culture and PCR methods are reliable for identification of MAP from milk samples. Copyright © 2016.

  9. Identification and molecular characterization of antimicrobial-resistant shiga toxin-producing Escherichia coli isolated from retail meat products.

    Science.gov (United States)

    Li, Ming-Cheng; Wang, Fang; Li, Fan

    2011-04-01

    Ten (2.7%) Shiga toxin-producing Escherichia coli (STEC) were isolated from 370 samples of raw minced beef, mutton, pork, and chicken from the Jilin region of China; and additional 10 E. coli O157:H7 isolates were previously isolated from different Jilin regions. Seventeen of the isolates were multiresistant, exhibiting resistance to ampicillin, ciprofloxacin, tetracycline, sulfamethoxazole-trimethoprim, gentamycin, and streptomycin. Class 1 integrons were detected in nine (45.0%) of the STEC isolates and consisted of serogroups O157, O62, O113, O149, and O70. Integrons containing amplicons of a 0.5-1.5 or 1.0 kb gene cassette were found in seven (77.8%) of the integron-containing isolates. Sequencing analysis revealed that these gene cassettes encode genes conferring resistance to trimethoprim (dfrA1) and streptomycin (aadA1). The 0.5 kb cassette described here was found to encode a putative transporter peptide in the STEC. Seventeen isolates contained plasmids with different bands, and transfer by conjugation between strains of E. coli demonstrated that class 1 integrons located on mobile plasmids could contribute to the emergence and dissemination of antimicrobial resistance to ampicillin, gentamycin, streptomycin, and sulfamethoxazole-trimethoprim amongst STEC. These data revealed the high prevalence of antimicrobial-resistant STEC isolates in Jilin's surrounding regions, providing important and useful surveillance information reflecting antimicrobial selection pressure. © Mary Ann Liebert, Inc.

  10. Molecular genetic identification and metal biosorption by a Geobacillus genospecies IRKM1 isolated from Deeymand hot spring, Kerman, Iran

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    Asieh Dokhani

    2012-01-01

    Conclusions: The above results showed that the isolate was a member of Geobacillus spp. and the thermophilic bacteria was moderately resistant to Cu and Ni metals even though there was not any previous contamination of that biological niche. The organism exhibited highest biosorption of Cu 2+ at 65°C and Ni 2+ at 27°C. No plasmid was detected in the Geobacillus isolate.

  11. Identification and molecular characterization of serological group C streptococci isolated from diseased pigs and monkeys in Indonesia.

    Science.gov (United States)

    Soedarmanto, I; Pasaribu, F H; Wibawan, I W; Lämmler, C

    1996-01-01

    The present study was designed to comparatively investigate 34 beta-hemolytic streptococci isolated from infected pigs and monkeys from various islands in Indonesia. According to the serological and biochemical data, all 34 isolates were Lancefield's serological group C streptococci and could be identified as Streptococcus equi subsp. zooepidemicus. Of the 34 group C streptococci investigated, 28 grew on solid media in large, mucoid colonies, in fluid media at a uniform turbidity, and in soft agar in diffuse colonies. A decapsulation test with a hyaluronidase-producing Staphylococcus aureus strain revealed the hyaluronic acid nature of the capsular material. The remaining six streptococci grew on solid media in small, nonmucoid colonies, in fluid media as sediment with clear supernatant, and in soft agar in compact colonies. Determination of surface hydrophobicity by salt aggregation revealed a hydrophilic surface for the encapsulated bacteria and a hydrophobic surface for the unencapsulated group C streptococci. To further analyze the epidemiological relationships, all 34 mucoid and nonmucoid isolates from pigs and monkeys were subjected to protein and DNA fingerprinting. The latter was performed by pulsed-field gel electrophoresis. The protein profiles of all 34 isolates and the DNA profiles of 32 isolates appeared to be identical, with the DNA profiles of 2 isolates being closely related, indicating that a single virulent clone is responsible for this disease outbreak in Indonesia. PMID:8862585

  12. Characterization of Phosphate Solubilizing Bacteria in Sediments from a Shallow Eutrophic Lake and a Wetland: Isolation, Molecular Identification and Phosphorus Release Ability Determination

    Directory of Open Access Journals (Sweden)

    Jie Tang

    2010-11-01

    Full Text Available The transformation of phosphorus (P is a major factor of lake eutrophication, and phosphate releasing bacteria play an important role in the release process. Experiments were conducted to investigate P content and characterize phosphate solubilizing bacterial composition at the molecular level in a shallow eutrophic lake and a wetland. Results showed that P concentrations were relatively high and derived from agricultural runoff and domestic or industrial pollution. Enumeration and molecular identification of these strains indicated that these bacterial groups were abundant in the ecosystem and various kinds of bacteria participated in the phosphorus release process. Twelve phosphate solubilizing bacteria, including eight organic P-solubilizing bacteria (OPBs and four inorganic P-solubilizing bacteria (IPBs, which belonged to three different families, were isolated and identified. Cupriavidus basilensis was found for the first time to have the ability to mineralize organic P (OP. Laboratory tests on P release ability revealed that IPBs were more effective at releasing P than OPBs. The most efficient IPB strain could accumulate over 170 mg·L-1 orthophosphate, while the equivalent OPB strain only liberated less than 4 mg·L-1 orthophosphate in liquid culture. The results obtained from this investigation should help clarify the roles of microorganisms in aquatic systems and the mechanisms of eutrophication.

  13. Molecular identification of phosphate-solubilizing native bacteria isolated from the rhizosphere of Prosopis glandulosa in Mexicali valley.

    Science.gov (United States)

    Moreno-Ramírez, L; González-Mendoza, D; Cecena-Duran, C; Grimaldo-Juarez, O

    2015-03-31

    One of the main limitations in intensive crop production in Northwestern Mexico is the dependence on the use of phosphate fertilizer. In this study, we isolated indigenous microorganisms with phosphate solubilization capacities from mesquite (Prosopis glandulosa) present in the Mexicali valley. In total, 4 bacteria were isolated from the rhizosphere of mesquite, including ICA01, ICA02Ba, ICA03Bs, and ICA04Ma. The bacterial isolates were identified based on their phenotypic and 16S rRNA gene sequencing data to be Acinetobacter calcoaceticus. The results showed that ICA01 was the most efficient in solubilizing phosphate, followed by ICA02Ba and ICA03Bs, while ICA04Ma showed the lowest phosphate-solubilizing activity. The pH value of the culture medium decreased with bacterial growth, suggesting that these strains produce organic acids that solubilize phosphorus. These results will be useful for biotechnological studies and A. calcoaceticus may be employed for biofertilization programs in northwest Mexico.

  14. Ultrastructural characteristics and molecular identification of Entamoeba suis isolated from pigs with hemorrhagic colitis: implications for pathogenicity.

    Science.gov (United States)

    Matsubayashi, Makoto; Suzuta, Fumiko; Terayama, Yoshimi; Shimojo, Kengo; Yui, Takeshi; Haritani, Makoto; Shibahara, Tomoyuki

    2014-08-01

    Protozoan parasites of the genus Entamoeba infect many classes of vertebrates and are primarily classified based on morphological criteria. To date, only a few species have been proven to cause disease. Here, we examined the pathology of infected pigs with hemorrhage and detected Entamoeba parasites. Isolates were characterized genetically and ultrastructurally to identify the species. Histopathologically, bleeding and thrombus formation were seen only in the large intestine mucosa, where a large number of trophozoites or some Entamoeba cysts were observed around breakdowns in the lamina propria. No screw-shaped bacteria were detected in the lesions, and no pathogenic bacteria such as Brachyspira spp. were detected in fecal cultures. Interestingly, electron microscopy revealed that the parasites possessed mitochondrial organelles, unlike other Entamoeba spp. The isolates were identified as Entamoeba suis by PCR analysis and sequencing of the small subunit ribosomal RNA (SSU rRNA) gene. In phylogenetic analyses based on the actin gene, the E. suis isolate formed a cluster with Entamoeba histolytica and Entamoeba invadens, as well as with other parasites of the Amoebidae. Whether the pathogenicity of the E. suis isolate is affected by the severity of infection or host health status remains unclear; however, our results suggest that E. suis could cause or exacerbate clinical symptoms such as hemorrhagic colitis or diarrhea.

  15. Identification and molecular characterization of Listeria monocytogenes isolated in raw milk in the region of Algiers (Algeria).

    Science.gov (United States)

    Hamdi, Taha Mossadak; Naïm, Malek; Martin, Paul; Jacquet, Christine

    2007-05-01

    Listeria monocytogenes was isolated from raw milk, whey and curdled milk produced and collected in the region of Algiers and Blida between September 2003 and July 2004. Four out of 153 (2.61%) farm milk samples and 6 out of 80 (7.50%) tankers' samples tested positive for L. monocytogenes. All samples of whey and curdled milk (n=12) tested negative for L. monocytogenes, but 2 of 22 (9%) samples of whey were contaminated by L. innocua. L. monocytogenes isolates were grouped by a multiplex PCR assay; all isolates belonged to the PCR-group IVb, which corresponds to serovars 4b, 4d and 4e. L. monocytogenes isolates were characterized by Pulsed-Field Gel Electrophoresis (PFGE). The combination of AscI and ApaI macrorestriction patterns yielded five different pulsovars (I to V). The results indicate that raw milk, and raw milk products are potential sources of the L. monocytogenes and represent a potential risk for consumers.

  16. Evaluation of molecular markers for Phytophthora ramorum detection and identification: Testing for specificity using a standardized library of isolates

    Science.gov (United States)

    F.N. Martin; M.D. Coffey; K. Zeller; R.C. Hamelin; P. Tooley; M. Garbelotto; K.J.D. Hughes; T. Kubisiak; G.J. Bilodeau; L. Levy; C. Blomquist; P.H. Berger

    2009-01-01

    Given the importance of Phytophthora ramorum from a regulatory standpoint, it is imperative that molecular markers for pathogen detection are fully tested to evaluate their specificity in detection of the pathogen. In an effort to evaluate 11 reported diagnostic techniques, we assembled a standardized DNA library using accessions from the World...

  17. First report of Streptococcus agalactiae isolated from Oreochromis niloticus in Piura, Peru: Molecular identification and histopathological lesions

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    Yessica Ortega Asencios

    2016-11-01

    Full Text Available The aim of this study was to identify the bacterium Streptococcus agalactiae isolated in farmed Nile tilapia (Oreochromis niloticus from Piura, Peru and to characterize the histopathological lesions caused by this pathogen. Sixteen tilapias were sampled with clinic signs of the disease such as erratic swimming, exophthalmia and haemorrhages on the body and fins. Qualitative PCR in real time and histopathological analysis were performed. Nine fishes positives to S. agalactiae were found. The main histopathological findings were fibrinosuppurative epicarditis, periesplenitis, meninigitis and panophtaltmitis with predominance of mononuclear infiltration in all tissues. The correlation between qPCR and histopathological findings demonstrated nine fish (prevalence of 56.25% with Cq lower than 30, associated to high degree of tissue injuries. This study reports the first isolation of S. agalactiae by PCR in real time in tilapia farmed in Peru and characterizes the major histopathological changes caused by this bacterium.

  18. Occurrence, isolation and DNA identification of involved in Algerian ...

    African Journals Online (AJOL)

    A total of 12 strains were tested by DNA identification analysis and these indigenous isolates were unambiguously characterised by their housekeeping gene profiles. Finally, four genotypes were recognised. To our knowledge, this is the first report on the isolation and molecular characterization of S. thermophilus strains ...

  19. Isolation and molecular identification of landfill bacteria capable of growing on di-(2-ethylhexyl) phthalate and deteriorating PVC materials.

    Science.gov (United States)

    Latorre, Isomar; Hwang, Sangchul; Montalvo-Rodriguez, Rafael

    2012-01-01

    Waste materials containing Di-(2-ethylhexyl) phthalate (DEHP), a suspected endocrine disruptor and reasonably anticipated human carcinogen, are typically disposed of in landfills. Despite this, very few studies had been conducted to isolate and identify DEHP-degrading bacteria in landfill leachate. Therefore, this study was conducted to isolate and characterize bacteria in landfill leachate growing on DEHP as the sole carbon source and deteriorating PVC materials. Four strains LHM1, LHM2, LHM3 and LHM4, not previously reported as DEHP-degraders, were identified via 16S rRNA gene sequence. Gram-positive strains LHM1 and LHM2 had a greater than 97% similarity with Chryseomicrobium imtechense MW 10(T) and Lysinibacillus fusiformis NBRC 15717(T), respectively. Gram-negative strains LHM3 and LHM4 were related to Acinetobacter calcoaceticus DSM 30006(T) (90.7% similarity) and Stenotrophomonas pavanii ICB 89(T) (96.0% similarity), respectively. Phylogenetic analysis also corroborated these similarities of strains LHM1 and LHM2 to the corresponding bacteria species. Strains LHM2 and LHM4 grew faster than strains LHM1 and LHM3 in the enrichment where DEHP was the sole carbon source. When augmented to the reactors with PVC shower curtains containing DEHP, strains LHM1 and LHM2 developed greater optical densities in the solution phase and thicker biofilm on the surfaces of the shower curtains.

  20. Molecular identification and analysis of human enteroviruses isolated from healthy children in Shenzhen, China from 2010 to 2011.

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    Wei Wu

    Full Text Available OBJECTIVE: To determine the prevalence and distribution of human enteroviruses (HEVs among healthy children in Shenzhen, China. METHOD: Clinical specimens were obtained from 320 healthy children under 5 years old in Shenzhen, China from 2010 to 2011. The specimens were evaluated using real-time PCR and cell cultures. The positive specimens were further tested using reverse transcription-seminested PCR (RT-snPCR. Molecular typing and phylogenetic analysis were based on the sequence determined. RESULTS: Among the 320 samples, 34 were tested positive for HEVs (10.6% and 22 different serotypes were identified using RT-snPCR. PV1 and PV2 were also detected. The predominant serotype observed was EV71 (17.6%, followed by CV-B4 (14.7%. HEV-B was detected most frequently, with an overall prevalence of 47.1%. HEV-A and HEV-C were found in 32.3% and 20.6% of the samples, respectively. No HEV-D was identified. Molecular phylogeny indicated that all EV71 strains were of C4 genotype. CONCLUSION: Although a variety of HEVs was detected in healthy children, HEV-B was relatively more prevalent than other HEV species. Considering HEV-A is more prevalent than HEV-B among patients with hand-foot-mouth disease, additional long-term surveillance of HEV is warranted in both asymptomatic and symptomatic populations.

  1. Isolation and identification of Salmonella spp. in drinking water, streams, and swine wastewater by molecular techniques in Taiwan

    Science.gov (United States)

    Kuo, C.; Hsu, B.; Shen, T.; Tseng, S.; Tsai, J.; Huang, K.; Kao, P.; Chen, J.

    2013-12-01

    Salmonella spp. is a common water-borne pathogens and its genus comprises more than 2,500 serotypes. Major pathogenic genotypes which cause typhoid fever, enteritis and other intestinal-type diseases are S. Typhimurium, S. Enteritidis, S. Stanley, S. Agona, S.Albany, S. Schwarzengrund, S. Newport, S. Choleraesuis, and S. Derby. Hence, the identification of the serotypes of Salmonella spp. is important. In the present study, the analytical procedures include direct concentration method, non-selective pre-enrichment method and selective enrichment method of Salmonella spp.. Both selective enrichment method and cultured bacteria were detected with specific primers of Salmonella spp. by polymerase chain reaction (PCR). At last, the serotypes of Salmonella were confirmed by using MLST (multilocus sequence typing) with aroC, dnaN, hemD, hisD, purE, sucA, thrA housekeeping genes to identify the strains of positive samples. This study contains 121 samples from three different types of water sources including the drinking water (51), streams (45), and swine wastewater (25). Thirteen samples with positive invA gene are separated from culture method. The strains of these positive samples which identified from MLST method are S. Albany, S. Typhimurium, S. Newport, S. Bareilly, and S. Derby. Some of the serotypes, S. Albany, S. Typhimurium and S. Newport, are highly pathogenic which correlated to human diarrhea. In our results, MLST is a useful method to identify the strains of Salmonella spp.. Keywords: Salmonella, PCR, MLST.

  2. Identification and molecular characterization of a new nonsegmented double-stranded RNA virus isolated from Culex mosquitoes in Japan.

    Science.gov (United States)

    Isawa, Haruhiko; Kuwata, Ryusei; Hoshino, Keita; Tsuda, Yoshio; Sakai, Kouji; Watanabe, Shumpei; Nishimura, Miho; Satho, Tomomitsu; Kataoka, Michiyo; Nagata, Noriyo; Hasegawa, Hideki; Bando, Hisanori; Yano, Kazuhiko; Sasaki, Toshinori; Kobayashi, Mutsuo; Mizutani, Tetsuya; Sawabe, Kyoko

    2011-01-01

    Two infectious agents were isolated from Culex species mosquitoes in Japan and were identified as distinct strains of a new RNA virus by a method for sequence-independent amplification of viral nucleic acids. The virus designated Omono River virus (OMRV) replicated in mosquito cells in which it produced a severe cytopathic effect. Icosahedral virus particles of approximately 40 nm in diameter were detected in the cytoplasm of infected cells. The OMRV genome was observed to consist of a nonsegmented, 7.6-kb double-stranded RNA (dsRNA) and contain two overlapping open reading frames (ORFs), namely ORF1 and ORF2. ORF1 was found to encode a putative dsRNA-binding protein, a major capsid protein, and other putative proteins, which might be generated by co- and/or post-translational processing of the ORF1 polyprotein precursor, and ORF2 was found to encode a putative RNA-dependent RNA polymerase (RdRp), which could be translated as a fusion with the ORF1 product by a -1 ribosomal frameshift. Phylogenetic analysis based on RdRp revealed that OMRV is closely related to penaeid shrimp infectious myonecrosis virus and Drosophila totivirus, which are tentatively assigned to the family Totiviridae. These results indicated that OMRV is a new member of the family of nonsegmented dsRNA viruses infecting arthropod hosts, but not fungal or protozoan hosts. Copyright © 2010 Elsevier B.V. All rights reserved.

  3. Molecular identification of aminoglycoside-modifying enzymes in clinical isolates of Escherichia coli resistant to amoxicillin/clavulanic acid isolated in Spain.

    Science.gov (United States)

    Fernández-Martínez, Marta; Miró, Elisenda; Ortega, Adriana; Bou, Germán; González-López, Juan José; Oliver, Antonio; Pascual, Alvaro; Cercenado, Emilia; Oteo, Jesús; Martínez-Martínez, Luis; Navarro, Ferran

    2015-08-01

    The activity of eight aminoglycosides (amikacin, apramycin, arbekacin, gentamicin, kanamycin, neomycin, netilmicin and tobramycin) against a collection of 257 amoxicillin/clavulanic acid (AMC)-resistant Escherichia coli isolates was determined by microdilution. Aminoglycoside resistance rates, the prevalence of aminoglycoside-modifying enzyme (AME) genes, the relationship between AME gene detection and resistance phenotype to aminoglycosides, and the association of AME genes with mechanisms of AMC resistance in E. coli isolates in Spain were investigated. Aminoglycoside-resistant isolates were screened for the presence of genes encoding common AMEs [aac(3)-Ia, aac(3)-IIa, aac(3)-IVa, aac(6')-Ib, ant(2″)-Ia, ant(4')-IIa and aph(3')-Ia] or 16S rRNA methylases (armA, rmtB, rmtC and npmA). In total, 105 isolates (40.9%) were resistant to at least one of the aminoglycosides tested. Amikacin, apramycin and arbekacin showed better activity, with MIC90 values of 2mg/L (arbekacin) and 8mg/L (amikacin and apramycin). Kanamycin presented the highest MIC90 (128mg/L). The most common AME gene was aac(6')-Ib (36 strains; 34.3%), followed by aph(3')-Ia (31 strains; 29.5%), ant(2″)-Ia (29 strains; 27.6%) and aac(3)-IIa (23 strains; 21.9%). aac(3)-Ia, aac(3)-IVa, ant(4')-IIa and the four methylases were not detected. The ant(2″)-Ia gene was usually associated with OXA-1 [21/30; 70%], whilst 23/25 (92%) strains producing CTX-M-15 had the aac(6')-Ib gene. The most prevalent AME gene was aac(6')-Ib (18/41; 44%) in nosocomial isolates, whilst ant(2″)-Ia and aph(3')-Ia genes (20/64; 31%) were more frequent in strains of community origin. In 64.6% isolates the phenotypic profile correlated with the presence of commonly encountered AMEs. Copyright © 2015 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved.

  4. Visual Analysis of DNA Microarray Data for Accurate Molecular Identification of Non-albicans Candida Isolates from Patients with Candidemia Episodes

    OpenAIRE

    De Luca Ferrari, Michela; Ribeiro Resende, Mariângela; Sakai, Kanae; Muraosa, Yasunori; Lyra, Luzia; Gonoi, Tohru; Mikami, Yuzuru; Tominaga, Kenichiro; Kamei, Katsuhiko; Zaninelli Schreiber, Angelica; Trabasso, Plinio; Moretti, Maria Luiza

    2013-01-01

    The performance of a visual slide-based DNA microarray for the identification of non-albicans Candida spp. was evaluated. Among 167 isolates that had previously been identified by Vitek 2, the agreement between DNA microarray and sequencing results was 97.6%. This DNA microarray platform showed excellent performance.

  5. Coagulase-positive Staphylococcus isolated from wildlife: Identification, molecular characterization and evaluation of resistance profiles with focus on a methicillin-resistant strain.

    Science.gov (United States)

    Nowakiewicz, Aneta; Ziółkowska, Grażyna; Zięba, Przemysław; Gnat, Sebastian; Wojtanowicz-Markiewicz, Katarzyna; Trościańczyk, Aleksandra

    2016-02-01

    The aim of the study was molecular analysis of coagulase-positive isolates of Staphylococcus bacteria obtained from wild animals and evaluation of their resistance to antimicrobial agents. A total of 76 rectal swabs were taken from wild animals. The species of the Staphylococcus isolates was determined by MALDI TOF MS, susceptibility to antimicrobials was evaluated by phenotypic and molecular methods, epidemiological analysis (ADSRRS-fingerprinting) was also carried out. MRSA isolate was typed by MLST and spa-typing. The animals tested, were carriers (n=38) of coagulase-positive Staphylococcus (S. aureus, S. pseudintermedius and S. delphini B). Analyzed isolates were resistant to 1 or 2 antimicrobials, which was confirmed by the presence of genes (blaZ, ermA, ermB, msrA, tetK and tetM). A multi-drug resistant and methicillin-resistant isolate of S. aureus was obtained as well (MRSA, ST8, t1635, PVL-positive and ACME-negative). The ADSRRS-fingerprinting method enabled interspecific and intraspecific differentiation of coagulase-positive Staphylococcus isolates, revealing a certain degree of correlation between the species of the isolate, and the degree of similarity between the isolates. The presence of resistance genes in 13% (5/38) of the isolates obtained from wild animals, including one methicillin-resistant isolate, is relatively small in comparison to the degree of colonization by resistant strains in humans, livestock or pets. Nevertheless, due to the possibility of contact between wild animals, domestic animals and humans, transmission of resistant strains is possible, as suggested by our isolation of a MRSA strain typed as ST8 and specific spa type t1635, which had previously been isolated exclusively from humans. Copyright © 2015 Elsevier Ltd. All rights reserved.

  6. Molecular identification of Candida species isolated from cases of neonatal candidemia using polymerase chain reaction-restriction fragment length polymorphism in a tertiary care hospital

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    Akeela Fatima

    2017-01-01

    Full Text Available Context: Candida spp. is an emerging cause of bloodstream infections worldwide. Delay in speciation of Candida isolates by conventional methods and resistance to antifungal drugs in various Candida species are responsible for the increase in morbidity and mortality due to candidemia. Hence, the rapid identification of Candida isolates is very important for the proper management of patients with candidemia. Aims: The aim was to re-evaluate the identification of various Candida spp. by polymerase chain reaction (PCR-restriction fragment length polymorphism (RFLP and to evaluate the accuracy, speed, and cost of phenotypic methodology versus PCR-RFLP. Settings and Design: Hospital-based cross-sectional study. Materials and Methods: Ninety consecutive clinical isolates of seven Candida species, isolated from blood of neonates and identified by routine phenotypic methods, were re-evaluated using universal primers internal transcribed spacer 1 (ITS1 and ITS4 for PCR amplification and Msp I restriction enzyme for RFLP. Statistical Analysis Used: Kappa test for agreement. Results: The results of PCR-RFLP were 100% in agreement with those obtained using conventional phenotypic methods. Identification could be achieved within 3 work days by both the methods. Our routine methods proved to be cost effective than PCR-RFLP. Conclusions: We can continue with our routine phenotypic methods and PCR-RFLP can be used for periodic quality control or when conventional methods fail to identify a species.

  7. Molecular identification of Aspergillus spp. isolated from coffee beans Identificação molecular de Aspergillus spp. isolados de grãos de café

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    Marciane Magnani

    2005-01-01

    Full Text Available Some species belonging to the genus Aspergillus are potential producers of ochratoxin A (OA, a mycotoxin with nephrotoxic, immunosuppressive, teratogenic and carcinogenic effects. The aim of the present study was to identify the species of Aspergillus that contaminate the inside of coffee beans collected in the stage of maturation and drying, from 16 producing areas located in the northern region of the State of Paraná, in the South of Brazil. A total of 108 isolates of Aspergillus spp. was identified at the species level, by sequencing the internal transcribed spacer (ITS1-5.8S-ITS2 of ribosomal DNA (rDNA. The results revealed the presence of potentially ochratoxigenic species in 82% of the geographic regions studied, among which Aspergillus niger was the species most frequently detected, followed by A. ochraceus and A. carbonarius. The presence of A. carbonarius in immature coffee fruits harvested from trees is reported for the first time.Algumas espécies pertencentes ao gênero Aspergillus possuem potencial para produção de Ocratoxina A (OA, uma micotoxina de efeitos nefrotóxicos, imunossupressivos, teratogênicos e carcinogênicos. Com o objetivo de identificar as espécies de Aspergillus que contaminam o interior de grãos de café, foram coletadas amostras em diferentes estádios de maturação do produto, em 16 propriedades produtoras do norte do estado do Paraná. Um total de 108 isolados de Aspergillus spp. foram identificados ao nível de espécie, pelo sequenciamento dos espaços internos transcritos (ITS1-5,8S-ITS2 do DNA ribossomal (rDNA. Os resultados revelaram a presença de espécies potencialmente ocratoxigênicas em 82% das regiões analisadas, sendo dentre estas, Aspergillus niger a espécie mais freqüentemente detectada,seguida por A. ochraceus, e A. carbonarius. É relatada pela primeira vez a presença de A. carbonarius em frutos de café coletados na árvore.

  8. Isolation and molecular identification of Vibrio spp. by sequencing of 16S rDNA from seafood, meat and meat products in Libya

    Directory of Open Access Journals (Sweden)

    S.M. Azwai

    2016-03-01

    Full Text Available The genus Vibrio includes several food-borne pathogens that cause a spectrum of clinical conditions including septicemia, cholera and milder forms of gastroenteritis. Several Vibrio spp. are commonly associated with food-borne transmission including Vibrio cholerae, Vibrio parahemolyticus, and Vibrio vulnificus. Microbiological analysis for enumeration and isolation of Vibrio spp. were carried out for a total of 93 samples of seafood, meat and meat products from different geographic localities in Libya (Tripoli, Regdalin, Janzour and Tobruk. Vibrio spp. were detected by conventional cultural and molecular method using PCR and sequencing of 16S rDNA. Out of the 93 cultured samples only 48 (51.6% yielded colonies on Thiosulfate Citrate Bile Salt agar (TCBS with culture characteristics of Vibrio spp. More than half (n=27 of processed seafood samples (n=46 yielded colonies on TCBS, while only 44.6% of samples of meat and meat products showed colonies on TCBS. Among cultured seafood samples, the highest bacterial count was recorded in clam with a count of 3.8 х104 CFU\\g. Chicken burger samples showed the highest bacterial count with 6.5 х104 CFU\\g. Molecular analysis of the isolates obtained in this study, showed that 11 samples out of 48 (22.9% were Vibrio spp. Vibrio parahemolyticus was isolated from camel meat for the first time. This study is an initial step to provide a baseline for future molecular research targeting Vibrio spp. foodborne illnesses. This data will be used to provide information on the magnitude of such pathogens in Libyan seafood, meat and meat products.

  9. Isolation and molecular identification of Vibrio spp. by sequencing of 16S rDNA from seafood, meat and meat products in Libya.

    Science.gov (United States)

    Azwai, S M; Alfallani, E A; Abolghait, S K; Garbaj, A M; Naas, H T; Moawad, A A; Gammoudi, F T; Rayes, H M; Barbieri, I; Eldaghayes, I M

    2016-01-01

    The genus Vibrio includes several food-borne pathogens that cause a spectrum of clinical conditions including septicemia, cholera and milder forms of gastroenteritis. Several Vibrio spp. are commonly associated with food-borne transmission including Vibrio cholerae, Vibrio parahemolyticus, and Vibrio vulnificus. Microbiological analysis for enumeration and isolation of Vibrio spp. were carried out for a total of 93 samples of seafood, meat and meat products from different geographic localities in Libya (Tripoli, Regdalin, Janzour and Tobruk). Vibrio spp. were detected by conventional cultural and molecular method using PCR and sequencing of 16S rDNA. Out of the 93 cultured samples only 48 (51.6%) yielded colonies on Thiosulfate Citrate Bile Salt agar (TCBS) with culture characteristics of Vibrio spp. More than half (n=27) of processed seafood samples (n=46) yielded colonies on TCBS, while only 44.6 % of samples of meat and meat products showed colonies on TCBS. Among cultured seafood samples, the highest bacterial count was recorded in clam with a count of 3.8 ×10(4) CFU\\g. Chicken burger samples showed the highest bacterial count with 6.5 ×10(4) CFU\\g. Molecular analysis of the isolates obtained in this study, showed that 11 samples out of 48 (22.9%) were Vibrio spp. Vibrio parahemolyticus was isolated from camel meat for the first time. This study is an initial step to provide a baseline for future molecular research targeting Vibrio spp. foodborne illnesses. This data will be used to provide information on the magnitude of such pathogens in Libyan seafood, meat and meat products.

  10. Molecular identification and physiological characterization of yeasts, lactic acid bacteria and acetic acid bacteria isolated from heap and box cocoa bean fermentations in West Africa.

    Science.gov (United States)

    Visintin, Simonetta; Alessandria, Valentina; Valente, Antonio; Dolci, Paola; Cocolin, Luca

    2016-01-04

    Yeast, lactic acid bacteria (LAB) and acetic acid bacteria (AAB) populations, isolated from cocoa bean heap and box fermentations in West Africa, have been investigated. The fermentation dynamicswere determined by viable counts, and 106 yeasts, 105 LAB and 82 AAB isolateswere identified by means of rep-PCR grouping and sequencing of the rRNA genes. During the box fermentations, the most abundant species were Saccharomyces cerevisiae, Candida ethanolica, Lactobacillus fermentum, Lactobacillus plantarum, Acetobacter pasteurianus and Acetobacter syzygii, while S. cerevisiae, Schizosaccharomyces pombe, Hanseniaspora guilliermondii, Pichia manshurica, C. ethanolica, Hanseniaspora uvarum, Lb. fermentum, Lb. plantarum, A. pasteurianus and Acetobacter lovaniensis were identified in the heap fermentations. Furthermore, the most abundant species were molecularly characterized by analyzing the rep-PCR profiles. Strains grouped according to the type of fermentations and their progression during the transformation process were also highlighted. The yeast, LAB and AAB isolates were physiologically characterized to determine their ability to grow at different temperatures, as well as at different pH, and ethanol concentrations, tolerance to osmotic stress, and lactic acid and acetic acid inhibition. Temperatures of 45 °C, a pH of 2.5 to 3.5, 12% (v/v) ethanol and high concentrations of lactic and acetic acid have a significant influence on the growth of yeasts, LAB and AAB. Finally, the yeastswere screened for enzymatic activity, and the S. cerevisiae, H. guilliermondii, H. uvarumand C. ethanolica species were shown to possess several enzymes that may impact the quality of the final product.

  11. Isolation, Characterization And Identification Of Enterobacteriaceae ...

    African Journals Online (AJOL)

    Isolation, Characterization And Identification Of Enterobacteriaceae From Well Water In Juja Town In Kenya. ... Of Enterobacteriaceae From Well Water In Juja Town In Kenya. RK Mwirichia, NM Budambulu, HI Boga ... All the water samples had total coliform counts >1100 per 100 ml of water. The Enterobacteriaceae ...

  12. Identification and differentiation of isolates of Colletotrichum ...

    African Journals Online (AJOL)

    Identification and differentiation of isolates of Colletotrichum gloeosporiodes from yam by random amplified polymorphic DNA markers. G Thottappilly, H D Mignouna, A Onasanya, M Abang, O Oyelakin, N K Singh. Abstract. (African Crop Science Journal: 1999 7(2): 195-206). AJOL African Journals Online. HOW TO USE ...

  13. Isolation, identification and characterization of Leuconostoc ...

    African Journals Online (AJOL)

    Isolation, identification and characterization of Leuconostoc mesenteroides as a new probiotic from intestine of snakehead fish ( Channa striatus ) ... This strain was able to survive and grow from pH 3 to 8 with the highest viability and growth rate at neutral conditions (pH 7). In addition, L. mesenteroides tolerated 0, 0.15 and ...

  14. Phenotypic and Molecular Identification of Bacteria Involved in Decubitus Ulcers

    Directory of Open Access Journals (Sweden)

    Mehran Dolati

    2017-07-01

    Full Text Available Background:    Bacterial secondary infection of pressure ulcers (bedsores, so called as decubitus ulcers, leads to ulcer development and it interferes with the healing process. Thus, such infections can be lethal due to the sepsis if no constructive medicinal measures regarded. Drug resistance of bacteria in pressure ulcers leads to healing inhibition. Molecular identification of bacteria involved in such infections seem necessary as culture and phenotypic approaches may result in misidentification. . The purpose of this study was to isolate and identify aerobic bacteria detected in bedsores in three Hospitals: Rasool-e-Akram, Imam Hossein and Tajrish Shohada Hospitals, Tehran, Iran.Methods:    To this end, decubitus ulcer samples of 49 patients were obtained using sterile swabs. After direct microscopic examination, the swabs were used to streak BHI agar plates supplemented with %5 defibrinated sheep blood for enrichment of probable aerobic cultures. Bacterial isolates diagnosed by biochemical tests. Antibiotic susceptibility of the isolates determined based on CLSI guideline. For molecular identification, PCR amplification of the 16S rRNA gene performed using Eubacterial universal primers. Then, the PCR products were sequenced and the nucleotide sequences of the PCR products were analyzed by BLASTN similarity search program available at NCBI. Results:   Among the isolates, Pseudomonas aeruginosa (36% had the highest frequency, followed by Staphylococcus aureus (32% and Escherichia coli (30%. The frequencies of Klebsiella pneumonia and Proteus spp. were 10% and 8%, respectively. Most of the isolated bacteria showed a widespread antibiotic resistance. Molecular identification of the bacterial isolates resulted in 6 isolates of Escherichia coli, two isolates of each of Proteus mirabilis and Shigella spp., 4 isolates of Enterobacter cloacae, and 1 isolate of each of Cronobacter sakazakii and Morganella morganii.Conclusion:

  15. Study of bacteria Gluconobacter sp.: isolation, purification, phenotypic and molecular identification Estudo de bactérias do gênero Gluconobacter: isolamento, purificação, identificação fenotípica e molecular

    Directory of Open Access Journals (Sweden)

    André Luiz Dorini de Oliveira

    2010-03-01

    Full Text Available The importance of the study of acetic bacteria, on species of the Gluconobacter genus is based on its industrial application, as these possess the capacity of bioconversion of sorbitol to sorbose, enabling the process of vitamin C production. The study involved samples collected in industries of soft drinks, flowers, fruits and honey, followed by purification, phenotypic identification, molecular identification with the use of primer defined from Nucleotide Sequence Database consultation. Strains preserved were identified as members of the Acetobacteraceae family, Gluconobacter genus. 110 strains had been isolated of substrate: Pyrostegia venusta (ker-gawler, honey, Vitis vinifera (grape, Pyrus communis (pear, Malus sp. (apple and in two samples of soft drinks. Of this total 57 strains had been recovered in manitol medium (manitol, yeast extract, peptone, 12 in YMG medium (glucose, manitol, yeast extract, ethanol, acetic acid, 41 in enrichment medium (De Ley and Swings and later in the GYC medium (glucose, yeast extract and calcium carbonate. 68 strains were identified as Gram negative bacilli rods. Of these, 31 were characterized biochemically as belonging to the Acetobacteriaceae family as they were catalase positive, oxidase negative and producers of acid from glucose. The characterization of these strains was complemented with the biochemistry tests: gelatin liquefaction, nitrate reduction, indole and H2S production, oxidation of ethanol to acetic acid and molecular tests for genus identification. Only eight strains were characterized as pertaining to the Gluconobacter genus. The strains are maintained in collection cultures at the Microbiology Laboratory of the Biology Department at the São Paulo State University (UNESP in Assis, stored in malt extract at -196 ºC.A importância do estudo de bactérias acéticas, em especial as do gênero Gluconobacter, está baseada em suas aplicações industriais, pois estas possuem a capacidade de

  16. Isolation and identification of Pseudomonas azotoformans for induced calcite precipitation.

    Science.gov (United States)

    Heidari Nonakaran, Siamak; Pazhouhandeh, Maghsoud; Keyvani, Abdullah; Abdollahipour, Fatemeh Zahra; Shirzad, Akbar

    2015-12-01

    Biomineralization is a process by which living organisms produce minerals. The extracellular production of these biominerals by microbes has potential for various bioengineering applications. For example, crack remediation and improvement of durability of concrete is an important goal for engineers and biomineral-producing microbes could be a useful tool in achieving this goal. Here we report the isolation, biochemical characterization and molecular identification of Pseudomonas azotoformans, a microbe that produces calcite and which potentially be used to repair cracks in concrete structures. Initially, 38 bacterial isolates were isolated from soil and cements. As a first test, the isolates were screened using a urease assay followed by biochemical tests for the rate of urea hydrolysis, calcite production and the insolubility of calcite. Molecular amplification and sequencing of a 16S rRNA fragment of selected isolates permitted us to identify P. azotoformans as a good candidate for preparation of biotechnological concrete. This species was isolated from soil and the results show that among the tested isolates it had the highest rate of urea hydrolysis, produced the highest amount of calcite, which, furthermore was the most adhesive and insoluble. This species is thus of interest as an agent with the potential ability to repair cracks in concrete.

  17. Molecular identification of Fusarium graminearum, sorghum pathogen in Serbia

    Directory of Open Access Journals (Sweden)

    Ristić Danijela

    2011-01-01

    Full Text Available A total of 39 samples of sorghum (Sorghum bicolor with symptoms of stem and root rot were collected and analyzed during 2009-2011 in Bački Petrovac and Čantavir, Serbia. Monosporic cultures were isolated from stem tissue, their pathogenicity was confirmed by the development of symptoms on artificially inoculated sorghum plants, and they were identified on the basis of macroscopic and microscopic morphological features as Fusarium graminearum. Molecular identification was performed utilizing polimerase chain reaction (PCR with primer pair ef1/ ef2 and by amplification of protein coding TEF 1-alpha gen. Sequence of TEF gene from the selected isolate 535- 10 (JF747146 showed 98-99% nucleotide identity with sequences of 63 Gibberella zeae isolates deposited in NCBI GenBank. Amplification of the barcoding region of F. graminearum genome of sorghum isolate, contributes to the fast and accurate identification and characterization of Fusarium species in Serbia.

  18. 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.

  19. Identificação molecular e suscetibilidade antifúngica de Candida parapsilosis isoladas no Ceará, Brasil Molecular identification and antifungal susceptibility of Candida parapsilosis isolates in Ceará, Brazil

    Directory of Open Access Journals (Sweden)

    Everardo Albuquerque Menezes

    2012-12-01

    Full Text Available INTRODUÇÃO: Candida parapsilosis é a segunda ou terceira levedura mais isolada de hemoculturas em várias partes do mundo. É um patógeno comumente isolado no Brasil e no Ceará. Apresenta capacidade de formar biofilmes em cateteres e outros dispositivos médicos e esses fatores contribuem para a disseminação dessa levedura. OBJETIVOS: Identificar e avaliar a suscetibilidade aos antifúngicos de C. parapsilosis isoladas de amostras de sangue e urina de pacientes atendidos em hospitais no Ceará. MÉTODOS: Foram isoladas e identificadas 57 cepas de C. parapsilosis. As cepas foram identificadas por testes fenotípicos e moleculares. A suscetibilidade foi avaliada pelo método de microdiluição em caldo (protocolo do Clinical and Laboratory Standards Institute [CLSI] M27-A3. Foram avaliados cinco antifúngicos (anfotericina B, caspofungina, fluconazol, itraconazol e voriconazol. RESULTADOS E CONCLUSÃO: As cepas foram identificadas como C. parapsilosis por testes fenotípicos e confirmadas pelos testes moleculares. Quanto ao perfil de sensibilidade, elas se mostraram sensíveis aos antifúngicos testados, sendo a resistência ainda um fenômeno raro entre cepas de C. parapsilosis isoladas no Ceará.INTRODUCTION: C. parapsilosis is the second or third most isolated yeast from blood cultures in various parts of the world. It is a commonly isolated pathogen in Brazil and Ceará. C. parapsilosis is liable to form biofilms on catheters and other medical devices, which contributes to the spread of this yeast. OBJECTIVE: The objective of this study was to identify and assess the antifungal susceptibility of C. parapsilosis isolates from blood and urine samples collected from patients in hospitals in Ceará. METHODS: We isolated and identified 57 strains of C. parapsilosis. The strains were identified by phenotypic and molecular tests. The susceptibility to antifungals was assessed by broth microdilution (Clinical and Laboratory Standards Institute

  20. Molecular Identification of Methicillin-Resistant Staphylococcus ...

    African Journals Online (AJOL)

    We use the molecular techniques of PCR and PFGE to identify MRSA from clinical isolates of Staphylococcus aureus causing infections among hospitalized patients in Benin-City, Nigeria. A total of 36 isolates were obtained from the University of Benin Teaching Hospital between July-September, 2007. The MRSA strains ...

  1. Molecular identification and characterization of the intervening sequences (IVSs) within 23S ribosomal RNA (rRNA) genes of Taylorella asinigenitalis isolated in France.

    Science.gov (United States)

    Tazumi, Akihiro; Petry, Sandrine; Hayashi, Kyohei; Moore, John E; Millar, Beverley C; Matsuda, Motoo

    2012-02-01

    In the helix 25 region, 32 French Taylorella asinigenitalis isolates carried at least one 23S rRNA gene not containing intervening sequences (IVSs). No IVSs in the region were identified in three isolates and the other remaining 29 isolates carried one or more IVSs (UCD-1(T)IVS1A, UCD-1(T)IVS1B and UK-1IVS1B) described already and two new kinds of IVS (TaIVS1C and TaIVS1D). In the helix 45 region, no T. asinigenitalis isolates not carrying any IVSs were identified. UK-1IVS2B was identified in the region from 26 isolates. Five new kinds of IVSs (TaIVS2D, E, F, G and H) occurred in the region in the 13 isolates. Distinctly different tandem repeat units (RS48 and RS32 and RS-A, -B and -C) were evident in both regions, respectively, from the French (n=32) and American (n=3) T. asinigenitalis isolates. Thus, several different kinds of tandem repeat units and their combinations in IVSs in both regions within the gene were shown in 32 French isolates. Copyright © 2010 Elsevier Ltd. All rights reserved.

  2. Molecular characterization of antimicrobial susceptibility of Salmonella isolates: First identification of a plasmid carrying qnrD or oqxAB in Taiwan

    Directory of Open Access Journals (Sweden)

    Cheng-Yen Kao

    2017-04-01

    Conclusion: GyrA mutations are the major mechanisms associated with quinolone-resistant Salmonella isolates in Taiwan. Overproduction of efflux pump genes and the presence of qnr and oqxAB play additional roles in reduced susceptibility to quinolones.

  3. Isolation and molecular characterization of phytase producing ...

    African Journals Online (AJOL)

    Isolation and molecular characterization of phytase producing bacteria from Malaysia hot springs. ... The strains were further analyzed in broth culture using sodium phytate as substrate. Among them, strain L3 was selected as the best producer (0.16 U/ml after 72 h of culture). This phytase showed optimal activity at 37 °C ...

  4. enumeration, isolation and identification of bacteria and fungi from ...

    African Journals Online (AJOL)

    userpc

    bioremediation of soil contaminated with petroleum products and possibly other oil polluted sites. Key words: Bioremediation .... bacterial isolates obtained from soil contaminated and layers chicken droppings. S/No Isolates identification. Grams. Reaction. Citrate Indole TSIA. Gas production. Urease Isolate identification. 1.

  5. Molecular typing of Chinese Streptococcus pyogenes isolates.

    Science.gov (United States)

    You, Yuanhai; Wang, Haibin; Bi, Zhenwang; Walker, Mark; Peng, Xianhui; Hu, Bin; Zhou, Haijian; Song, Yanyan; Tao, Xiaoxia; Kou, Zengqiang; Meng, Fanliang; Zhang, Menghan; Bi, Zhenqiang; Luo, Fengji; Zhang, Jianzhong

    2015-06-01

    Streptococcus pyogenes causes human infections ranging from mild pharyngitis and impetigo to serious diseases including necrotizing fasciitis and streptococcal toxic shock syndrome. The objective of this study was to compare molecular emm typing and pulsed field gel electrophoresis (PFGE) with multiple-locus variable-number tandem-repeat analysis (MLVA) for genotyping of Chinese S. pyogenes isolates. Molecular emm typing and PFGE were performed using standard protocols. Seven variable number tandem repeat (VNTR) loci reported in a previous study were used to genotype 169 S. pyogenes geographically-diverse isolates from China isolated from a variety of disease syndromes. Multiple-locus variable-number tandem-repeat analysis provided greater discrimination between isolates when compared to emm typing and PFGE. Removal of a single VNTR locus (Spy2) reduced the sensitivity by only 0.7%, which suggests that Spy2 was not informative for the isolates screened. The results presented support the use of MLVA as a powerful epidemiological tool for genotyping S. pyogenes clinical isolates. Copyright © 2015 Elsevier Ltd. All rights reserved.

  6. Molecular DNA Analysis in Forensic Identification.

    Science.gov (United States)

    Dumache, Raluca; Ciocan, Veronica; Muresan, Camelia; Enache, Alexandra

    2016-01-01

    Serological and biochemical identification methods used in forensics have several major disadvantages, such as: long time in processing biological sample and lack of sensitivity and specificity. In the last 30 years, DNA molecular analysis has become an important tool in forensic investigations. DNA profiling is based on the short tandem repeats (STR) and aids in human identification from biological samples. Forensic genetics, can provide information on the events which occurred at the crime scene or to supplement other methods of forensic identification. Currently, the methods used in identification are based on polymerase chain reaction (PCR) analyses. This method analyses the autosomal STRs, the Y-chromosome, and the mitochondrial DNA. Correlation of biological samples present at the crime scene with identification, selection, and the probative value factor is therefore the first aspect to be taken into consideration in the forensic genetic analysis. In the last decade, because of the advances in the field of molecular biology, new biomarkers such as: microRNAs (miR), messenger RNA (mRNA), and DNA methylation have been studied and proposed to be used in the forensic identifications of body fluids.

  7. Isolation and molecular identification of lactic acid bacteria and Bifidobacterium spp. from faeces of the blue-fronted Amazon parrot in Brazil.

    Science.gov (United States)

    Allegretti, L; Revolledo, L; Astolfi-Ferreira, C S; Chacón, J L; Martins, L M; Seixas, G H F; Ferreira, A J P

    2014-12-01

    In Brazil, the blue-fronted Amazon parrot (Amazona aestiva) is a common pet. The faecal microbiota of these birds include a wide variety of bacterial species, the majority of which belong to the Gram-positive lactic acid bacteria (LAB) clade. The aim of this study was to investigate differences in the diversity and abundance of LAB and Bifidobacterium spp. in the cloacae between wild and captive birds and to select, identify and characterise LAB for consideration as a parrot probiotic. Cloacal swabs were collected from 26 wild and 26 captive birds. Bacterial DNA was extracted, and the 16S rRNA genes were amplified. The numbers of PCR-positive Enterococcus, Pediococcus, and Lactobacillus species isolated from wild and captive birds were significantly different (P<0.05). Enterococcus was the most frequently isolated genus, followed by Pediococcus, Lactobacillus, Lactococcus and Bifidobacterium. Enterococcus faecium, Pediococcus pentosaceus, Lactococcus lactis, Lactobacillus coryniformis, Lactobacillus sanfranciscensis and Bifidobacterium bifidum were the most frequently isolated species from all birds. This study increases our understanding of the faecal microbiota, and may help to improve the nutrition and habitat management of captive and wild parrots. The bacterial population identified in the faecal microbiota of clinically healthy wild and captive parrots can serve as a database to analyse variations in the gut microbiota of pathogen-infected parrots and to develop probiotics specific to these genera.

  8. Molecular cloning of cellulase genes from indigenous bacterial isolates

    International Nuclear Information System (INIS)

    Jong Bor Chyan; Pauline Liew Woan Ying; Mat Rasol Awang

    2006-01-01

    Indigenous cellulolytic bacterial isolates having high activities in degrading carboxymethyl cellulose (CMC) were isolated from local environments. Identification of these isolates were performed by molecular techniques. By using polymerase chain reaction (PCR) techniques, PCR products encoding cellulase gene were amplified from the total genomic DNAs. Purified PCR product was successfully cloned and expressed in Escherichia coli host system. The complete nucleotide sequences of the cellulase genes determined. The analysis of amino acid sequences deduced from the genes indicated that the cloned DNA fragments show high homology to those of endoglucanase genes of family GH5. All cloned genes consist of an N-terminal signal peptide, a catalytic domain of family 5 glycosyl hydrolase and a cellulose-binding domain of family III. (Author)

  9. Molecular identification and biodegradation of 3-chloropropionic acid (3CP by filamentous fungi-Mucor and Trichoderma species isolated from UTM agricultural land

    Directory of Open Access Journals (Sweden)

    Parvizpour, S.

    2013-01-01

    Full Text Available Aims: This study was carried out to further characterize fungal species that could degrade 3-chloropropionic acid (3CPas sole source of carbon and energy. Methodology and Results: Both fungi were able to grow on 3CP after 10 days on solid minimal media. Based on sequencing of its segment of 18S rRNA these isolates were identified as Mucor sp. SP1 and Trichoderma sp. SP2. The isolated strains were not able to grow on media plates containing 10 mM of 2,2-dichloropropionate (2,2DCP as sole source of carbon. 3CP degradation was observed in liquid minimal medium containing 10 mM 3CP after 18 days cultureperiod. The chloride ion released was detected in both growth medium containing Mucor sp. SP1 and Trichoderma sp. SP2. At least 80% of 10 mM 3CP was utilized in the growth medium. Conclusion, significance and impact of study: Dehalogenase enzyme that can degrade α-chloro-substituted haloalkanoic acids for example 2,2DCP is well studied up to protein crystallization. Very few reports on the degradationof β-chloro-substituted haloalkanoic acids such as 3CP and none from fungi. This study is considered important because it can be compared to that of well-documented α-chloro-substituted haloalkanoic acids degradation. This is the first study to indicate fungal growth on 3CP as sole carbon and energy sources.

  10. The isolation and identification of predatory bacteria from a ...

    African Journals Online (AJOL)

    evaluate their lytic activities on Microcystis cells and identification. Isolates B2 and B16 had a lytic effect on the Microcystis cells with isolate B16 having a greater effect than isolate B2. Isolate B2 was identified as Pseudomonas stutzeri with 99.9% certainty and B16 as Bacillus mycoides with 99.7% certainty using the API ...

  11. Isolation and identification of pathogenic free-living amoeba from surface and tap water of Shiraz City using morphological and molecular methods.

    Science.gov (United States)

    Armand, B; Motazedian, M H; Asgari, Q

    2016-01-01

    Free-living amoebae (FLA) are the most abundant and widely distributed protozoa in the environment. An investigation was conducted to determine the presence of free-living amoebae (FLA), Acanthamoeba and Vermamoeba in waterfronts of parks and squares and tap water of Shiraz City, Iran. FLA are considered pathogenic for human. These ubiquitous organisms have been isolated from different environments such as water, soil, and air. Eighty-two water samples were collected from different places of Shiraz City during the summer of 2013. All samples were processed in Dept. of Parasitology and Mycology, Shiraz University of Medical Sciences, Fars, Iran. Samples were screened for FLA and identified by morphological characters in the cultures, PCR amplification targeting specific genes for each genus and sequencing determined frequent species and genotypes base on NCBI database. Overall, 48 samples were positive for Acanthamoeba and Vermamoeba in non-nutrient agar culture based on morphological characteristics. The PCR examination was done successfully. Sequencing results were revealed T4 (62.96 %) genotypes as the most common genotype of Acanthamoeba in the Shiraz water sources. In addition, T5 (33.33 %) and T15 (3.71 %) were isolated from water supplies. Vermamoeba vermiformis was known the dominant species from this genus. The high frequency of Acanthamoeba spp. and Vermamoeba in different environmental water sources of Shiraz is an alert for the public health related to water sources. The result highlights a need for taking more attention to water supplies in order to prevent illnesses related to free-living amoebae.

  12. Molecular identification and technological characterization of lactic acid bacteria isolated from fermented kidney beans flours (Phaseolus vulgaris L. and P. coccineus) in northwestern Argentina.

    Science.gov (United States)

    Sáez, Gabriel D; Hébert, Elvira M; Saavedra, Lucila; Zárate, Gabriela

    2017-12-01

    Legumes are an important protein source in developing countries and their flours represent an attractive alternative for the manufacture of gluten free products. In the present study, 4 kidney bean varieties (Alubia, Pallar, Black and Red beans) commonly cultivated in northwestern Argentina, were milled and spontaneously fermented in order to isolate and select autochthonous lactic acid bacteria (LAB) with relevant technological and functional properties for usage as starter cultures. Twelve doughs were fermented with daily back-slopping at 37°C for 6days and evolution of total mesophiles, lactic acid bacteria, and yeasts and molds populations were followed by plate counting. A combination of phenotypic and genotypic methods including (GTG) 5 -based PCR fingerprinting and 16S rRNA gene sequencing were used to differentiate and identify the isolated LAB to species level. LAB counts ranged from around 0.89±0.81 to 8.74±0.03logcfu/g with a pH decline from 6.4 to 3.9 throughout fermentation. Four genera and nine species of LAB: Enterococcus durans, E. faecium, E. mundtii, E. casseliflavus; Lactobacillus rhamnosus, Lactococcus garvieae, Weissella cibaria and W. paramesenteroides were found on kidney beans. Twenty five LAB strains were assessed for their abilities to grow on kidney bean extracts, acidifying capacities (pH and acidification rates), amylolytic, proteolytic, tannase and gallate decarboxylase activities as well as pathogens inhibition by antimicrobials. Based on these properties E. durans CRL 2178 and W. paramesenteroides CRL 2182 were inoculated singly and combined in Alubia kidney bean flour and fermented for 24h at 37°C. LAB strains were beneficial for removing trypsin inhibitors and tannins from sourdoughs and for improving amino acids and phenolics contents, increasing the antioxidant activities of kidney bean matrices. Selected strains have potential as starter cultures for obtaining fermented bean products with high nutritional and functional

  13. Isolation and Molecular Identification of Auxotrophic Mutants to Develop a Genetic Manipulation System for the Haloarchaeon Natrinema sp. J7-2

    Directory of Open Access Journals (Sweden)

    Jie Lv

    2015-01-01

    Full Text Available Our understanding of the genus Natrinema is presently limited due to the lack of available genetic tools. Auxotrophic markers have been widely used to construct genetic systems in bacteria and eukaryotes and in some archaeal species. Here, we isolated four auxotrophic mutants of Natrinema sp. J7-2, via 1-methyl-3-nitro-1-nitroso-guanidin mutagenesis, and designated them as J7-2-1, J7-2-22, J7-2-26, and J7-2-52, respectively. The mutant phenotypes were determined to be auxotrophic for leucine (J7-2-1, arginine (J7-2-22 and J7-2-52, and lysine (J7-2-26. The complete genome and the biosynthetic pathways of amino acids in J7-2 identified that the auxotrophic phenotype of three mutants was due to gene mutations in leuB (J7-2-1, dapD (J7-2-26, and argC (J7-2-52. These auxotrophic phenotypes were employed as selectable makers to establish a transformation method. The transformation efficiencies were determined to be approximately 103 transformants per µg DNA. And strains J7-2-1 and J7-2-26 were transformed into prototrophic strains with the wild type genomic DNA, amplified fragments of the corresponding genes, or the integrative plasmids carrying the corresponding genes. Additionally, exogenous genes, bgaH or amyH gene, were expressed successfully in J7-2-1. Thus, we have developed a genetic manipulation system for the Natrinema genus based on the isolated auxotrophic mutants of Natrinema sp. J7-2.

  14. Molecular diversity of Scottish Cryptosporidium hominis isolates.

    Science.gov (United States)

    Deshpande, A; Alexander, C L; Coyne, M; Brownlie, S; Smith-Palmer, A; Jones, B L

    2015-04-01

    Cryptosporidium hominis is one of the most prevalent protozoan parasites to infect humans where transmission is via the consumption of infective oocysts. This study describes sporadic cases in addition to the molecular diversity of outbreak cases in Scotland using the glycoprotein-60 subtyping tool. From a total of 187 C. hominis isolates, 65 were subjected to further molecular analysis and 46 were found to be the common IbA10G2 subtype. Unusual subtypes included four isolates belonging to the Ia family (IaA14R3, n = 12; IaA14R2, n = 1; IaA9G3, n = 1; IaA25R3, n = 2), two from the Id family (IdA24, n = 1; IdA17, n = 1) and one belonging to the Ie family, namely IeA11G3T3. These data contribute significantly to our knowledge and understanding of the molecular diversity of C. hominis isolates from outbreak investigations involving Scottish residents which will be beneficial for the management of future outbreaks.

  15. Molecular Identification of a Newly Isolated Bacillus subtilis BI19 and Optimization of Production Conditions for Enhanced Production of Extracellular Amylase.

    Science.gov (United States)

    Dash, Biplab Kumar; Rahman, M Mizanur; Sarker, Palash Kumar

    2015-01-01

    A study was carried out with a newly isolated bacterial strain yielding extracellular amylase. The phylogenetic tree constructed on the basis of 16S rDNA gene sequences revealed this strain as clustered with the closest members of Bacillus sp. and identified as Bacillus subtilis BI19. The effect of various fermentation conditions on amylase production through shake-flask culture was investigated. Rice flour (1.25%) as a cheap natural carbon source was found to induce amylase production mostly. A combination of peptone and tryptone as organic and ammonium sulfate as inorganic nitrogen sources gave highest yield. Maximum production was obtained after 24 h of incubation at 37 °C with an initial medium pH 8.0. Addition of surfactants like Tween 80 (0.25 g/L) and sodium lauryl sulfate (0.2 g/L) resulted in 28% and 15% increase in enzyme production, respectively. Amylase production was 3.06 times higher when optimized production conditions were used. Optimum reaction temperature and pH for crude amylase activity were 50 °C and 6.0, respectively. The crude enzyme showed activity and stability over a fair range of temperature and pH. These results suggest that B. subtilis BI19 could be exploited for production of amylase at relatively low cost and time.

  16. Molecular Identification of a Newly Isolated Bacillus subtilis BI19 and Optimization of Production Conditions for Enhanced Production of Extracellular Amylase

    Directory of Open Access Journals (Sweden)

    Biplab Kumar Dash

    2015-01-01

    Full Text Available A study was carried out with a newly isolated bacterial strain yielding extracellular amylase. The phylogenetic tree constructed on the basis of 16S rDNA gene sequences revealed this strain as clustered with the closest members of Bacillus sp. and identified as Bacillus subtilis BI19. The effect of various fermentation conditions on amylase production through shake-flask culture was investigated. Rice flour (1.25% as a cheap natural carbon source was found to induce amylase production mostly. A combination of peptone and tryptone as organic and ammonium sulfate as inorganic nitrogen sources gave highest yield. Maximum production was obtained after 24 h of incubation at 37°C with an initial medium pH 8.0. Addition of surfactants like Tween 80 (0.25 g/L and sodium lauryl sulfate (0.2 g/L resulted in 28% and 15% increase in enzyme production, respectively. Amylase production was 3.06 times higher when optimized production conditions were used. Optimum reaction temperature and pH for crude amylase activity were 50°C and 6.0, respectively. The crude enzyme showed activity and stability over a fair range of temperature and pH. These results suggest that B. subtilis BI19 could be exploited for production of amylase at relatively low cost and time.

  17. Extracellular polymeric substances (EPS) producing bacterial strains of municipal wastewater sludge: isolation, molecular identification, EPS characterization and performance for sludge settling and dewatering.

    Science.gov (United States)

    Bala Subramanian, S; Yan, S; Tyagi, R D; Surampalli, R Y

    2010-04-01

    Wastewater treatment plants often face the problems of sludge settling mainly due to sludge bulking. Generally, synthetic organic polymer and/or inorganic coagulants (ferric chloride, alum and quick lime) are used for sludge settling. These chemicals are very expensive and further pollute the environment. Whereas, the bioflocculants are environment friendly and may be used to flocculate the sludge. Extracellular polymeric substances (EPS) produced by sludge microorganisms play a definite role in sludge flocculation. In this study, 25 EPS producing strains were isolated from municipal wastewater treatment plant. Microorganisms were selected based on EPS production properties on solid agar medium. Three types of EPS (slime, capsular and bacterial broth mixture of both slime and capsular) were harvested and their characteristics were studied. EPS concentration (dry weight), viscosity and their charge (using a Zetaphoremeter) were also measured. Bioflocculability of obtained EPS was evaluated by measuring the kaolin clay flocculation activity. Six bacterial strains (BS2, BS8, BS9, BS11, BS15 and BS25) were selected based on the kaolin clay flocculation. The slime EPS was better for bioflocculation than capsular EPS and bacterial broth. Therefore, extracted slime EPS (partially purified) from six bacterial strains was studied in terms of sludge settling [sludge volume index (SVI)] and dewatering [capillary suction time (CST)]. Biopolymers produced by individual strains substantially improved dewaterability. The extracted slime EPS from six different strains were partially characterized. Copyright (c) 2009 Elsevier Ltd. All rights reserved.

  18. Identification and molecular characterization of a novel sugarcane streak mastrevirus and an isolate of the A-strain of maize streak virus from sugarcane in Nigeria.

    Science.gov (United States)

    Yahaya, Adama; Dangora, Danladi B; Alegbejo, Matthew D; Kumar, P Lava; Alabi, Olufemi J

    2017-02-01

    Sugarcane and maize plants showing symptoms typical of those described for the so-called "African streak viruses" (AfSVs) were encountered during field surveys conducted from February to July 2015 to document viruses infecting both crops across the northern Guinea savannah region of Nigeria. As part of this study, two categories of complete mastrevirus-like genome sequences were obtained from nine samples (maize = 2; sugarcane = 7). In pairwise comparisons, the full-length genomes of the first sequence category (2,687 nt each; maize = 2; sugarcane = 2) shared 96 to 99% identity with global isolates of the A-strain of maize streak virus (MSV-A), indicating that sugarcane may also serve as a reservoir host to MSV-A. Analysis of the complete genomes belonging to the second sequence category (2,757 nt each; sugarcane = 5) showed that they shared 42 to 67% identity with their closest AfSV relatives, thus indicating that they represent sequences of a novel mastrevirus. Both sequence categories shared 61-62% sequence identity with each other. Further analysis revealed that the novel sugarcane-infecting virus, tentatively named as sugarcane chlorotic streak virus (SCSV), arose from a putative interspecific recombination event involving two grass-infecting mastreviruses, eragrostis streak virus and urochloa streak virus, as putative parental sequences. The results of this study add to the repertoire of diverse AfSVs present in cereal and sugarcane mixed cropping landscapes in the northern Guinea savannah region of Nigeria, with implications for disease epidemiology.

  19. Molecular identification and population dynamic of Anisakis pegreffii (Nematoda: Anisakidae Dujardin, 1845) isolated from the European anchovy (Engraulis encrasicolus L.) in the Adriatic Sea.

    Science.gov (United States)

    Mladineo, Ivona; Simat, Vida; Miletić, Jelena; Beck, Relja; Poljak, Vedran

    2012-07-02

    Anchovy Engraulis encrasicolus (L.) is a coastal pelagic and euryhaline species that represents the only European species of the family Engraulidae, with a widespread distribution. In Croatia, it is marketed fresh, frozen, salted or marinated and mainly exported to Italy and Spain, however Anisakis sp. larval infection is frequently the reason for border rejection. Since it is known that the prevalence and intensity of Anisakis infection varies with fish species, fishing area and season, the aim of our study was to identify Anisakis sp. parasitizing European anchovy and infer its population dynamic through a 2.5-year period. Larvae were found coiled and encysted on the external wall of intestine (94%) and reproductive organs (6%), rarely in fillets. Prevalence was 76.1% (95% confidence limits 74.51-77.56%), mean abundance 6.59 (bootstrap 95% confidence limits 5.81-7.26) and mean intensity 8.67 (bootstrap 95% confidence limits 7.82-9.35). The partial CO2 mitochondrial DNA sequence of the isolated anisakids confirmed clustering of the anchovy parasite within A. pegreffii sister group. Parasite population structure showed plasticity inferred by fishing ground, sampling year and fish gender and size. Compared to anisakid prevalence/abundance in other fish, the European anchovy in the Adriatic Sea represents a moderately high-infected paratenic host, although in the Mediterranean and Atlantic waters, anchovies have shown strikingly lesser values of prevalence. Since this host represents one of the most attractive Mediterranean fisheries products traditionally consumed without thermal preparation that in any case would not disrupt larval antigenicity and prevent human allergies, and given the high prevalence of the anisakid within the host, it is necessary to include anchovy into more firm risk assessment frames in order to develop measures that will support the safe alimentary production and consumption of seafood. Copyright © 2012 Elsevier B.V. All rights reserved.

  20. Isolation, identification and characterization of regional indigenous Saccharomyces cerevisiae strains.

    Science.gov (United States)

    Šuranská, Hana; Vránová, Dana; Omelková, Jiřina

    2016-01-01

    In the present work we isolated and identified various indigenous Saccharomyces cerevisiae strains and screened them for the selected oenological properties. These S. cerevisiae strains were isolated from berries and spontaneously fermented musts. The grape berries (Sauvignon blanc and Pinot noir) were grown under the integrated and organic mode of farming in the South Moravia (Czech Republic) wine region. Modern genotyping techniques such as PCR-fingerprinting and interdelta PCR typing were employed to differentiate among indigenous S. cerevisiae strains. This combination of the methods provides a rapid and relatively simple approach for identification of yeast of S. cerevisiae at strain level. In total, 120 isolates were identified and grouped by molecular approaches and 45 of the representative strains were tested for selected important oenological properties including ethanol, sulfur dioxide and osmotic stress tolerance, intensity of flocculation and desirable enzymatic activities. Their ability to produce and utilize acetic/malic acid was examined as well; in addition, H2S production as an undesirable property was screened. The oenological characteristics of indigenous isolates were compared to a commercially available S. cerevisiae BS6 strain, which is commonly used as the starter culture. Finally, some indigenous strains coming from organically treated grape berries were chosen for their promising oenological properties and these strains will be used as the starter culture, because application of a selected indigenous S. cerevisiae strain can enhance the regional character of the wines. Copyright © 2015 Sociedade Brasileira de Microbiologia. Published by Elsevier Editora Ltda. All rights reserved.

  1. Isolation, identification and characterization of regional indigenous Saccharomyces cerevisiae strains

    Directory of Open Access Journals (Sweden)

    Hana Šuranská

    2016-03-01

    Full Text Available Abstract In the present work we isolated and identified various indigenous Saccharomyces cerevisiae strains and screened them for the selected oenological properties. These S. cerevisiae strains were isolated from berries and spontaneously fermented musts. The grape berries (Sauvignon blanc and Pinot noir were grown under the integrated and organic mode of farming in the South Moravia (Czech Republic wine region. Modern genotyping techniques such as PCR-fingerprinting and interdelta PCR typing were employed to differentiate among indigenous S. cerevisiae strains. This combination of the methods provides a rapid and relatively simple approach for identification of yeast of S. cerevisiae at strain level. In total, 120 isolates were identified and grouped by molecular approaches and 45 of the representative strains were tested for selected important oenological properties including ethanol, sulfur dioxide and osmotic stress tolerance, intensity of flocculation and desirable enzymatic activities. Their ability to produce and utilize acetic/malic acid was examined as well; in addition, H2S production as an undesirable property was screened. The oenological characteristics of indigenous isolates were compared to a commercially available S. cerevisiae BS6 strain, which is commonly used as the starter culture. Finally, some indigenous strains coming from organically treated grape berries were chosen for their promising oenological properties and these strains will be used as the starter culture, because application of a selected indigenous S. cerevisiae strain can enhance the regional character of the wines.

  2. Molecular identification of the Sporothrix schenckii complex.

    Science.gov (United States)

    Oliveira, Manoel Marques Evangelista; Almeida-Paes, Rodrigo; Gutierrez-Galhardo, Maria Clara; Zancope-Oliveira, Rosely M

    2014-01-01

    Sporothrix schenckii, an ascomycetous dimorphic organism that for over a century was recognized as the sole agent of sporotrichosis, a subcutaneous mycosis with a worldwide distribution. However, it has been proposed, based on physiologic and molecular aspects, that S. schenckii is a complex of distinct species: Sporothrix brasiliensis, Sporothrix mexicana, Sporothrix globosa, S. schenckii sensu strictu, Sporothrix luriei, and Sporothrix albicans (formerly Sporothrix pallida). Human disease has a broad range of clinical manifestations and can be classified into fixed cutaneous, lymphocutaneous, disseminated cutaneous, and extracutaneous sporotrichosis. The gold standard for the diagnosis of sporotrichosis is the culture; however, serologic, histopathologic and molecular approaches have been recently adopted for the diagnosis of this mycosis. Few molecular methods have been applied to the diagnosis of sporotrichosis to detect S. schenckii DNA from clinical specimens, and to identify Sporothrix spp. in culture. Until now, Sporothrix is the unique clinically relevant dimorphic fungus without an elucidated genome sequence, thus limiting molecular knowledge about the cryptic species of this complex, and the sexual form of all S. schenckii complex species. In this review we shall focus on the current diagnosis of the sporotrichosis, and discuss the current molecular tools applied to the diagnosis and identification of the Sporothrix complex species. This manuscript is part of the series of works presented at the "V International Workshop: Molecular genetic approaches to the study of human pathogenic fungi" (Oaxaca, Mexico, 2012). Copyright © 2013 Revista Iberoamericana de Micología. Published by Elsevier Espana. All rights reserved.

  3. Detection, Isolation, and Identification of Vibrio cholerae from the Environment

    Science.gov (United States)

    Huq, Anwar; Haley, Bradd J.; Taviani, Elisa; Chen, Arlene; Hasan, Nur A.; Colwell, Rita R.

    2012-01-01

    Recent molecular advances in microbiology have greatly improved the detection of bacterial pathogens in the environment. Improvement and a downward trend in the cost of molecular detection methods have contributed to increased frequency of detection of pathogenic microorganisms where traditional culture-based detection methods have failed. Culture methods also have been greatly improved and the confluence of the two suites of methods provides a powerful tool for detection, isolation, and characterization of pathogens. While molecular detection provides data on the presence and type of pathogens, culturing methods allow a researcher to preserve the organism of interest for “–omics” studies, such as genomic, metabolomic, secretomic, and transcriptomic analysis, which are rapidly becoming more affordable. This has yielded a clearer understanding of the ecology and epidemiology of microorganisms that cause disease. Specifically, important advances have been made over the past several years on isolation, detection, and identification of Vibrio cholerae, the causative agent of cholera in humans. In this unit, we present commonly accepted methods for isolation, detection, and characterization of V. cholerae, providing more extensive knowledge of the ecology and epidemiology of this organism. This unit has been fully revised and updated from the earlier unit (Huq, Grim et al. 2006) with the latest knowledge and additional information not previously included. We have also taken into account of cost of reagents and equipment that may be prohibitive for many researchers and have, therefore, included protocols for all laboratories, including those with limited resources, likely to be located in regions of cholera endemicity. PMID:22875567

  4. Isolation, characterization and identification of actinomycetes from ...

    African Journals Online (AJOL)

    A total of 62 isolates of actinomycetes were isolated from 7 soil samples collected from Agriculture Research Center Semongok, Sarawak. All 62 isolates exhibited a range of colony colours (dark grey, grey, dark brown, brownish, whitish and yellowish white). All the isolates were later purified and subjected to a few ...

  5. Species identification of Streptococcus bovis group isolates causing bacteremia

    DEFF Research Database (Denmark)

    Agergaard, Charlotte N; Knudsen, Elisa; Dargis, Rimtas

    2017-01-01

    This study compared two MALDI-TOF MS systems (Biotyper and VITEK MS) on clinical Streptococcus bovis group isolates (n=66). The VITEK MS gave fewer misidentifications and a higher rate of correct identifications than the Biotyper. Only the identification of S. lutetiensis by the VITEK MS was reli......This study compared two MALDI-TOF MS systems (Biotyper and VITEK MS) on clinical Streptococcus bovis group isolates (n=66). The VITEK MS gave fewer misidentifications and a higher rate of correct identifications than the Biotyper. Only the identification of S. lutetiensis by the VITEK MS...

  6. Isolation and identification of radiation resistant yeasts from sea water

    International Nuclear Information System (INIS)

    Park, Jong Cheon; Jeong, Yong Uk; Kim, Du Hong; Jo, Eun A

    2011-12-01

    This study was conducted to isolate radiation-resistant yeasts from sea water for development of application technology of radiation-resistant microorganism. · Isolation of 656 yeasts from sea water and selection of 2 radiation-resistant yeasts (D 10 value >3) · Identification of isolated yeasts as Filobasidium elegans sharing 99% sequence similarity · Characterization of isolated yeast with ability to repair of the DNA damage and membrane integrity to irradiation

  7. Isolation, genetic diversity and identification of a virulent pathogen of ...

    African Journals Online (AJOL)

    Isolation, genetic diversity and identification of a virulent pathogen of eriophyid mite, Aceria guerreronis (Acari: Eriophyidae) by DNA marker in Karnataka, India. Basavaraj Kalmath, B Mallik, S Onkarappa, R Girish, N Srinivasa ...

  8. Isolation and identification of two galangin metabolites from rat urine ...

    African Journals Online (AJOL)

    Isolation and identification of two galangin metabolites from rat urine and determination of their in vitro hypolipidemic activity. Xuguang Zhang, Shouqian Cheng, Hailong Li, Xiaopo Zhang, Feng Chen, Youbin Li, Junqing Zhang, Yinfeng Tan ...

  9. Isolation and identification of flavonoids from anticancer and ...

    African Journals Online (AJOL)

    Isolation and identification of flavonoids from anticancer and neuroprotective extracts of Trigonella foenum graecum. Shabina Ishtiaq Ahmed, Muhammad Qasim Hayat, Saadia Zahid, Muhammad Tahir, Qaisar Mansoor, Muhammad Ismail, Kristen Keck, Robert Bates ...

  10. Isolation and identification of Metarhizium anisopliae from Chilo ...

    African Journals Online (AJOL)

    *

    2012-04-12

    Apr 12, 2012 ... 1Environment and Plant Protection Institute, Chinese Academy of Tropical Agricultural Sciences (Key Laboratory of ... Keywords: Metarhizium anisopliae, isolation, identification, Chilo venosatus, culture medium, biological control. .... with a Leica microscope and average values were compared for all.

  11. Identification, isolation and in vitro antimicrobial susceptibility testing ...

    African Journals Online (AJOL)

    Identification, isolation and in vitro antimicrobial susceptibility testing of Aeromonas veronii associated with an acute death of Channel Catfish (Ictalurus lunetas) in China. X Huang, K Wang, D Zong-Jun, Y Geng, Y Deng ...

  12. MORPHOLOGICAL AND MOLECULAR IDENTIFICATION OF Fusarium SPECIES AND THEIR PATHOGENICITY FOR WHEAT

    Directory of Open Access Journals (Sweden)

    Jelena Poštić

    2012-12-01

    Full Text Available From the root and lower stem parts of weeds and plant debris of maize, wheat, oat and sunflower we isolated 300 isolates of Fusarium spp. and performed morphological and molecular identification. With molecular identification using AFLP method we determined 14 Fusarium species: F. acuminatum, F. avenaceum, F. concolor, F. crookwellense, F. equiseti, F. graminearum, F. oxysporum, F. proliferatum, F. semitectum, F. solani, F. sporotrichioides, F. subglutinans, F. venenatum and F. verticillioides.By comparing results of morphological and molecular identification we found out that determination of 16,7% isolates was incorrect. Out of 300 isolates identified with molecular methods, 50 did not belong to the species determined with morphological determination.With pathogenicity tests of 30 chosen Fusarium isolates we determined that many of them were pathogenic to wheat and maize seedlings and to wheat heads. The most pathogenic were isolates of F. graminearum from A. retroflexus, A. theophrasti and C. album, F. venenatum from maize debris and and A. theophrasti, F. crookwellense from A. lappa. Antifungal influence of 11 essential oils on mycelia growth and sporulation of chosen Fusarium isolates determined that essential oils of T. vulgaris, P. anisum and E. caryophyllus had the strongest effect on mycelial growth. Influence of essential oils on sporulation was not statistically significant.

  13. Antifungal activity and molecular identification of endophytic fungi ...

    African Journals Online (AJOL)

    Antifungal activity and molecular identification of endophytic fungi from the angiosperm Rhodomyrtus tomentosa. Juthatip Jeenkeawpieam, Souwalak Phongpaichit, Vatcharin Rukachaisirikul, Jariya Sakayaroj ...

  14. [Isolation, identification and insecticidal activity of endophyte from Achnatherum inebrians].

    Science.gov (United States)

    Zhang, Xuebing; Shi, Yingwu; Wang, Xiaoxia; Zhang, Wei; Lou, Kai

    2010-04-01

    To study endophyte species of Achnatherum inebrians and to screen strains with insecticidal activity against cotton insect. We isolated endophytic from roots,stems,leaves and seeds of health A. inebrians by grinding separation method and identified by a dual approach of morphological and physiological observation and 16S rDNA gene (for bacteria) and ITS sequence (for fungi) based molecular identification. Then,those endophytes were inoculated into liquid media for fermentation and the crude extracts were used to test insecticidal activities by slide disc immersion and nebulization methods. We isolated bacteria species classified into 8 genera of Bacillus, Streptomyces, Corynebacterium, Phyllobacterium, sphingomonnas, Paenibacillus, Pseudomonas, Acinetobacter and 2 fungi of Claviceps purpure and Claviceps Chaetomium. Of them, the strain Streptomyces rochei (GA) and Claviceps purpurea (PF-2) had more than 85% of mortality to cotton aphis. Two strains of PF-2 and GA associated within the A. inebrians had significant insecticidal activity to cotton aphis (Aphis gossypii), which may provide a new biological resource to explore new microbial insecticide.

  15. Isolation, cloning and molecular characterization of a thermotolerant ...

    African Journals Online (AJOL)

    Isolation, cloning and molecular characterization of a thermotolerant xylanase from Streptomyces sp. THW31. Thayat Sriyapai, Peechapack Somyoonsap, Supatra Areekit, Paisarn Khawsak, Arda Pakpitcharoen, Kosum Chansiri ...

  16. Isolation and identification of fungi responsible for leaf spots disease ...

    African Journals Online (AJOL)

    The diseased plant leaves were taken to the laboratory for culture, isolation, and identification of pathogens. Plant disease incidence was determined using disease index and severity scale of 0-4 rating. Eleven fungal pathogens were isolated and identified. These included; Alternaria longipes, Aspergillus fumigatus, ...

  17. Molecular characterization of lactobacilli isolated from fermented idli batter

    Directory of Open Access Journals (Sweden)

    Perumal Jayaprabha Agaliya

    2013-12-01

    Full Text Available Lactic acid bacteria are non pathogenic organism widely distributed in nature typically involved in a large number of spontaneous food fermentation. The purpose of this study was to characterize the bacteriocinogenic lactobacilli from fermented idli batter which can find application in biopreservation and biomedicine. Eight most promising lactobacilli were chosen from twenty two isolates based on their spectrum of activity against other lactic acid bacteria and pathogens. The eight lactobacilli were characterized based on the various classical phenotypic tests, physiological tests and biochemical tests including various carbohydrate utilization profiles. All isolates were homo fermentative, catalase, and gelatin negative. Molecular characterization was performed by RAPD, 16S rRNA analysis, 16S ARDRA, and Multiplex PCR for species identification. RAPD was carried out using the primer R2 and M13. Five different clusters were obtained based on RAPD indicating strain level variation. 16S rRNA analysis showed 99 to 100% homology towards Lactobacillus plantarum. The restriction digestion pattern was similar for all the isolates with the restriction enzyme AluI. The subspecies were identified by performing Multiplex PCR using species specific primer. Among the five clusters, three clusters were clearly identified as Lactobacillus plantarum subsp. plantarum, Lactobacillus pentosus, and Lactobacillus plantarum subsp. argentoratensis.

  18. Molecular characterization of lactobacilli isolated from fermented idli batter.

    Science.gov (United States)

    Agaliya, Perumal Jayaprabha; Jeevaratnam, Kadirvelu

    2013-12-01

    Lactic acid bacteria are non pathogenic organism widely distributed in nature typically involved in a large number of spontaneous food fermentation. The purpose of this study was to characterize the bacteriocinogenic lactobacilli from fermented idli batter which can find application in biopreservation and biomedicine. Eight most promising lactobacilli were chosen from twenty two isolates based on their spectrum of activity against other lactic acid bacteria and pathogens. The eight lactobacilli were characterized based on the various classical phenotypic tests, physiological tests and biochemical tests including various carbohydrate utilization profiles. All isolates were homo fermentative, catalase, and gelatin negative. Molecular characterization was performed by RAPD, 16S rRNA analysis, 16S ARDRA, and Multiplex PCR for species identification. RAPD was carried out using the primer R2 and M13. Five different clusters were obtained based on RAPD indicating strain level variation. 16S rRNA analysis showed 99 to 100% homology towards Lactobacillus plantarum. The restriction digestion pattern was similar for all the isolates with the restriction enzyme AluI. The subspecies were identified by performing Multiplex PCR using species specific primer. Among the five clusters, three clusters were clearly identified as Lactobacillus plantarum subsp. plantarum, Lactobacillus pentosus, and Lactobacillus plantarum subsp. argentoratensis.

  19. Isolation, identifications and antimicrobial susceptibility pattern of ...

    African Journals Online (AJOL)

    A cross-sectional study was conducted from November 2013 to April 2014 to isolate coagulase positive Staphylococcus (CPS) from subclinical mastitic (SCM) lactating cows, to establishing prevalence, to identify risk factors and to determine the antimicrobial susceptibility pattern of CPS isolates in and around Haramaya ...

  20. Identificación Molecular de Bacterias Productoras de Polihidroxialcanoatos en Subproductos de Lácteos y Caña de Azúcar / Molecular Identification of Polyhydroxyalkanoate-Producing Bacteria Isolated from Dairy and Sugarcane Residues

    Directory of Open Access Journals (Sweden)

    Ana Carolina Cardona Echavarría

    2013-12-01

    . In this work, the presence of PHAs-producing bacteria in whey, sugarcane molasses, cachaza and bagasse was investigated. Bacteria were isolated using a minimal medium supplemented with 2% glucoseand 1 μL mL-1 Nile red (0.1%. Colonies exhibitingfluorescence at 340 nm were further analyzed by fluorescence microscopy with Nileblue. When positive for both tests, bacterial isolates were classified as PHAs producers and their 16S rDNA sequenced. For selected isolates, the presence of the phaC gene was confirmed by PCR. A total of 38 strains, grouped into 18 different morphotypes, were identified as PHA producers. Bacteria belonging to Lactococcus, Klebsiella, Pseudomonas, Enterobacter and Enterococcus genera were isolated from whey and Bacillus, Enterobacter, Pantoea, Klebsiella and Gluconobacter from sugar cane residues. The phaC gene was detected by PCR in 16 bacteria withtype I and IV arrangements. This work opens up the possibility of using the isolated bacteria as an environmentally sustainablealternative for the disposal of agro-industrial residues as well as an additional income source.

  1. Isolation and Identification of the Chitinolytic Bacteria from Rumen Ecosystem

    Directory of Open Access Journals (Sweden)

    Sri Rahayu

    2003-05-01

    Full Text Available Rumen is an interesting ecosystem for microbial exploration and their products. Isolation of the chitinolytic bacteria from the rumen ecosystem found 109 colonies that produced clear zone, 84 colonies (86% anaerobic and 17 colonies (14% aerobic. Clear zone appeared in the third and fourth days incubation. Four potential isolates were chosen for identification purposes. Results showed that the bacteria were sticky, gram-positive, motile, endospore-forming, mesophilic and aerobic. It was supposed to Bacillus spp. the optimal pH and temperature to produce chitinase from isolate 18 are pH 6.0 and temperature of 35-40ºC. Divalent cations Mg, Ca, Zn, and Mn increase chitinase activity, while Cu and Co inhibit enzyme activity. When isolate 18 was grown on shrimp waste meal, it showed aptimal activity on the fifth days incubation. (Animal Production 5(2: 73-78 (2003   Key Words : Isolation, Identification, Chitinolytic Bacteria, Rumen

  2. Characterization and identification of streptococci isolated from bovine mammary glands.

    Science.gov (United States)

    Watts, J L

    1988-06-01

    A total of 317 gram-positive, catalase-negative cocci isolated from bovine mammary glands were characterized and identified using current species descriptions. Two hundred eighty-seven isolates (90.5%) could be placed in 11 distinct species. Streptococcus uberis was the most frequently encountered species and could be separated into two previously described genetic types based upon sucrose utilization. Streptococcus dysgalactiae and a newly described species, Streptococcus saccharolyticus, were the most frequently isolated organisms from teat canal swabs. Thirty isolates could not be placed in currently described species. A proposed identification scheme based upon serological grouping and seven biochemical tests would permit 24 h identification of streptococci isolated from bovine mammary glands.

  3. Identification of human pathogens isolated from blood using microarray hybridisation and signal pattern recognition

    Directory of Open Access Journals (Sweden)

    Presterl Elisabeth

    2007-08-01

    Full Text Available Abstract Background Pathogen identification in clinical routine is based on the cultivation of microbes with subsequent morphological and physiological characterisation lasting at least 24 hours. However, early and accurate identification is a crucial requisite for fast and optimally targeted antimicrobial treatment. Molecular biology based techniques allow fast identification, however discrimination of very closely related species remains still difficult. Results A molecular approach is presented for the rapid identification of pathogens combining PCR amplification with microarray detection. The DNA chip comprises oligonucleotide capture probes for 25 different pathogens including Gram positive cocci, the most frequently encountered genera of Enterobacteriaceae, non-fermenter and clinical relevant Candida species. The observed detection limits varied from 10 cells (e.g. E. coli to 105 cells (S. aureus per mL artificially spiked blood. Thus the current low sensitivity for some species still represents a barrier for clinical application. Successful discrimination of closely related species was achieved by a signal pattern recognition approach based on the k-nearest-neighbour method. A prototype software providing this statistical evaluation was developed, allowing correct identification in 100 % of the cases at the genus and in 96.7 % at the species level (n = 241. Conclusion The newly developed molecular assay can be carried out within 6 hours in a research laboratory from pathogen isolation to species identification. From our results we conclude that DNA microarrays can be a useful tool for rapid identification of closely related pathogens particularly when the protocols are adapted to the special clinical scenarios.

  4. Testing of DNA isolation for the identification of hemp

    Directory of Open Access Journals (Sweden)

    Tomáš Vyhnánek

    2015-12-01

    Full Text Available Hemp is diploid organism (2n = 2x = 20, genome size 534 Mb with nine pairs of autosomes plus XX (♀ or XY (♂ chromosomes. Cannabis sativa L. is an important economic plant for the production of food, fibre, oils, and intoxicants. Genotypes (varieties or chemovar of hemp with low Δ9-tetrahydrocannabinol content are used for industrial applications. Varieties with high Δ9-tetrahydrocannabinol or high cannabidiol content are used for medicinal applications. Biochemical and molecular methods can be used for identification and classification. An important step for molecular biology methods is to obtain the matrix of the native and sufficiently pure DNA. We tested two different experimental variant of samples (20 mg and 100 mg of seeds, oilcake and dried flowers for analysis of the Italian variety Carmagnola for analysis (harvested in 2014, Hempoint Ltd., Czech Republic. The DNeasy® Plant Mini Kit (Qiagen, GE was used to isolate the DNA. The DNA concentration and purity was assessed by agarose electrophoresis and via a spectrophotometer. Samples of lower weight yielded lower values of DNA concentration (average 16.30 - 38.90 ng.µL-1, but with better purity than samples of higher weight (ratio A260nm/A280nm for low-weight samples was near 1.80. To test the applicability of DNA analysis, we used two SSR markers (CAN1347 and CAN2913. PCR products were separated on 1% agarose and on 8% polyacrylamide electrophoresis. DNA samples obtained from samples of higher weight exhibited less PCR amplification than samples of lower weight. We found no effect of sample weight on the formation of non-specific amplification products during the PCR reaction. Based on our results we can be recommended for practical isolation procedure using DNeasy® Plant Mini Kit with lower of sample weight (20 mg. In future work the procedure for DNA isolating from wheat-cannabis products, e. g. breads, rolls or pasta, will be optimized.

  5. Sporothrix schenckii complex in Iran: Molecular identification and antifungal susceptibility.

    Science.gov (United States)

    Mahmoudi, Shahram; Zaini, Farideh; Kordbacheh, Parivash; Safara, Mahin; Heidari, Mansour

    2016-08-01

    Sporotrichosis is a global subcutaneous fungal infection caused by the Sporothrix schenckii complex. Sporotrichosis is an uncommon infection in Iran, and there have been no phenotypic, molecular typing or antifungal susceptibility studies of Sporothrix species. This study aimed to identify nine Iranian isolates of the S. schenckii complex to the species level using colony morphology, carbohydrate assimilation tests, and PCR-sequencing of the calmodulin gene. The antifungal susceptibilities of these Sporothrix isolates to five antifungal agents (amphotericin B (AMB), voriconazole (VRC), itraconazole (ITC), fluconazole (FLC), and terbinafine (TRB)) were also evaluated according to the M27-A3 and M38-A2 protocols of the Clinical and Laboratory Standards Institute for yeast and mycelial phases, respectively. Five of seven clinical isolates were identified as S. schenckii, and two clinical and two environmental isolates were identified as S. globosa. This is the first report of S. globosa in Iran. There was significant agreement (73%) between the results of the phenotypic and genotypic identification methods. TRB and ITC were the most effective antifungals against the Sporothrix isolates. The minimum inhibitory concentration (MIC) values of TRB for the yeast and mycelial phases of S. schenckii differed significantly. There was also a significant difference in the minimum fungicidal concentration (MFC) values of AMB and TRB for the two phases. Considering the low efficacy of VRC and FLC and the wide MIC ranges of AMB (1-16 μg/ml and 1-8 μg/ml for yeast and mycelial forms, respectively) observed in the present study, in vitro antifungal susceptibility testing should be performed to determine appropriate therapeutic regimens. © The Author 2016. Published by Oxford University Press on behalf of The International Society for Human and Animal Mycology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  6. Occurrence, isolation and DNA identification of Streptococcus ...

    African Journals Online (AJOL)

    ajl yemi

    2011-11-28

    Nov 28, 2011 ... Streptococcus thermophilus isolates from traditional butter 'Smen', a fermented product from cow's ... temperature of milk. The fresh butter is removed manually in a single lump called 'Zebda' which is trans- formed into 'Smen' by washing with salt ... Egyptian artisanal butter 'zabady' was reported recently.

  7. Isolation, characterization and identification of actinomycetes from ...

    African Journals Online (AJOL)

    STORAGESEVER

    Further study into the utilization of the actinomycetes for agriculture industry will be done to fully utilize ... Previous study showed that actinomycetes isolated from Malaysia soil have the potential to inhibit the growth of several tested plant pathogens (Jeffrey et al., ..... environment the microbes (in this case actinomycetes).

  8. Isolation and identification of Profenofos degrading bacteria

    Directory of Open Access Journals (Sweden)

    Saadatullah Malghani

    2009-12-01

    Full Text Available An enrichment culture technique was used to isolate bacterial strains responsible for the biodegradation of profenofos in a soil from Hubei province of central China. Two pure bacterial cultures, named W and Y, were isolated and subsequently characterized by sequencing of 16S rRNA genes and biochemical tests. Isolate W showed 96% similarity to the 16S rRNA gene of a Pseudomonas putida unlike Y which showed 99% similarity to the 16S rRNA gene of Burkholderia gladioli. Both strains grew well at pH 5.5-7.2 with a broad temperature profile ranging from 28º to 36 ºC. Bioremediation of profenofos-contaminated soil was examined using soil treated with 200 ug g-1; profenofos resulted in a higher degradation rate than control soils without inoculation. In a mineral salt medium (FTW reduction in profenofos concentration was 90% within 96 hours of incubation. A literature survey revealed that no data is available regarding the role of Burkholderia gladioli on pesticide biodegradation as well as on profenofos.

  9. Molecular characterization of Danish Cryptosporidium parvum isolates

    DEFF Research Database (Denmark)

    Enemark, Heidi L.; Ahrens, Peter; Juel, Cynthia Dawn

    2002-01-01

    The genetic polymorphism among 271 Danish Cryptosporidium isolates of human and animal origin was studied by partial amplification and sequencing of the Cryptosporidium oocyst wall protein (COWP) gene, the 18S rDNA, and a microsatellite locus.dagger Furthermore, the microsatellite locus was studied...... (P Cryptosporidium isolates of human origin the anthroponotic subgenotype H1 was identified, in addition to the zoonotic subgenotypes C1, C2, and C3. Of 44 human samples, 56.8% were anthroponotic, whereas 40.9% were zoonotic genotypes. One human isolate...... was characterized as C. meleagridis. The porcine Cryptosporidium isolates (N = 4) revealed a pattern which was genetically distinct from human and bovine isolates. Cryptosporidium in a hedgehog (Erinaceus europaeus L.) was identified for the first time. By microsatellite sequencing the hedgehog isolate showed...

  10. Isolation and identification of Mycoplasma mycoides cluster strains from goats in Chongqing, China

    Directory of Open Access Journals (Sweden)

    Wang Haoju

    2014-03-01

    Full Text Available In order to evaluate the prevalence of the Mycoplasma mycoides cluster in goats in Chongqing, China, an epidemiological survey in this area was carried out. A total of 68 samples were subjected to bacteria isolation on Hartley’s medium. Four isolates (three from lung tissue and one from nasal discharges were recovered from the samples and identified as the Mycoplasma species by their morphological and biochemical characteristics. They were further confirmed by PCR using 16S rRNA specific primer pairs and by restriction enzyme analysis. In vitro antimicrobial susceptibility of the isolates indicated that some strains had developed resistance to the antibiotics tested. This is the first report on the isolation, identification, and molecular characterisation of Mycoplasma species isolated from goats in Chongqing. This study also revealed a prevalence of Mycoplasma species infection in goats in this area.

  11. Molecular identification of trypanosomatids in wild animals.

    Science.gov (United States)

    Tenório, M S; Oliveira e Sousa, L; Alves-Martin, M F; Paixão, M S; Rodrigues, M V; Starke-Buzetti, W A; Araújo Junior, J P; Lucheis, S B

    2014-06-16

    Diverse wild animal species can be reservoirs of zoonotic flagellate parasites, which can cause pathologic Chagas disease. The present study aimed to detect the natural occurrence of flagellate parasites through direct microscopic examination of the parasites in blood samples and through PCR of whole blood and blood culture (haemoculture) samples from 38 captive and 65 free-living wild animals in the Centre for Conservation of Wild Fauna (CCWF), an area endemic for leishmaniasis. For this study, PCR was accomplished using primers for the ribosomal region (ITS-1) of the flagellate parasites. The amplified fragments were cloned and sequenced to identify DNA of the Trypanosomatid parasite species, observed in blood cultures from 3.9% (04/103) of the animals. Through these techniques, Trypanosoma cruzi was identified in haemoculture samples of the following three free-living species: common agouti (Dasyprocta aguti), white-eared opossum (Didelphis albiventris), and nine-banded armadillo (Dasypus novemcinctus). Furthermore, Trypanosoma minasense was identified in whole blood samples from 01 (0.9%) captive animal (black howler monkey-Alouatta caraya). These results demonstrated the first report of T. cruzi isolation in wild species from the CCWF using blood culture, which can be applied in addition to molecular tools for epidemiological studies and to identify trypanosomatids in wild animals. Copyright © 2014 Elsevier B.V. All rights reserved.

  12. Occurrence, isolation and DNA identification of streptococcus thermophilus involved in algerian traditional butter 'Smen'

    OpenAIRE

    Labtar, Asmaa; Delorme, Christine; Renault, Pierre

    2011-01-01

    Streptococcus thermophilus isolates from traditional butter 'Smen', a fermented product from cow's and ewe's milk in arid area was subjected to taxonomical investigations. The identification procedure included phenotypic approaches, molecular characterization by using genus polymerase chain reaction (PCR) amplifications for sodA gene encoding the manganese-dependant superoxide dismutase A, and species-specific primers from gene encoding glucose kinase (glcK), gene encoding DNA polymerase III ...

  13. Antifungal susceptibilities and identification of species of the Sporothrix schenckii complex isolated in Brazil.

    Science.gov (United States)

    Ottonelli Stopiglia, Cheila Denise; Magagnin, Cibele Massotti; Castrillón, Mauricio Ramírez; Mendes, Sandra Denise Camargo; Heidrich, Daiane; Valente, Patricia; Scroferneker, Maria Lúcia

    2014-01-01

    Sporotrichosis is a subacute or chronic mycosis caused worldwide by the dimorphic species complex, Sporothrix schenckii. We studied 85 isolates recovered in Brazil to verify their identification and evaluate their in vitro antifungal susceptibility patterns. Based on phenotypic tests (microscopic features, ability to grow at 30°C and 37°C, colony diameters, as well as assimilation of sucrose and raffinose) and molecular assays (amplification of a fragment of the calmodulin gene), the strains were identified as S. schenckii, S. brasiliensis and S. globosa, with a predominance of S. schenckii isolates. There was 37.7% disagreement between the phenotypic and genotypic identification methodologies. In general, terbinafine was the most active drug, followed by ketoconazole and itraconazole, and the less active fluconazole and voriconazole. Five isolates (one S. globosa and four S. schenckii) were found to be itraconazole-resistant strains but, in general, there were no differences in the in vitro antifungal susceptibility profiles among the Sporothrix species.

  14. Molecular characterization of Trichoderma sp. isolated from ...

    African Journals Online (AJOL)

    The genetic relatedness among 15 isolates of Trichoderma sp. was analyzed with six micro-satellite primers. ISSR profiles showed 83.7% genetic diversity among the isolates with the formation of four clusters. Analysis of dendrogram revealed that similarity coefficient ranged from 0.27 to 0.95. ITS-PCR of rDNA region with ...

  15. Identification of Bacitracin Produced by Local Isolate of Bacillus ...

    African Journals Online (AJOL)

    Bacillus licheniformis was isolated from soil of different house gardens. Diagnosis was performed according to Gram stain, motility, shape forming, aerobic condition and other tests. Bacitracin was primary identified after its activity was tested against some species of Gram positive and Gram negative bacteria. Identification ...

  16. Enumeration, isolation and identification of bacteria and fungi from ...

    African Journals Online (AJOL)

    Enumeration, isolation and identification of bacteria and fungi from soil contaminated with petroleum products using layer chicken droppings as an amendment The media used were nutrient agar for total heterotrophic bacterial count, potato dextrose agar for fungi count, serial dilution was carried out and the pour plate ...

  17. Isolement et identification de microorganismes indigènes de ...

    African Journals Online (AJOL)

    Isolement et identification de microorganismes indigènes de cacaoyères en côte d'ivoire et mise en évidence de leurs effets antagonistes vis-àvis de Phytophthora palmivora , agent de la pourriture brune des cabosses.

  18. Molecular identification of Coccidioides spp. in soil samples from Brazil

    Directory of Open Access Journals (Sweden)

    Filho Antônio D

    2011-05-01

    Full Text Available Abstract Background Since 1991 several outbreaks of acute coccidioidomycosis (CM were diagnosed in the semi-arid Northeast of Brazil, mainly related to disturbance of armadillo burrows caused by hunters while digging them for the capture of these animals. This activity causes dust contaminated with arthroconidia of Coccidioides posadasii, which, once inhaled, cause the mycosis. We report on the identification of C. posadasii in soil samples related to outbreaks of CM. Results Twenty four soil samples had their DNA extracted and subsequently submitted to a semi-nested PCR technique using specific primers. While only 6 (25% soil samples were positive for C. posadasii by mice inoculation, all (100% were positive by the molecular tool. Conclusion This methodology represents a simple, sensitive and specific molecular technique to determine the environmental distribution of Coccidioides spp. in endemic areas, but cannot distinguish the species. Moreover, it may be useful to identify culture isolates. Key-words: 1. Coccidioidomycosis. 2. Coccidioides spp. 3. C. posadasii. 4. Semi-arid. 5. Semi-nested PCR

  19. Molecular identification of Coccidioides spp. in soil samples from Brazil.

    Science.gov (United States)

    de Macêdo, Regina C L; Rosado, Alexandre S; da Mota, Fabio F; Cavalcante, Maria A S; Eulálio, Kelsen D; Filho, Antônio D; Martins, Liline M S; Lazéra, Márcia S; Wanke, Bodo

    2011-05-16

    Since 1991 several outbreaks of acute coccidioidomycosis (CM) were diagnosed in the semi-arid Northeast of Brazil, mainly related to disturbance of armadillo burrows caused by hunters while digging them for the capture of these animals. This activity causes dust contaminated with arthroconidia of Coccidioides posadasii, which, once inhaled, cause the mycosis. We report on the identification of C. posadasii in soil samples related to outbreaks of CM. Twenty four soil samples had their DNA extracted and subsequently submitted to a semi-nested PCR technique using specific primers. While only 6 (25%) soil samples were positive for C. posadasii by mice inoculation, all (100%) were positive by the molecular tool. This methodology represents a simple, sensitive and specific molecular technique to determine the environmental distribution of Coccidioides spp. in endemic areas, but cannot distinguish the species. Moreover, it may be useful to identify culture isolates. Key-words: 1. Coccidioidomycosis. 2. Coccidioides spp. 3. C. posadasii. 4. Semi-arid. 5. Semi-nested PCR.

  20. Identification of bacterial strains in viili by molecular taxonomy and ...

    African Journals Online (AJOL)

    Identification of bacterial strains in viili by molecular taxonomy and their synergistic effects on milk curd and exopolysaccharides production. T Chen, Q Tan, M Wang, S Xiong, S Jiang, Q Wu, S Li, C Luo, H Wei ...

  1. Molecular and pathological identification of feline coronavirus type I ...

    African Journals Online (AJOL)

    Molecular and pathological identification of feline coronavirus type I. Alazawy Amer, Arshad Siti-Suri, Hair-Bejo Mohd, Omar Abdul-Rahman, Tengku-Ibrahim Tengku-Azmi, Bande Faruku, Assumaidaee Ajwad ...

  2. Identification and Pathogenic Potential of Clinical Bacillus and Paenibacillus Isolates.

    Directory of Open Access Journals (Sweden)

    Francesco Celandroni

    Full Text Available The soil-related Bacillus and Paenibacillus species have increasingly been implicated in various human diseases. Nevertheless, their identification still poses problems in the clinical microbiology laboratory and, with the exception of Bacillus anthracis and Bacillus cereus, little is known on their pathogenicity for humans. In this study, we evaluated the use of matrix-assisted laser desorption-ionization time of flight mass spectrometry (MALDI-TOF MS in the identification of clinical isolates of these genera and conducted genotypic and phenotypic analyses to highlight specific virulence properties. Seventy-five clinical isolates were subjected to biochemical and MALDI-TOF MS identification. 16S rDNA sequencing and supplemental tests were used to solve any discrepancies or failures in the identification results. MALDI-TOF MS significantly outperformed classical biochemical testing for correct species identification and no misidentification was obtained. One third of the collected strains belonged to the B. cereus species, but also Bacillus pumilus and Bacillus subtilis were isolated at high rate. Antimicrobial susceptibility testing showed that all the B. cereus, B. licheniformis, B. simplex, B. mycoides, Paenibacillus glucanolyticus and Paenibacillus lautus isolates are resistant to penicillin. The evaluation of toxin/enzyme secretion, toxin-encoding genes, motility, and biofilm formation revealed that B. cereus displays the highest virulence potential. However, although generally considered nonpathogenic, most of the other species were shown to swim, swarm, produce biofilms, and secrete proteases that can have a role in bacterial virulence. In conclusion, MALDI-TOF MS appears useful for fast and accurate identification of Bacillus and Paenibacillus strains whose virulence properties make them of increasing clinical relevance.

  3. Identification and Pathogenic Potential of Clinical Bacillus and Paenibacillus Isolates.

    Science.gov (United States)

    Celandroni, Francesco; Salvetti, Sara; Gueye, Sokhna Aissatou; Mazzantini, Diletta; Lupetti, Antonella; Senesi, Sonia; Ghelardi, Emilia

    2016-01-01

    The soil-related Bacillus and Paenibacillus species have increasingly been implicated in various human diseases. Nevertheless, their identification still poses problems in the clinical microbiology laboratory and, with the exception of Bacillus anthracis and Bacillus cereus, little is known on their pathogenicity for humans. In this study, we evaluated the use of matrix-assisted laser desorption-ionization time of flight mass spectrometry (MALDI-TOF MS) in the identification of clinical isolates of these genera and conducted genotypic and phenotypic analyses to highlight specific virulence properties. Seventy-five clinical isolates were subjected to biochemical and MALDI-TOF MS identification. 16S rDNA sequencing and supplemental tests were used to solve any discrepancies or failures in the identification results. MALDI-TOF MS significantly outperformed classical biochemical testing for correct species identification and no misidentification was obtained. One third of the collected strains belonged to the B. cereus species, but also Bacillus pumilus and Bacillus subtilis were isolated at high rate. Antimicrobial susceptibility testing showed that all the B. cereus, B. licheniformis, B. simplex, B. mycoides, Paenibacillus glucanolyticus and Paenibacillus lautus isolates are resistant to penicillin. The evaluation of toxin/enzyme secretion, toxin-encoding genes, motility, and biofilm formation revealed that B. cereus displays the highest virulence potential. However, although generally considered nonpathogenic, most of the other species were shown to swim, swarm, produce biofilms, and secrete proteases that can have a role in bacterial virulence. In conclusion, MALDI-TOF MS appears useful for fast and accurate identification of Bacillus and Paenibacillus strains whose virulence properties make them of increasing clinical relevance.

  4. A PCR-based strategy for simple and rapid identification of rough presumptive Salmonella isolates

    DEFF Research Database (Denmark)

    Hoorfar, Jeffrey; Baggesen, Dorte Lau; Porting, P.H.

    1999-01-01

    The purpose of the present study was to investigate the application of ready-to-go Salmonella PCR tests, based on dry chemistry, for final identification of rough presumptive Salmonella isolates. The results were compared with two different biotyping methods performed at two different laboratorie...... (serogroup D). In conclusion, rough presumptive Salmonella isolates can be conveniently confirmed to the serogroup-revel, using the pre-mixed PCR tests. The system can be easily implemented in accredited laboratories with limited experience in molecular biology....

  5. Molecular characterization of Danish Cryptosporidium parvum isolates

    DEFF Research Database (Denmark)

    Enemark, Heidi L.; Ahrens, Peter; Juel, Cynthia Dawn

    2002-01-01

    The genetic polymorphism among 271 Danish Cryptosporidium isolates of human and animal origin was studied by partial amplification and sequencing of the Cryptosporidium oocyst wall protein (COWP) gene, the 18S rDNA, and a microsatellite locus.dagger Furthermore, the microsatellite locus was studied...... directly using fragment analysis. A comparative analysis of DNA sequences showed the presence of 3 different subgenotypes (C1, C2 and C3) in C. parvum isolates from Danish cattle, with prevalences of 16.7, 17.2 and 73.1% including 13 (7.0%) mixed infections. Subgenotype C1 was significantly more prevalent...... (P isolates of human origin the anthroponotic subgenotype H1 was identified, in addition to the zoonotic subgenotypes C1, C2, and C3. Of 44 human samples, 56.8% were anthroponotic, whereas 40.9% were zoonotic genotypes. One human isolate...

  6. Molecular identification of polydorid polychaetes (Annelida ...

    African Journals Online (AJOL)

    The early detection and correct identification of polydorid polychaete species is essential as they are often encountered as invasive alien pests in aquaculture facilities or the intertidal where they may modify the ecosystem. Accurate identification is, however, often hampered by high levels of morphological similarity among ...

  7. Identification of Isolates of Streptococcus canis Infecting Humans

    Science.gov (United States)

    Whatmore, Adrian M.; Engler, Kathryn H.; Gudmundsdottir, Gudny; Efstratiou, Androulla

    2001-01-01

    During a survey of Group G and C streptococcal infections of humans two epidemiologically unrelated Group G streptococcal isolates were identified, one from a case of bacteremia and one from a wound infection. These isolates were atypical among this sample in that the emm gene could not be amplified from them by PCR. Biochemical characterization identified the isolates as Streptococcus canis, an organism normally associated with animal hosts. The biochemical identification was confirmed by sequencing of the 16S rRNA gene from both isolates and comparison with sequences of the S. canis type strain and other related streptococci of animals and humans. Comparative sequencing of fragments of two other housekeeping genes, sodA and mutS, confirmed that the isolates are most closely related to S. canis. The identification of two isolates of S. canis from a relatively small sample set suggests that the practice of identifying streptococci only by the Lancefield serological group may result in underestimation of the presence of S. canis in the human population. PMID:11682560

  8. Isolation and molecular characterization of Cryptococcus species isolated from pigeon nests and Eucalyptus trees.

    Science.gov (United States)

    Kamari, A; Sepahvand, A; Mohammadi, R

    2017-06-01

    Cryptococcus species are pathogenic and non-pathogenic basidiomycete yeasts that are found widely in the environment. Based on phenotypic methods, this genus has many species; however, its taxonomy is presently being re-evaluated by modern techniques. The Cryptococcus species complex includes two sibling taxa of Cryptococcus neoformans and Cryptococcus gattii . We aimed to investigate the possible distribution of Cryptococcus species in pigeon nests and Eucalyptus trees in Ilam, Iran, using molecular techniques. Two hundred and seventy-four specimens were collected from pigeon nests and Eucalyptus trees during 2016-2017. All the specimens were sub-cultured on Sabouraud Glucose Agar with chloramphenicol and bird seed agar. For molecular identification, the ITS15.8SITS2 rDNA region was amplified using the first and fourth internal transcribed spacer (ITS1 and ITS4, respectively) primers. The purified products were applied for cycle sequencing reactions in forward direction with ITS1 primer. The obtained results were analyzed with Chromas 2.3. Thirty-three out of 186 cultures (17.7%) and 11 out of 88 cultures (12.5%) were positive among pigeon nest and Eucalyptus tree specimens, respectively. Cryptococcus albidus (17.2%), C. albidus var. kuetzingii (3.4%), C. adeliensis (3.4%), C. uzbekistanensis (3.4%), and C. neoformans var. grubii (3.4%) were isolated from pigeon nests, and Cryptococcus adeliensis (25%) was the only Cryptococcus species isolated from Eucalyptus trees. The presence of pigeons and Eucalyptus trees in the vicinity of some particular places such as rest homes and hospitals should be considered as a risk factor for the immunocompromised population.

  9. Molecular characterization of Cryptosporidium isolates from humans in Equatorial Guinea.

    Science.gov (United States)

    Blanco, María Alejandra; Iborra, Asunción; Vargas, Antonio; Nsie, Eugenia; Mbá, Luciano; Fuentes, Isabel

    2009-12-01

    The aim of the study was to perform a molecular characterization of clinical isolates of Cryptosporidium species from Equatorial Guinea. Standard laboratory methods were used to identify 35 cryptosporidiosis cases among 185 patients. PCR-RFLP successfully identified 34 Cryptosporidium species from these 35 cases, comprising C. parvum (52.9%), C. hominis (44.1%) and C. meleagridis (2.9%); over 90% of the species were isolated from HIV-positive patients. This is the first report of the molecular characterization of Cryptosporidium species isolated from humans in Equatorial Guinea and shows that zoonotic and anthroponotic transmission is present in this country.

  10. Molecular identification of pathogenic Fusarium species, the causal agents of tomato wilt in western Iran

    Directory of Open Access Journals (Sweden)

    Chehri Khosrow

    2016-04-01

    Full Text Available Fusarium species are causal agents of fungal diseases occurring frequently in numerous agriculturally important plants, including potato, garlic and are one of the common pathogens of tomato, causing root rot in the west part of Iran. Therefore, the objectives of this study were to isolate and identify disease-causing Fusarium species from infected tomatoes based on the morphological and molecular characteristics. Twenty-five isolates of Fusarium were obtained from infected root of tomato plants collected from the fields in different regions of western Iran. Based on morphological features, the strains were classified into four following Fusarium species: F. oxysporum, F. redolens, F. proliferatum and F. verticillioides. The phylogenetic trees based on tef1 and tub2 dataset clearly distinguished closely related species. All of the isolates were evaluated for their pathogenicity on healthy tomato seedlings in the greenhouse. This is the first report on molecular identification of Fusarium species isolated from tomato plants cultivated in Iran.

  11. Identification of nonlinear anti-vibration isolator properties

    Science.gov (United States)

    Mezghani, Fares; Del Rincón, Alfonso Fernández; Souf, Mohamed Amine Ben; Fernandez, Pablo García; Chaari, Fakher; Viadero Rueda, Fernando; Haddar, Mohamed

    2017-06-01

    Vibrations are classified among the major problems for engineering structures. Anti-vibration isolators are used to absorb vibration energy and minimise transmitted force which can cause damage. The isolator is modelled as a parallel combination of stiffness and damping elements. The main purpose of the model is to enable designers to predict the dynamic response of systems under different structural excitations and boundary conditions. A nonlinear identification method, discussed in this paper, aims to provide a tool for engineers to extract information about the nonlinear dynamic behaviour using measured data from experiments. The proposed method is demonstrated and validated with numerical simulations. Thus, this technique is applied to determine the nonlinear parameters of a commercial metal mesh isolator. Nonlinear stiffness and nonlinear damping can decrease with the increase in the amplitude of the base excitation. The softening behaviour of the mesh isolator is clearly visible.

  12. Identification and molecular characterisation of Colletotrichum ...

    African Journals Online (AJOL)

    The isolates were identified using PCR with speciesspecific primers, complemented by phylogenetic analysis of nucleotide sequences of the internal transcribed spacer region and partial glyceraldehyde-3-phosphate dehydrogenase gene. The pathogenicity of the isolated fungi was determined on detached matured fruits.

  13. Isolation, molecular and biochemical characterization of oil ...

    African Journals Online (AJOL)

    Biochemical and physiological characterization performed on the 34 bacterial isolates, revealed the presence of oil biodegrading bacterial genera and species of Pseudomonas Acidovorans, P. aerugi-nosa, P. alcaligenes, P. fluorescens, P. cepacia, P. mallei, P. maltophilia, P. oleovorans, P. putida, P. stutzeri P. vesicularis, ...

  14. Isolation and molecular characterisation of Mycobacterium bovis ...

    African Journals Online (AJOL)

    Background: Consumption of raw milk and unpasteurized dairy products is common in Tunisia where bovine tuberculosis remains enzootic. We herein investigated the frequency of M. bovis isolation from raw milk. Methods: Three hundred and six milk samples collected from 102 infected cows in different Tunisian regions ...

  15. Isolation, characterization and molecular weight determination of ...

    African Journals Online (AJOL)

    Collagen is a major structural protein of connective tissues. It can be used as a prosthetic biomaterial applicable to artificial skin, tendon ligaments and development collagen implants. In the present study, an attempt was made to isolate and characterize collagen from the marine sponge, Spirastrella inconstans. The total ...

  16. Isolation, biochemical and molecular characterization of 2 ...

    African Journals Online (AJOL)

    SERVER

    2007-12-03

    Dec 3, 2007 ... Research and Technology Applications, New-Burgelarab City, Alexandria, Egypt. Accepted 30 October, 2007 ... REP-PCR results strongly confirmed that the bacterial isolates from different Qatari soils produced different fingerprinting ..... dermatitis from homatropine and phenylephrine eyedrops. Contact.

  17. Molecular analysis of Aspergillus section Flavi isolated from Brazil nuts.

    Science.gov (United States)

    Gonçalves, Juliana Soares; Ferracin, Lara Munique; Carneiro Vieira, Maria Lucia; Iamanaka, Beatriz Thie; Taniwaki, Marta Hiromi; Pelegrinelli Fungaro, Maria Helena

    2012-04-01

    Brazil nuts are an important export market in its main producing countries, including Brazil, Bolivia, and Peru. Approximately 30,000 tons of Brazil nuts are harvested each year. However, substantial nut contamination by Aspergillus section Flavi occurs with subsequent production of aflatoxins. In our study, Aspergillus section Flavi were isolated from Brazil nuts (Bertholletia excelsa), and identified by morphological and molecular means. We obtained 241 isolates from nut samples, 41% positive for aflatoxin production. Eighty-one isolates were selected for molecular investigation. Pairwise genetic distances among isolates and phylogenetic relationships were assessed. The following Aspergillus species were identified: A. flavus, A. caelatus, A. nomius, A. tamarii, A. bombycis, and A. arachidicola. Additionally, molecular profiles indicated a high level of nucleotide variation within β-tubulin and calmodulin gene sequences associated with high genetic divergence from RAPD data. Among the 81 isolates analyzed by molecular means, three of them were phylogenetically distinct from all other isolates representing the six species of section Flavi. A putative novel species was identified based on molecular profiles.

  18. Molecular characterization of a Korean bovine parainfluenza virus type 3 isolate.

    Science.gov (United States)

    Oem, Jae-Ku; Lee, Eun-Yong; Lee, Kyoung-Ki; Kim, Seong-Hee; Lee, Myoung-Heon; Hyun, Bang-Hun

    2013-02-22

    Bovine parainfluenza virus type 3 (BPIV-3) was isolated from Korean native cattle that presented clinical signs of mild pneumonia. The complete genome of a representative isolate (12Q061) was sequenced. The newly identified strain, which was found to be distinct from the previously reported genotypes A (BPIV-3a) and B (BPIV-3b) and closely related to the Chinese strain SD0835, was tentatively classified as genotype C (BPIV-3c). Our results suggest a relationship between BPIV-3 genetic variation and the geographic location of its isolation. Identification of these new BPIV-3 genotypes may facilitate the development of improved diagnostic methods and vaccines. This is to our knowledge the first report of the identification and molecular characterization of BPIV-3 in Korea. Copyright © 2012 Elsevier B.V. All rights reserved.

  19. Identification and characterization of Daldinia eschscholtzii isolated from skin scrapings, nails, and blood

    Directory of Open Access Journals (Sweden)

    Kee Peng Ng

    2016-12-01

    Full Text Available Background Daldinia eschscholtzii is a filamentous wood-inhabiting endophyte commonly found in woody plants. Here, we report the identification and characterization of nine D. eschscholtzii isolates from skin scrapings, nail clippings, and blood. Methods The nine isolates were identified based on colony morphology, light microscopy, and internal transcribed spacer (ITS-based phylogeny. In vitro antifungal susceptibility of the fungal isolates was evaluated by the Etest to determine the minimum inhibitory concentration (MIC. Results The nine isolates examined were confirmed as D. eschscholtzii. They exhibited typical features of Daldinia sp. on Sabouraud Dextrose Agar, with white felty colonies and black-gray coloration on the reverse side. Septate hyphae, branching conidiophore with conidiogenous cells budding from its terminus, and nodulisporium-like conidiophores were observed under the microscope. Phylogenetic analysis revealed that the nine isolates were clustered within the D. eschscholtzii species complex. All the isolates exhibited low MICs against azole agents (voriconazole, posaconazole, itraconazole, and ketoconazole, as well as amphotericin B, with MIC of less than 1 µg/ml. Discussion Early and definitive identification of D. eschscholtzii is vital to reducing misuse of antimicrobial agents. Detailed morphological and molecular characterization as well as antifungal profiling of D. eschscholtzii provide the basis for future studies on its biology, pathogenicity, and medicinal potential.

  20. Isolation and molecular characterization of Acanthamoeba strains isolated from the oral cavity of immunosuppressed individuals in Tehran, Iran.

    Science.gov (United States)

    Memari, Fatemeh; Niyyati, Maryam; Lorenzo-Morales, Jacob; Jonaydi, Zaynab

    2016-09-01

    Acanthamoeba spp. is an opportunistic protozoan parasite which is the causative agent of granulomatous amoebic encephalitis (GAE) and Acanthamoeba keratitis (AK). GAE usually occurs in immunocompromised patients which in most cases is fatal. The present study was conducted to determine the genotypes of Acanthamoeba isolated from patients with compromised immunological status. For this purpose, 90 samples from the oral cavity of these individuals were collected in different hospitals of Tehran, Iran using sterile cotton swabs. Samples were cultured in 2% Non-Nutrient Agar (NNA) plates in order to check for the presence of amoebae. Identification of isolates was carried out using both morphological and molecular tools. The pathogenic potential of the obtained strains was assessed by performing osmo- and thermotolerance assays as previously described. Genotyping of the isolates was carried out by PCR/sequencing of the DF3 region of the 18S rDNA gene of Acanthamoeba. From the 90 collected samples, 11 (13.4%) were positive for Acanthamoeba genus. Molecular analysis revealed the presence of genotypes T3, T4 and T11, although most of the isolates belonged to genotype T4. Only 3 of the isolates genotyped as T4 were positive for the pathogenic potential assays. To this end if the immunological status is considered as one of the key factors for the development of GAE due to Acanthamoeba in the previous reported cases, individuals suffering from the conditions mentioned in this study should be considered as a high risk group of population in Iran and worldwide.

  1. Isolation, characterization and Molecular weight Determination of ...

    African Journals Online (AJOL)

    Salwee

    2013-07-10

    Jul 10, 2013 ... chromatography on DEAE-Sepharose. SDS-PAGE revealed molecular mass of 87 kDa. .... activity protein profile on the SDS-PAGE prior to anion exchange chromatography. The anion exchange ... of non-ionic detergent, Triton-X100, at a final concentration of 0.1% and also 0.2 and 5% ethanol and after 3 ...

  2. Signal Identification and Isolation Utilizing Radio Frequency Photonics

    Science.gov (United States)

    2017-09-01

    the drawings, specifications, or other data does not license the holder or any other person or corporation; or convey any rights or permission to...burden, to Department of Defense, Washington Headquarters Services, Directorate for Information Operations and Reports (0704-0188), 1215 Jefferson...identification and isolation are important to applications such as radio astronomy. Radio frequency (RF) photonics can provide solutions to these areas. Spectrum

  3. Isolation and identification of native lower fungi for polyunsaturated ...

    African Journals Online (AJOL)

    The minors were linoleic acid (C18:2n6) (8.26±0.59%) and γ-linoleic acid (GLA; C18:3n6) (5.48±0.08%). Besides the morphological characterization, taxonomic identification by the 636 bp-ITS region sequencing and phylogenetic analysis were performed. It was demonstrated that the fungal isolate NR06 was classified in ...

  4. Isolation, Characterization and Molecular weight determination of ...

    African Journals Online (AJOL)

    Enzyme purification to homogeneity was carried out by anion exchange chromatography on DEAE-Sepharose. SDS-PAGE revealed molecular mass of 87 kDa. Maximal activity of the enzymes was observed at 50°C at pH 4 and was stimulated by Ca2+, Co2+, Mg2+ (test at 10 Mm each) and inhibited by Fe2+. Ethanol at an ...

  5. Identification and isolation of Genotype-I Japanese Encephalitis virus from encephalitis patients

    Directory of Open Access Journals (Sweden)

    Gao Xiaoyan

    2010-11-01

    Full Text Available Abstract Historically, Japanese Encephalitis virus (JEV genotype III (GIII has been responsible for human diseases. In recent years, JEV genotype I (GI has been isolated from mosquitoes collected in numerous countries, but has not been isolated from patients with encephalitis. In this study, we report recovery of JEV GI live virus and identification of JEV GI RNA from cerebrospinal fluid (CSF of encephalitis patients in JE endemic areas of China. Whole-genome sequencing and molecular phylogenetic analysis of the JEV isolate from the CSF samples was performed. The isolate in this study is highly similar to other JEV GI strains which isolated from mosquitoes at both the nucleotide and deduced amino acid levels. Phylogenetic analysis based on the genomic sequence showed that the isolate belongs to JEV GI, which is consistent with the phylogenetic analysis based on the pre-membrane (PrM and Glycoprotein genes. As a conclusion, this is the first time to isolate JEV GI strain from CSF samples of encephalitis patients, so continuous survey and evaluate the infectivity and pathogenecity of JEV GI strains are necessary, especially for the JEV GI strains from encephalitis patients. With respect to the latter, because all current JEV vaccines (live and inactivated are derived from JEV GIII strains, future studies should be aimed at investigating and monitoring cross-protection of the human JEV GI isolates against widely used JEV vaccines.

  6. Morphological and molecular identification of phytophthora species from maple trees in Serbia

    Directory of Open Access Journals (Sweden)

    Milenković Ivan

    2014-01-01

    Full Text Available The paper presents the results of the study performed with aims to determine the presence and diversity of Phytophthora species on maple trees in Serbia. Due to high aggressiveness and their multicyclic nature, presence of these pathogens is posing significant threat to forestry and biodiversity. In total, 29 samples of water, soil and tissues were taken from 10 different localities, and six different maple hosts were tested. After the isolation tests, 17 samples from five different maple hosts were positive for the presence of Phytophthora spp., and 31 isolates were obtained. After the detailed morphological and physiological classification, four distinct groups of isolates were separated. DNA was extracted from selected representative isolates and molecular identification with sequencing of ITS region was performed. Used ITS4 and ITS6 primers successfully amplified the genomic DNA of chosen isolates and morphological identification of obtained isolates was confirmed after the sequencing. Four different Phytophthora species were detected, including P. cactorum, P. gonapodyides, P. plurivora and P. lacustris. The most common isolated species was homothallic, and with very variable and semipapillate sporangia, P. plurivora with 22 obtained isolates. This is the first report of P. plurivora and P. gonapodyides on A. campestre, P. plurivora and P. lacustris on Acer heldreichii and first report of P. lacustris on A. pseudoplatanus and A. tataricum in Serbia. [Projekat Ministarstva nauke Republike Srbije, br. TR 37008

  7. [Research progress on molecular identification and biologic behavior of Taenia saginata in Western China].

    Science.gov (United States)

    Bao, Huai-En; Mou, Rong

    2009-12-01

    This article reviews the epidemiological investigation of taeniasis saginata in 10 counties of 7 provinces/ autonomous regions in western China. The morphological observation of adult worms, molecular identification of 10 isolates of the worms, experimental infection on pigs and cattle with Taenia saginata and T. asiatica, observation on development and biology behavior of cysticercus, and pathological changes in the intermediate host pig and cattle revealed that T. asiatica is a new species, instead of a subspecies of T. saginata.

  8. Isolation and identification of marine fish tumour (odontoma associated bacteria

    Directory of Open Access Journals (Sweden)

    Ramalingam Vijayakumar

    2015-09-01

    Full Text Available Objective: To identify fish tumour associated bacteria. Methods: The marine fish Sphyraena jello with odontoma was collected from in Tamil Nadu (Southeast India, and tumour associated bacteria were isolated. Then the isolated bacteria were identified based on molecular characters. Results: A total of 4 different bacterial species were isolated from tumour tissue. The bacterial species were Bacillus sp., Pontibacter sp., Burkholderia sp. and Macrococcus sp., and the sequences were submitted in DNA Data Bank of Japan with accession numbers of AB859240, AB859241, AB859242 and AB859243 respectively. Conclusions: Four different bacterial species were isolated from Sphyraena jello, but the role of bacteria within tumour needs to be further investigated.

  9. Molecular and antimicrobial susceptibility characterization of Globicatella sulfidifaciens isolated from sow's urinary tract infection.

    Science.gov (United States)

    Matajira, Carlos E C; Moreno, Luisa Z; Gomes, Vasco T M; Silva, Ana Paula S; Mesquita, Renan E; Amigo, Cristina R; Christ, Ana Paula G; Sato, Maria Inês Z; Moreno, Andrea M

    2017-12-01

    The Globicatella genus comprises Gram-positive, facultative anaerobic, α-hemolytic and catalase negative cocci morphologically and phenotypically very similar to Streptococcus and Aerococcus genus which can lead to misidentification and underestimation of this pathogen. Globicatella species have already been isolated from human and animals with heart and brain disorders. Their clinical relevance in animals, and its zoonotic potential, remains unknown due to the difficulty in their identification. To present the isolation, phenotypic and molecular characterization of G. sulfidifaciens from urinary tract infection in sows. Urine samples from 140 sows of two swine herds located in São Paulo State (Brazil) yielded the isolation of three presumptive G. sulfidifaciens strains. Identification and species confirmation were done by MALDI-TOF MS and 16S rRNA sequencing. Strains were further characterized by single enzyme amplified fragments length polymorphism (SE-AFLP) and broth microdilution techniques. All three isolates were confirmed as G. sulfidifaciens. The SE-AFLP genotyping resulted in distinct fingerprint patterns for each strain. All isolates presented high MIC values to tetracycline, sulphonamides, aminoglycosides and tylosin tartrate, which present high usage in human and animal medicine. Globicatella sulfidifaciens could be related to sporadic urinary tract infections in swine and appear to present alarming antimicrobial susceptibility profile. It is necessary to differentiate Streptococcus-like microorganisms in routine laboratory diagnostics for the correct identification of underestimated species potentially pathogenic to animals.

  10. Phenotypic and molecular characterization of Malassezia japonica isolated from psoriasis vulgaris patients.

    Science.gov (United States)

    Honnavar, Prasanna; Chakrabarti, Arunaloke; Dogra, Sunil; Handa, Sanjeev; Rudramurthy, Shivaprakash M

    2015-03-01

    Malassezia species, which are skin colonizers, are being debated as to their pathogenic role in various cutaneous diseases. Species identification of Malassezia is important as particular species have been implicated in or associated with specific diseases. Malassezia japonica, a relatively newly described species, has not been completely characterized owing to the rarity of its isolation. In the present study we describe phenotypic and molecular characterization of six M. japonica strains isolated from patients with psoriasis vulgaris. In contrast to the physiological and biochemical properties of the M. japonica type strain, CBS9348, all our isolates assimilated Tween 20 and showed positive β-glucosidase activity, and the Cremophor EL utilization test was negative. However, the sequences of the D1/D2 region of rDNA, ITS2 and IGS1 regions of all our isolates clustered with the type strain of M. japonica. © 2015 The Authors.

  11. Isolation, Identification, and Evaluation of Novel Probiotic Strains Isolated from Feces of Breast-Fed Infants.

    Science.gov (United States)

    Panya, Marutpong; Lulitanond, Viraphong; Rattanachaikunsopon, Pongsak; Srivoramas, Thanyakarn; Chaiwong, Tarinee

    2016-01-01

    To isolate, identify, and evaluate the probiotic properties of lactic acid bacteria (LAB) isolated from the feces of breast-fed infants. The probiotic tests included investigation of hemolysis activity, survival in simulated gastrointestinal tract conditions (acid and bile salt tolerance), susceptibility to antibiotics, and ability to inhibit selected bacterial pathogens (Escherichia coli O157:H7, Vibrio cholerae and Salmonella enterica subsp enterica serovar Typhimurium). The bacterial species identification was performed by both carbohydrate utilization and partial 16S ribosomal RNA sequencing. Five of fifty LAB isolates (UBU-03, UBU-06, UBU-09, UBU-34, and UBU-37) showed good probiotic properties. These five isolates showed non-hemolysis type (gamma-hemolysis), susceptibility to all antibiotics tested except for vancomycin, ability to survive in the simulated gastrointestinal conditions of both acid and bile salt solution, and ability to inhibit growth of E. coli O157: H7 and V. cholerae. Bacterial species identification revealed that all five isolates were firmly identified as Lactobacillus rhamnosus species. The L. rhamnosus strains that were isolated and characterized in this study could be considered as probiotic strains, and then used for further probiotic characterization in human cell cultures or animal models.

  12. Molecular Identification of Methicillin-Resistant Staphylococcus ...

    African Journals Online (AJOL)

    Antimicrobial resistance has become a great public health problem worldwide and multi-drug resistant Staphylococcus aureus has been widely reported. Methods: The presence or absence of methicillin resistance gene (mecA) in 48 clinical wound isolates of S. aureus was examined by the polymerase chain reaction ...

  13. Molecular identification of phosphate solubilizing bacterium ...

    African Journals Online (AJOL)

    A phosphate solubilizing bacterium was isolated from the rhizosphere soil of upland rice and identified by 16S rRNA gene sequencing. The gene sequence showed 99% homology with Alcaligenes faecalis. Based on the gene sequence homology, it was identified as A. faecalis. Interaction effect of this bacterium on growth ...

  14. Molecular species identification and population genetics of ...

    African Journals Online (AJOL)

    Molecular genetic techniques, such as DNA barcoding and genotyping, are increasingly being used to assist with the conservation and management of chondrichthyans worldwide. Southern Africa is a shark biodiversity hotspot, with a large number of endemic species. According to the IUCN Red List, a quarter of South ...

  15. Molecular identification versus local people's information for ...

    African Journals Online (AJOL)

    The species identity was verified in 118 bushmeat samples through molecular sequencing of the cytochrome oxidase subunit I (COI) and phylogenetic assignments to established reference sequences of the respective species. The species diversity among the bushmeat samples was high with 15 identified species ...

  16. Molecular Typing of Salmonella Isolates in Poultry by Pulsed-Field Gel Electrophoresis in Iran

    Directory of Open Access Journals (Sweden)

    Narges Golab

    2014-11-01

    Full Text Available Background: Salmonella is one of the most widespread zoonotic enter pathogenic microorganisms found in the global food chain. Poultryand Poultry products have been identified as one of the important foodborne sources of Salmonella. Pulsed-Field Gel Electrophoresis (PFGE is a gold standard typing method for identification of Salmonella isolates during outbreaks and epidemiological investigations. Objectives: The aim of this study was to carry out molecular typing of Salmonella enterica spp. by PFGE technique. Materials and Methods: All 47 Salmonella isolates were serotyped and then subjected to PFGE. Total isolates were analyzed by means of the molecular technique XbaI PFGE. Results: In the current work, PFGE and serotyping were used to subtype 47 Salmonella isolates belonging to 22 different serotypes and derived from poultry. Thirty-nine PFGE patterns out of 47 isolates were obtained. The Discrimination Index (DI by serotyping (0.93 was lower than PFGE (DI = 0.99. Conclusions: In conclusion, molecular methods such as PFGE can be used for epidemiological characterization of Salmonella serotypes.

  17. Contemporary nucleic acid-based molecular techniques for detection, identification, and characterization of Bifidobacterium.

    Science.gov (United States)

    Mianzhi, Yao; Shah, Nagendra P

    2017-03-24

    Bifidobacteria are one of the most important bacterial groups found in the gastrointestinal tract of humans. Medical and food industry researchers have focused on bifidobacteria because of their health-promoting properties. Researchers have historically relied on classic phenotypic approaches (culture and biochemical tests) for detection and identification of bifidobacteria. Those approaches still have values for the identification and detection of some bifidobacterial species, but they are often labor-intensive and time-consuming and can be problematic in differentiating closely related species. Rapid, accurate, and reliable methods for detection, identification, and characterization of bifidobacteria in a mixed bacterial population have become a major challenge. The advent of nucleic acid-based molecular techniques has significantly advanced isolation and detection of bifidobacteria. Diverse nucleic acid-based molecular techniques have been employed, including hybridization, target amplification, and fingerprinting. Certain techniques enable the detection, characterization, and identification at genus-, species-, and strains-levels, whereas others allow typing of species or strains of bifidobacteria. In this review, an overview of methodological principle, technique complexity, and application of various nucleic acid-based molecular techniques for detection, identification, and characterization of bifidobacteria is presented. Advantages and limitations of each technique are discussed, and significant findings based on particular techniques are also highlighted.

  18. Isolation and identification of iron ore-solubilising fungus

    Directory of Open Access Journals (Sweden)

    Damase Khasa

    2010-09-01

    Full Text Available Potential mineral-solubilising fungi were successfully isolated from the surfaces of iron ore minerals. Four isolates were obtained and identified by molecular and phylogenetic methods as close relatives of three different genera, namely Penicillium (for isolate FO, Alternaria (for isolates SFC2 and KFC1 and Epicoccum (for isolate SFC2B. The use of tricalcium phosphate (Ca3(PO42in phosphate-solubilising experiments confirmed isolate FO as the only phosphate solubiliser among the isolated fungi. The bioleaching capabilities of both the fungus and its spent liquid medium were tested and compared using two types of iron ore materials, conglomerate and shale, from the Sishen Iron Ore Mine as sources of potassium (K and phosphorus (P. The spent liquid medium removed more K (a maximum of 32.94% removal, from conglomerate, than the fungus (a maximum of 21.36% removal, from shale. However, the fungus removed more P (a maximum of 58.33% removal, from conglomerate than the spent liquid medium (a maximum of 29.25% removal, from conglomerate. The results also indicated a potential relationship between the removal of K or P and the production of organic acids by the fungus. A high production of gluconic acid could be related to the ability of the fungus to reduce K and P. Acetic, citric and maleic acids were also produced by the fungus, but in lower quantities. In addition, particle size and iron ore type were also shown to have significant effects on the removal of potassium and phosphorus from the iron ore minerals. We therefore conclude that the spent liquid medium from the fungal isolate FO can potentially be used for biobeneficiation of iron ore minerals.

  19. Isolation and identification of Mycobacterium avium subspecies silvaticum from a horse.

    Science.gov (United States)

    Chiers, Koen; Deschaght, Pieter; De Baere, Thierry; Dabrowski, Slawomir; Kotlowski, Roman; De Clercq, Dominique; Ducatelle, Richard; Vaneechoutte, Mario

    2012-07-01

    Routine cultivation methods are able to distinguish between isolates of the Mycobacterium avium and the Mycobacterium tuberculosis complex. However, molecular tools are needed to further identify the several subspecies in the M. avium complex, especially for the subspecies avium and silvaticum. A rapid technique using HhaI restriction digestion of a 349 bp amplification product of the 85B antigen (α-antigen) gene was used for the identification of M. avium subsp. silvaticum in a three-year-old gelding presenting with caseous, necrotizing, granulomatous lesions. The result was confirmed by sequencing of the 85B antigen gene. Copyright © 2012 Elsevier Ltd. All rights reserved.

  20. Molecular screening and isolation of Newcastle disease virus from ...

    African Journals Online (AJOL)

    Molecular screening and isolation of Newcastle disease virus from live poultry markets and chickens from commercial poultry farms in Zaria, Kaduna state, Nigeria. ... The PDF file you selected should load here if your Web browser has a PDF reader plug-in installed (for example, a recent version of Adobe Acrobat Reader).

  1. Molecular Microbial Analysis of Lactobacillus Strains Isolated from the Gut of Calves for Potential Probiotic Use

    Directory of Open Access Journals (Sweden)

    Lorena P. Soto

    2010-01-01

    Full Text Available The intestinal microbiota has an influence on the growth and health status of the hosts. This is of particular interest in animals reared using intensive farming practices. Hence, it is necessary to know more about complexity of the beneficial intestinal microbiota. The use of molecular methods has revolutionized microbial identification by improving its quality and effectiveness. The specific aim of the study was to analyze predominant species of Lactobacillus in intestinal microbial ecosystem of young calves. Forty-two lactic acid bacteria (LAB isolated from intestinal tract of young calves were characterized by: Amplified Ribosomal DNA Restriction Analysis (ARDRA, by using Hae III, Msp I, and Hinf I restriction enzymes, and 16S rDNA gene sequencing. ARDRA screening revealed nine unique patterns among 42 isolates, with the same pattern for 29 of the isolates. Gene fragments of 16S rDNA of 19 strains representing different patterns were sequenced to confirm the identification of these species. These results confirmed that ARDRA is a good tool for identification and discrimination of bacterial species isolated from complex ecosystem and between closely related groups. This paper provides information about the LAB species predominant in intestinal tract of young calves that could provide beneficial effects when administered as probiotic.

  2. Isolation and molecular characterization of Acanthamoeba from patients with keratitis in Spain.

    Science.gov (United States)

    Martín-Pérez, T; Criado-Fornelio, A; Martínez, J; Blanco, M A; Fuentes, I; Pérez-Serrano, J

    2017-10-01

    In order to improve our knowledge on the epidemiology of amoebic keratitis, as well as the identification of Acanthamoeba isolates, we have isolated Acanthamoeba spp. from five symptomatic patients in Spain in the present study. All isolates were grown in axenic liquid medium, with only one exception. The morphology of these isolates were characterized by optical and scanning electron microscopy. Their structural features corresponded to those of amphizoic amoebae (namely Acanthamoeba spp.). The molecular characterization of the five Acanthamoeba isolates yielded six sequences. Almost complete 18S rRNA gene sequences (>2000bp) were obtained from three isolates and partial sequences (∼1500bp) from the other two. A robust phylogenetic analysis based on the almost complete 18S rRNA sequence showed that four isolates belonged to the T4 genotype and the other one to the T3 genotype. However, all isolates were identified as T4 genotype using the ASA.S1 fragment. As previously suggested by other researchers, only a robust phylogenetic approach may be helpful in identifying Acanthamoeba genotypes. In addition, new data on the phylogenetic relationships among the Acanthamoeba genotypes is provided and discussed. Copyright © 2017 Elsevier GmbH. All rights reserved.

  3. Metabolite identification and molecular fingerprint prediction through machine learning.

    Science.gov (United States)

    Heinonen, Markus; Shen, Huibin; Zamboni, Nicola; Rousu, Juho

    2012-09-15

    Metabolite identification from tandem mass spectra is an important problem in metabolomics, underpinning subsequent metabolic modelling and network analysis. Yet, currently this task requires matching the observed spectrum against a database of reference spectra originating from similar equipment and closely matching operating parameters, a condition that is rarely satisfied in public repositories. Furthermore, the computational support for identification of molecules not present in reference databases is lacking. Recent efforts in assembling large public mass spectral databases such as MassBank have opened the door for the development of a new genre of metabolite identification methods. We introduce a novel framework for prediction of molecular characteristics and identification of metabolites from tandem mass spectra using machine learning with the support vector machine. Our approach is to first predict a large set of molecular properties of the unknown metabolite from salient tandem mass spectral signals, and in the second step to use the predicted properties for matching against large molecule databases, such as PubChem. We demonstrate that several molecular properties can be predicted to high accuracy and that they are useful in de novo metabolite identification, where the reference database does not contain any spectra of the same molecule. An Matlab/Python package of the FingerID tool is freely available on the web at http://www.sourceforge.net/p/fingerid. markus.heinonen@cs.helsinki.fi.

  4. Molecular sex identification of painted storks (Mycteria leucocephala ...

    Indian Academy of Sciences (India)

    ONLINE RESOURCES. Molecular sex identification of painted storks (Mycteria leucocephala): using FTA cards, horizontal PAGE and quick silver staining. ELSIE Y. S. YEE1∗, ZAINAL ZAHARI ZAINUDDIN2, AHMAD ISMAIL3, CHEE KONG YAP3 and SOON GUAN TAN1. 1Faculty of Biotechnology and Biomolecular Science ...

  5. Molecular approaches for the identification and characterisation of ...

    African Journals Online (AJOL)

    Molecular approaches for the identification and characterisation of oenological lactic acid bacteria. ... Frequently Asked Questions about PDFs. Alternatively, you can download the PDF file directly to your computer, from where it can be opened using a PDF reader. To download the PDF, click the Download link above.

  6. Development of molecular map and identification of QTLs linked to ...

    Indian Academy of Sciences (India)

    Development of molecular map and identification of QTLs linked to. Fusarium wilt resistance in chickpea. Pavankumar Jingade and R. L. Ravikumar. J. Genet. 94, 723-729. Table 1. List of SSR markers used for screening parental polymorphism in the present study. SSR ID. Forward primers (5'-3'). Reverse primers (5'-3').

  7. Molecular study for the sex identification in Japanese quails ...

    African Journals Online (AJOL)

    In many birds' species such as Japanese quail, sex determination in young and many adult birds is very difficult. Nowadays, sex identification of animals throughout their lives is possible by molecular genetic techniques such as polymerase chain reaction (PCR). The aim of this study was to determine the sex of Japanese ...

  8. Molecular sex identification of painted storks (Mycteria leucocephala ...

    Indian Academy of Sciences (India)

    [Yee E. Y. S., Zainuddin Z. Z., Ismail A., Yap C. K. and Tan S. G. 2013 Molecular sex identification of painted storks. (Mycteria leucocephala): ... or blood collected in anticoagulant tubes as source of DNA. Here, we used dried blood .... of Higher Education, Malaysia and partially by Malaysian Wildlife. Department. Thanks to ...

  9. Molecular cytogenetic identification of a novel dwarf wheat line with ...

    Indian Academy of Sciences (India)

    2012-01-08

    Jan 8, 2012 ... 2State Key Laboratory of Plant Cell and Chromosome Engineering, Institute of. Genetics and ... [Chen G, Zheng Q, Bao Y, Liu S, Wang H and Li X 2012 Molecular cytogenetic identification of a novel dwarf wheat line with introgressed. Thinopyrum ..... tance to diseases and pests: Current status. Eyphytica 91 ...

  10. Molecular identification and genotyping of Microsporidia in selected hosts

    Czech Academy of Sciences Publication Activity Database

    Valenčáková, A.; Balent, P.; Ravaszová, P.; Horák, Aleš; Oborník, Miroslav; Halanová, M.; Malčeková, B.; Novotný, F.; Goldová, M.

    2012-01-01

    Roč. 110, č. 2 (2012), s. 689-693 ISSN 0932-0113 Institutional research plan: CEZ:AV0Z60220518 Keywords : ENCEPHALITOZOON-CUNICULI * RIBOSOMAL-RNA * SPECIES IDENTIFICATION * AIDS PATIENTS * PET RABBITS Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.852, year: 2012

  11. Molecular identification and food source inference of constructive ...

    African Journals Online (AJOL)

    user

    2011-01-10

    Jan 10, 2011 ... Ophiocordyceps sinensis is a precious and effectual Traditional Chinese Medicine, native to Tibetan ... primer design and correlation analysis and so on. In consequence, molecular identification system of constructive plants native to the O. sinensis habitat was established, and then feeding food habit of.

  12. Sequencing identification and antimicrobial resistance of Campylobacter spp. isolated from poultry and other animal sources

    Directory of Open Access Journals (Sweden)

    Ioanna eMarinou

    2012-02-01

    Full Text Available Background:Campylobacter spp. are together with Salmonella spp. the leading causes of human bacterial gastroenteritis worldwide. The most commonly isolated species in humans are Campylobacter jejuni and C. coli The isolation, identification and antimicrobial resistance of Campylobacter spp. from poultry and raw meat from slaughterhouses, has been investigated for the first time in Greece. During the period from August 2005 to November 2008 a total of 1080 samples were collected: a 830 fecal samples from five poultry farms, b 150 cecal samples from chicken carcasses in a slaughterhouse and c 100 fecal samples from one pig farm near the region of Attica. The identification of the isolates was performed with conventional, as well as with and molecular methods.Results: Sixteen Campylobacter strains were isolated, all from the poultry farms. None of the strains was identified as C. jejuni. Antimicrobial susceptibility to six antimicrobials was performed and all the strains were susceptible to ciprofloxacin, amoxicillin-clavulanic acid and gentamicin. Thirteen out of 14 C.coli were resistant to erythromycin and all C.coli strains were resistant to ampicillin. Conclusions:Our results emphasize the need for a surveillance and monitoring system with respect to the prevalence and antimicrobial resistance of Campylobacter in poultry, as well as for the use of antimicrobials in veterinary medicine in Greece.

  13. Molecular characterization of Rhodococcus equi isolates in equines

    Science.gov (United States)

    Javed, Rabyia; Taku, A. K.; Sharma, R. K.; Badroo, Gulzaar Ahmed

    2017-01-01

    Aim: The aim was to determine the occurrence of Rhodococcus equi in equines and their environment in Jammu (R.S. Pura, Katra), molecular characterization and to determine the antibiotic resistance pattern of R. equi. Materials and Methods: A total of 96 nasopharyngeal swab samples were collected from equines. The organism was isolated on Columbia nalidixic acid agar containing 5% sheep blood as well as on sheep blood agar and was later confirmed by cultural characteristics and biochemical tests. Molecular detection of R. equi isolates was done by 16S rRNA gene amplification followed by virulence associated protein A (Vap A) gene amplification. Antibiogram was performed against five antibiotics, viz., amoxicillin, penicillin G, streptomycin, rifampicin, and methicillin. Results: During the study, 9 R. equi isolates were identified on the basis of cultural and biochemical tests. In the polymerase chain reaction based detection, 3 among the 9 rhodococcal isolates were positive for species-specific 16S rRNA gene and revealed amplicon of 450 bp for confirmation of 16S rRNA gene. None of the sample was found positive for Vap A gene. In antibiogram, R. equi isolates were found sensitive for amoxicillin, while some isolates were also found resistant to the most conventional antibiotic penicillin G. Conclusion: From this study, it was concluded that R. equi infection is prevalent in equines in Jammu region of India and the indiscriminate use of the antibiotics is leading toward the development of resistant strains of R. equi. PMID:28246441

  14. Molecular mechanism of fluoroquinolones resistance in Mycoplasma hominis clinical isolates

    Directory of Open Access Journals (Sweden)

    Meng Dong-Ya

    2014-01-01

    Full Text Available To evaluate the molecular mechanism of fluoroquinolones resistance in Mycoplasma hominis (MH clinical strains isolated from urogenital specimens. 15 MH clinical isolates with different phenotypes of resistance to fluoroquinolones antibiotics were screened for mutations in the quinolone resistance-determining regions (QRDRs of DNA gyrase (gyrA and gyrB and topoisomerase IV (parC and parE in comparison with the reference strain PG21, which is susceptible to fluoroquinolones antibiotics. 15 MH isolates with three kinds of quinolone resistance phenotypes were obtained. Thirteen out of these quinolone-resistant isolates were found to carry nucleotide substitutions in either gyrA or parC. There were no alterations in gyrB and no mutations were found in the isolates with a phenotype of resistance to Ofloxacin (OFX, intermediate resistant to Levofloxacin (LVX and Sparfloxacin (SFX, and those susceptible to all three tested antibiotics. The molecular mechanism of fluoroquinolone resistance in clinical isolates of MH was reported in this study. The single amino acid mutation in ParC of MH may relate to the resistance to OFX and LVX and the high-level resistance to fluoroquinolones for MH is likely associated with mutations in both DNA gyrase and the ParC subunit of topoisomerase IV.

  15. Molecular mechanism of fluoroquinolones resistance in Mycoplasma hominis clinical isolates.

    Science.gov (United States)

    Meng, Dong-Ya; Sun, Chang-Jian; Yu, Jing-Bo; Ma, Jun; Xue, Wen-Cheng

    2014-01-01

    To evaluate the molecular mechanism of fluoroquinolones resistance in Mycoplasma hominis (MH) clinical strains isolated from urogenital specimens. 15 MH clinical isolates with different phenotypes of resistance to fluoroquinolones antibiotics were screened for mutations in the quinolone resistance-determining regions (QRDRs) of DNA gyrase (gyrA and gyrB) and topoisomerase IV (parC and parE) in comparison with the reference strain PG21, which is susceptible to fluoroquinolones antibiotics. 15 MH isolates with three kinds of quinolone resistance phenotypes were obtained. Thirteen out of these quinolone-resistant isolates were found to carry nucleotide substitutions in either gyrA or parC. There were no alterations in gyrB and no mutations were found in the isolates with a phenotype of resistance to Ofloxacin (OFX), intermediate resistant to Levofloxacin (LVX) and Sparfloxacin (SFX), and those susceptible to all three tested antibiotics. The molecular mechanism of fluoroquinolone resistance in clinical isolates of MH was reported in this study. The single amino acid mutation in ParC of MH may relate to the resistance to OFX and LVX and the high-level resistance to fluoroquinolones for MH is likely associated with mutations in both DNA gyrase and the ParC subunit of topoisomerase IV.

  16. Molecular characterization and antimicrobial susceptibility of nasal Staphylococcus aureus isolates from a Chinese medical college campus.

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    Jimei Du

    Full Text Available Staphylococcus aureus colonization and infection occur more commonly among persons living or working in crowded conditions, but characterization of S. aureus colonization within medical communities in China is lacking. A total of 144 (15.4%, 144/935 S. aureus isolates, including 28 (3.0%, 28/935 MRSA isolates, were recovered from the nares of 935 healthy human volunteers residing on a Chinese medical college campus. All S. aureus isolates were susceptible to vancomycin, quinupristin/dalfopristin and linezolid but the majority were resistant to penicillin (96.5%, ampicillin/sulbactam (83.3% and trimethoprim/sulfamethoxazole (93.1%. 82%, (23/28 of the MRSA isolates and 66% (77/116 of the MSSA isolates were resistant to multiple antibiotics, and 3 MRSA isolates were resistant to mupirocin--an agent commonly used for nasal decolonization. 16 different sequence types (STs, as well as SCCmec genes II, III, IVd, and V, were represented among MRSA isolates. We also identified, for the first time, two novel STs (ST1778 and ST1779 and 5 novel spa types for MRSA. MRSA isolates were distributed in different sporadic clones, and ST59-MRSA-VId- t437 was found within 3 MRSA isolates. Moreover, one isolate with multidrug resistance belonging to ST398-MRSA-V- t571 associated with animal infections was identified, and 3 isolates distributed in three different clones harbored PVL genes. Collectively, these data indicate a high prevalence of nasal MRSA carriage and molecular heterogeneity of S. aureus isolates among persons residing on a Chinese medical college campus. Identification of epidemic MRSA clones associated with community infection supports the need for more effective infection control measures to reduce nasal carriage and prevent dissemination of MRSA to hospitalized patients and health care workers in this community.

  17. Systematic internal transcribed spacer sequence analysis for identification of clinical mold isolates in diagnostic mycology: a 5-year study.

    Science.gov (United States)

    Ciardo, Diana E; Lucke, Katja; Imhof, Alex; Bloemberg, Guido V; Böttger, Erik C

    2010-08-01

    The implementation of internal transcribed spacer (ITS) sequencing for routine identification of molds in the diagnostic mycology laboratory was analyzed in a 5-year study. All mold isolates (n = 6,900) recovered in our laboratory from 2005 to 2009 were included in this study. According to a defined work flow, which in addition to troublesome phenotypic identification takes clinical relevance into account, 233 isolates were subjected to ITS sequence analysis. Sequencing resulted in successful identification for 78.6% of the analyzed isolates (57.1% at species level, 21.5% at genus level). In comparison, extended in-depth phenotypic characterization of the isolates subjected to sequencing achieved taxonomic assignment for 47.6% of these, with a mere 13.3% at species level. Optimization of DNA extraction further improved the efficacy of molecular identification. This study is the first of its kind to testify to the systematic implementation of sequence-based identification procedures in the routine workup of mold isolates in the diagnostic mycology laboratory.

  18. Systematic Internal Transcribed Spacer Sequence Analysis for Identification of Clinical Mold Isolates in Diagnostic Mycology: a 5-Year Study▿ †

    Science.gov (United States)

    Ciardo, Diana E.; Lucke, Katja; Imhof, Alex; Bloemberg, Guido V.; Böttger, Erik C.

    2010-01-01

    The implementation of internal transcribed spacer (ITS) sequencing for routine identification of molds in the diagnostic mycology laboratory was analyzed in a 5-year study. All mold isolates (n = 6,900) recovered in our laboratory from 2005 to 2009 were included in this study. According to a defined work flow, which in addition to troublesome phenotypic identification takes clinical relevance into account, 233 isolates were subjected to ITS sequence analysis. Sequencing resulted in successful identification for 78.6% of the analyzed isolates (57.1% at species level, 21.5% at genus level). In comparison, extended in-depth phenotypic characterization of the isolates subjected to sequencing achieved taxonomic assignment for 47.6% of these, with a mere 13.3% at species level. Optimization of DNA extraction further improved the efficacy of molecular identification. This study is the first of its kind to testify to the systematic implementation of sequence-based identification procedures in the routine workup of mold isolates in the diagnostic mycology laboratory. PMID:20573873

  19. Selective Isolation and Rapid Identification of Members of the Genus Micromonospora▿ †

    OpenAIRE

    Qiu, Danheng; Ruan, Jisheng; Huang, Ying

    2008-01-01

    Improved methods for selective isolation of diverse actinomycetes of the genus Micromonospora and a genus-specific nested PCR for rapid identification of putative Micromonospora isolates were developed. The robustness of both the isolation and the identification approach was underpinned by phylogenetic analysis based on 16S rRNA gene sequences.

  20. Molecular Characterization and Analysis of 16S Ribosomal DNA in some Isolates of Demodex folliculorum

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    Afrooz DANESHPARVAR

    2017-06-01

    Full Text Available Background: Demodicosis is one of the most prevalent skin diseases resulting from infestation by Demodex mites. This parasite usually inhabits in follicular infundibulum or sebaceous duct transmitted through close contact with an infested host.Methods: This study was carried from September 2014 to January 2016 at Tehran University of Medical Sciences, Tehran, Iran. DNA extraction and amplification of 16S ribosomal RNA was performed on four isolates, obtained from four patients and identified morphologically through clearing with 10% Potassium hydroxide (KOH and microscopical examination. Amplified fragments from the isolates were compared with GenBank database and phylogenetic analysis was carried out using MEGA6 software.Results: A 390 bp fragment of 16S rDNA was obtained in all isolates and analysis of generated sequences showed high similarity with those submitted to GenBank, previously. Intra-species similarity and distance also showed 99.983% and 0.017, respectively, for the studied isolates. Multiple alignments of the isolates showed Single Nucleotide Polymorphisms (SNPs in 16S rRNA fragment. Phylogenetic analysis revealed that all 4 isolates clustered with other D. folliculorum, recovered from GenBank database. Our accession numbers KF875587 and KF875589 showed more similarity together in comparison with two other studied isolates. Conclusion: Mitochondrial 16S rDNA is one of the most suitable molecular barcodes for identification D. folliculorum and this fragment can use for intra-species characterization of the most human-infected mites.

  1. Biochemical and Molecular Analysis of Staphylococcus aureus Clinical Isolates from Hospitalized Patients

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    Amit Karmakar

    2016-01-01

    Full Text Available Staphylococcus aureus is opportunistic human as well as animal pathogen that causes a variety of diseases. A total of 100 Staphylococcus aureus isolates were obtained from clinical samples derived from hospitalized patients. The presumptive Staphylococcus aureus clinical isolates were identified phenotypically by different biochemical tests. Molecular identification was done by PCR using species specific 16S rRNA primer pairs and finally 100 isolates were found to be positive as Staphylococcus aureus. Screened isolates were further analyzed by several microbiological diagnostics tests including gelatin hydrolysis, protease, and lipase tests. It was found that 78%, 81%, and 51% isolates were positive for gelatin hydrolysis, protease, and lipase activities, respectively. Antibiogram analysis of isolated Staphylococcus aureus strains with respect to different antimicrobial agents revealed resistance pattern ranging from 57 to 96%. Our study also shows 70% strains to be MRSA, 54.3% as VRSA, and 54.3% as both MRSA and VRSA. All the identified isolates were subjected to detection of mecA, nuc, and hlb genes and 70%, 84%, and 40% were found to harbour mecA, nuc, and hlb genes, respectively. The current investigation is highly important and informative for the high level multidrug resistant Staphylococcus aureus infections inclusive also of methicillin and vancomycin.

  2. Preliminary physiological characteristics of thermotolerant Saccharomyces cerevisiae clinical isolates identified by molecular biology techniques.

    Science.gov (United States)

    Siedlarz, P; Sroka, M; Dyląg, M; Nawrot, U; Gonchar, M; Kus-Liśkiewicz, M

    2016-03-01

    The aim of the study was a molecular identification and physiological characteristic of the five Saccharomyces cerevisiae strains isolated from patients. The tested isolates were compared with control strains (which are of laboratory or commercial origin). The relation of the isolates to baker's yeast S. cerevisiae was studied using species-specific primers in PCR analysis of the ITS-26S region of DNA. Five isolates were genetically identified as the yeast belonging to the genus S. cerevisiae. The effects of temperature and carbon sources on the growth of the yeast strains were analysed. A quantitative characterization of growth kinetics approve that some tested isolates are thermotolerant and are able to grow at range 37-39°C. Among them, one representative is characterized by the highest specific growth rate (0·637 h(-1) ). In conclusions, some strains are defined as potential candidates to use in the biotechnology due to a higher growth rate at elevated temperatures. Screening for further evaluation of biotechnological significance of the tested isolates will be done (e.g. ethanol and trehalose production at higher temperatures). The physiological characterization and confirmation of species identification by molecular methods for yeasts important in the context of biotechnology industry were demonstrated. Thermotolerant microbial strains are required in various industrial applications, for improving productivity and for decreasing the risk of undesirable contaminations when higher temperatures are used. It is important to search for such strains in extreme environments or exotic niches. In this paper, new thermotolerant strains were identified belonging to the Saccharomyces cerevisiae, but differed from typical bakers' yeast, essentially by their growth rate at higher temperature. The described yeast strains are promising for using in biotechnological industry, especially, for production of ethanol and other products at higher temperatures. © 2015 The

  3. Selection, isolation, and identification of fungi for bioherbicide production

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    Angélica Rossana Castro de Souza

    Full Text Available Abstract Production of a bioherbicide for biological control of weeds requires a series of steps, from selection of a suitable microbial strain to final formulation. Thus, this study aimed to select fungi for production of secondary metabolites with herbicidal activity using biological resources of the Brazilian Pampa biome. Phytopathogenic fungi were isolated from infected tissues of weeds in the Pampa biome. A liquid synthetic culture medium was used for production of metabolites. The phytotoxicity of fungal metabolites was assessed via biological tests using the plant Cucumis sativus L., and the most promising strain was identified by molecular analysis. Thirty-nine fungi were isolated, and 28 presented some phytotoxic symptoms against the target plant. Fungus VP51 belonging to the genus Diaporthe showed the most pronounced herbicidal activity. The Brazilian Pampa biome is a potential resource for the development of new and sustainable chemical compounds for modern agriculture.

  4. Isolation and identification of sulfate reducing bacteria (SRB) from the sediment pond after a coal mine in Samarinda, East Kalimantan

    Science.gov (United States)

    Kusumawati, Eko; Sudrajat, Putri, Junita Susilaning

    2017-02-01

    Title isolation and identification of sulfate reducing bacteria (SRB) of sediment pond former coal mine in Samarinda, East Kalimantan. Sulfate reducing bacteria (SRB) is a group of microbes that can be used to improve the quality of sediment former coal mine. In the metabolic activities, the SRB can reduce sulfate to H2S which immediately binds to metals that are widely available on mined lands and precipitated in the form of metal sulfides reductive. Isolation and identification of sulfate reducing bacteria carried out in the Laboratory of Microbiology and Molecular Genetics, Faculty of Mathematics and Natural Sciences, University of Mulawarman, Samarinda. Postgate B is a liquid medium used for isolation through serial dilution. Physiological and biochemical characterization was done by Bergey's Manual of Determinative Bacteriology. Six isolates of sulfate reducing bacteria were isolated from the sediment pond former coal mine in Samarinda. Several groups of bacteria can grow at 14 days of incubation, however, another group of bacteria which takes 21 days to grow. The identification results showed that two isolates belong to the genus Desulfotomaculum sp., and each of the other isolates belong to the genus Desulfococcus sp., Desulfobacter sp., Desulfobulbus sp. and Desulfobacterium sp.

  5. Molecular and Genetic Aspects of Congenital Isolated Hypogonadotropic Hypogonadism.

    Science.gov (United States)

    Lima Amato, Lorena Guimaraes; Latronico, Ana Claudia; Gontijo Silveira, Leticia Ferreira

    2017-06-01

    Congenital isolated hypogonadotropic hypogonadism (IHH) is a clinically and genetically heterogenous disorder characterized by abnormal synthesis, secretion, or action of gonadotropin-releasing hormone, a key hypothalamic decapeptide that orchestrates the reproductive axis. Several modes of inheritance have been identified. A growing list of causative genes has been implicated in the molecular pathogenesis of syndromic and nonsyndromic IHH, largely contributing for better understanding the complex neuroendocrine control of reproduction. This article summarizes the great advances of molecular genetics of IHH and pointed up the heterogeneity and complexity of the genetic basis of this condition. Copyright © 2017 Elsevier Inc. All rights reserved.

  6. Isolation and Metagenomic Identification of Avian Leukosis Virus Associated with Mortality in Broiler Chicken.

    Science.gov (United States)

    Bande, Faruku; Arshad, Siti Suri; Omar, Abdul Rahman

    2016-01-01

    Avian leukosis virus (ALV) belongs to the family Retroviridae and causes considerable economic losses to the poultry industry. Following an outbreak associated with high mortality in a broiler flock in northern part of Malaysia, kidney tissues from affected chickens were submitted for virus isolation and identification in chicken embryonated egg and MDCK cells. Evidence of virus growth was indicated by haemorrhage and embryo mortality in egg culture. While viral growth in cell culture was evidenced by the development of cytopathic effects. The isolated virus was purified by sucrose gradient and identified using negative staining transmission electron microscopy. Further confirmation was achieved through next-generation sequencing and nucleotide sequence homology search. Analysis of the viral sequences using the NCBI BLAST tool revealed 99-100% sequence homology with exogenous ALV viral envelope protein. Phylogenetic analysis based on partial envelope sequences showed the Malaysian isolate clustered with Taiwanese and Japanese ALV strains, which were closer to ALV subgroup J, ALV subgroup E, and recombinant A/E isolates. Based on these findings, ALV was concluded to be associated with the present outbreak. It was recommended that further studies should be conducted on the molecular epidemiology and pathogenicity of the identified virus isolate.

  7. Isolation and Metagenomic Identification of Avian Leukosis Virus Associated with Mortality in Broiler Chicken

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    Faruku Bande

    2016-01-01

    Full Text Available Avian leukosis virus (ALV belongs to the family Retroviridae and causes considerable economic losses to the poultry industry. Following an outbreak associated with high mortality in a broiler flock in northern part of Malaysia, kidney tissues from affected chickens were submitted for virus isolation and identification in chicken embryonated egg and MDCK cells. Evidence of virus growth was indicated by haemorrhage and embryo mortality in egg culture. While viral growth in cell culture was evidenced by the development of cytopathic effects. The isolated virus was purified by sucrose gradient and identified using negative staining transmission electron microscopy. Further confirmation was achieved through next-generation sequencing and nucleotide sequence homology search. Analysis of the viral sequences using the NCBI BLAST tool revealed 99-100% sequence homology with exogenous ALV viral envelope protein. Phylogenetic analysis based on partial envelope sequences showed the Malaysian isolate clustered with Taiwanese and Japanese ALV strains, which were closer to ALV subgroup J, ALV subgroup E, and recombinant A/E isolates. Based on these findings, ALV was concluded to be associated with the present outbreak. It was recommended that further studies should be conducted on the molecular epidemiology and pathogenicity of the identified virus isolate.

  8. Phenotypic and molecular characterization of Brachyspira spp. isolated from laying hens in different housing systems.

    Science.gov (United States)

    Jansson, D S; Fellström, C; Råsbäck, T; Vågsholm, I; Gunnarsson, A; Ingermaa, F; Johansson, K-E

    2008-08-25

    Several species of intestinal spirochaetes, Brachyspira (B.) alvinipulli, B. intermedia and B. pilosicoli, may cause reduced egg production and faecal staining of eggshells in chickens. The aim of this study was to characterize potentially pathogenic and presumably non-pathogenic Brachyspira spp. from commercial laying hens. Selective culture, phenotyping, PCR and 16S rRNA gene sequencing were used and clinical data were collected. Phenotypic profiles were obtained for 489 isolates and 351 isolates obtained after subculture, and 30 isolates were selected for molecular characterization. Seven isolates were positive by a B. intermedia-specific PCR based on the nox gene, and two were positive in a B. hyodysenteriae-specific 23S rRNA gene based PCR. By comparative phylogenetic analysis in combination with PCR and phenotyping, seven isolates were identified as B. intermedia, eight isolates as B. innocens, five as B. murdochii, and three isolates each as B. alvinipulli and "B. pulli". The remaining four isolates could not be assigned to any presently recognized species. Co-infection with several species or genetic variants of Brachyspira spp. were detected in some flocks and samples, suggesting a high level of diversity. Organic flocks with access to outdoor areas were at higher risk (RR=2.3; 95% CI 1.5-3.6) for being colonized than chickens in other housing systems. No significant differences between colonized and non-colonized flocks were found regarding clinical parameters, i.e. mortality, egg production, faecally contaminated eggshells, and wet litter. Our results show that a combination of traditional laboratory diagnostics, molecular tests and phylogeny is needed for identification of Brachyspira sp. from chickens.

  9. Phenotypic and genotypic identification of lactic acid bacteria isolated from traditional pickles of the Çubuk region in Turkey.

    Science.gov (United States)

    Bağder Elmacı, Simel; Tokatlı, Mehmet; Dursun, Derya; Özçelik, Filiz; Şanlıbaba, Pınar

    2015-05-01

    A total of 152 lactic acid bacteria (LAB) were isolated from pickles produced in the Ankara-Çubuk region. These isolates were clustered into eight groups on the basis of their phenotypic characteristics including cell morphology, CO2 production from glucose, growth at 10 and 45 °C, growth in 6.5 % NaCl, and growth at pH 9.6. API 50 CH carbohydrate fermentation test, 16S ribosomal RNA (rRNA) sequence analysis, and sodium dodecyl sulfate-acrylamide gel electrophoresis (SDS-PAGE) whole-cell protein profile analysis were also performed for precise identification of the isolates at the species level. Molecular identification revealed that the most prevalent LAB species involved in pickle fermentation were Pediococcus ethanolidurans (46 isolates, 30.3 %), Lactobacillus brevis (37 isolates, 24.3 %), Lactobacillus plantarum (37 isolates, 24.3 %), and Lactobacillus buchneri (15 isolates, 9.9 %). Other LAB were found in minor frequencies such as Pediococcus parvulus (8 isolates, 5.3 %), Lactobacillus namurensis (6 isolates, 3.9 %), Lactobacillus diolivorans (1 isolate, 0.7 %), Lactobacillus parabrevis (1 isolate, 0.7 %), and Enterococcus casseliflavus (1 isolate, 0.7 %). When results of phenotypic and genotypic identification methods were compared, differences in the species distribution of LAB associated with pickles were defined between the API and the 16S rRNA sequencing. The API 50 CHL test coincided with the 16S rRNA results in 71 out of the 152 tested isolates, indicating that API gave unreliable identification results. A clear correlation could not be found between the results of whole-cell SDS profiles and 16S rRNA sequencing. Therefore, molecular characterization by 16S rRNA sequencing was considered to be the most reliable method for identifying isolates. The results presented in this work provide insight in to the LAB population associated with traditional Çubuk pickles and constitute a LAB strain resource for further studies involving the development of

  10. Study of the Mechanical Properties and Vibration Isolation Performance of a Molecular Spring Isolator

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    Muchun Yu

    2016-01-01

    Full Text Available Molecular Spring Isolator (MSI is a novel passive vibration isolation technique, providing High-Static-Low-Dynamic (HSLD stiffness based on the use of molecular spring material. The molecular spring material is a solid-liquid mixture consisting of water and hydrophobic nanoporous materials. Under a certain level of external pressure, water molecules can intrude into the hydrophobic pores of nanoporous materials, developing an additional solid-liquid interface. Such interfaces are able to store, release, and transform mechanical energy, providing properties like mechanical spring. Having been only recently developed, the basic mechanic properties of a MSI have not been studied in depth. This paper focuses on the stiffness influence factors, the dynamic frequency response, and the vibration isolation performance of a MSI; these properties help engineers to design MSIs for different engineering applications. First, the working mechanism of a MSI is introduced from a three-dimensional general view of the water infiltration massive hydrophobic nanoporous pores. Next, a wide range of influence factors on the stiffness properties of MSI are studied. In addition, the frequency response functions (FRFs of the MSI vibration isolation system are studied utilizing the matching method based on equivalent piecewise linear (EPL system. Finally, the vibration isolation properties of MSI are evaluated by force transmissibility.

  11. Identification and Structural Characterization of Naturally-Occurring Broad-Spectrum Cyclic Antibiotics Isolated from Paenibacillus

    Science.gov (United States)

    Knolhoff, Ann M.; Zheng, Jie; McFarland, Melinda A.; Luo, Yan; Callahan, John H.; Brown, Eric W.; Croley, Timothy R.

    2015-08-01

    The rise of antimicrobial resistance necessitates the discovery and/or production of novel antibiotics. Isolated strains of Paenibacillus alvei were previously shown to exhibit antimicrobial activity against a number of pathogens, such as E. coli, Salmonella, and methicillin-resistant Staphylococcus aureus (MRSA). The responsible antimicrobial compounds were isolated from these Paenibacillus strains and a combination of low and high resolution mass spectrometry with multiple-stage tandem mass spectrometry was used for identification. A group of closely related cyclic lipopeptides was identified, differing primarily by fatty acid chain length and one of two possible amino acid substitutions. Variation in the fatty acid length resulted in mass differences of 14 Da and yielded groups of related MSn spectra. Despite the inherent complexity of MS/MS spectra of cyclic compounds, straightforward analysis of these spectra was accomplished by determining differences in complementary product ion series between compounds that differ in molecular weight by 14 Da. The primary peptide sequence assignment was confirmed through genome mining; the combination of these analytical tools represents a workflow that can be used for the identification of complex antibiotics. The compounds also share amino acid sequence similarity to a previously identified broad-spectrum antibiotic isolated from Paenibacillus. The presence of such a wide distribution of related compounds produced by the same organism represents a novel class of broad-spectrum antibiotic compounds.

  12. Molecular Assay for Fraud Identification of Handmade Hamburgers

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    Gilda Eslami

    2014-11-01

    Full Text Available Background: Meat products could be sources of enter pathogens. Identification of meat species in different foods could help us in molecular epidemiological studies of pathogens transmitted by meat. Objectives: In this study, we targeted cytochrome b for identification of beef in handmade hamburgers. Patients and Methods: A total of 110 raw handmade hamburgers were collected from different areas of Yazd city, Iran, during spring of 2013. Genomic DNA was extracted using the salting out method. The beef cytochrome b gene was amplified using specific primers. Analysis of the amplicons was done with agarose gel electrophoresis usinga100 base pair (bp DNA ladder. Results: The results showed that among the 110 handmade hamburger samples, 10 (9.09% samples did not containany cow meat while 100 samples contained cow meat. Conclusions: We used an appropriate molecular method for controlling raw and processed products. Therefore, this study would be useful for control of correct labeling and protection of consumer’s rights.

  13. Molecular typing of Sporothrix schenckii isolates from cats in Malaysia.

    Science.gov (United States)

    Kano, Rui; Okubo, Miki; Siew, Han Hock; Kamata, Hiroshi; Hasegawa, Atsuhiko

    2015-04-01

    Epidemiological data on the aetiologic agents of feline sporotrichosis in Malaysia have not been reported, though human sporotrichosis in Malaysia is reported to be transmitted primarily via cat scratch. To the best of our knowledge, the present report is the first study of the molecular epidemiology of Sporothrix schenckii isolates from cats with sporotrichosis in Malaysia. In the present work, we characterised 18 clinical isolates from cats in Malaysia based on molecular properties, including sequence analyses of the calmodulin gene and the rDNA ITS region and selective PCR of mating type (MAT) loci. In this study, isolates from feline sporotrichosis were identified as a S. schenckii sensu stricto by sequence analyses of the calmodulin gene and the internal transcribed spacer (ITS) region. Notably, phylogenetic analysis of the ITS confirmed assignment to clinical clade D (and not C) of S. schenckii sensu stricto. Therefore, clinical clade D of S. schenckii sensu stricto appeared to be the prevailing source of feline sporotrichosis in Malaysia. The ratio of MAT1-1-1:MAT1-2-1 in these Malaysian isolates was found to be 1 : 0. This result suggested that a clonal strain of S. schenckii is the prevailing causative agent of feline sporotrichosis in Malaysia. © 2015 Blackwell Verlag GmbH.

  14. Molecular Identification of SHV,TEM, CTX-M β lactamases Genes and Antibiotics Resistance Pattern of k.pneumoniae Isolates Collected from ICU Patients of Namazi Hospital, Shiraz, Iran

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    T Archin

    2014-01-01

    Full Text Available Abstract Background and aim: β-lactamase enzymes producing bacteria ESBL have spread widely throughout the world. The production of enzymes induces bacterial resistance to a wide range of antibiotics which is leading to the limitation of infection control and correct treatment. The aim of the present study was to investigate patterns of antibiotic susceptibility to antibiotics and the presence of β-lactamase genes SHV, TEM, CTX-M, in Klebsiella pneumoniae isolates from clinical specimens of intensive care. Methods: Susceptibility of isolated bacteria against 10 antibiotics was determined by agar disk diffusion method according to the CLSI guidelines. The strains (DDST were examined for the presence of the spectrum β-lactamase enzymes. Using E-test, the minimum inhibitory concentration (MIC of the antibiotic was determined to cefotaxime. Moreover, SHV, TEM, CTX-M genes were identified by, Multiplex PCR method, and some of them were sequenced. Results: The antibiotic resistance against 10 antibiotics was determined. The highest percentage of isolates was resistant to ampicillin (100% and sensitivity to imipenem was 1.66%. In this study, the majority of strains produced ESBL (60%. TEM gene in 34.38% and all three genes (TEM and SHV and CTX at 33.13% of isolates were observed. Conclusion: The present study showed that the K. pneumoniae producing ESBL in patients in ICU are common. Therefore, the use of procedures and policies for infection control in hospitals and especially ICU is necessary. Key words: Klebsiella pneumoniae, ESBL, Multiplex PCR, antibiotic sensitivity

  15. PREVALENCE AND IDENTIFICATION OF VIBRIO SPP. ISOLATED ON AQUACULTURED GILTHEAD SEA BREAM

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    C. Scarano

    2011-01-01

    Full Text Available The aim of the study was to investigate the prevalence of Vibrio spp isolated from gilthead sea bream (Sparus aurata farmed on sea cages and to identify and characterize the pathogen by molecular techniques. Eighty fish were collected from two hatcheries located on the North-Est Sardinian Mediterranean coast, and microbiological analysis were performed on different body parts such as skin, gills, muscle and intestinal tract. Subsequently 100 pure colonies with typical morphology and phenotypic characteristics were selected and submitted to the molecular identification. The analysis on the prevalence of Vibrio spp showed the effect of the hatchery rearing system (P<0.001, of the date of sampling (P<0.001, and of the body part (P<0.001. All the strains selected were confirmed to be members of the genus Vibrio spp by the molecular method/techinique/identification, whereas the rpoA gene sequence analyses allowed to identify 89 strains belonging to the species Vibrio harveyi, 6 to V. diabolicus, 2 to V. parahaemolyticus and 1 to V. mediterranei.

  16. NEW PRODUCER STRAINS OF BIOBUTANOL. І. ISOLATION AND IDENTIFICATION

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    O. A. Tigunova

    2013-02-01

    Full Text Available Getting new, more productive strains of microorganisms that produce butanol is a topical problem. Studing of morphological and physiological characteristics of the isolated strains, improvement of their cultivation conditions, optimization of biobutanol synthesis gives the possibility to organize a cost-effective butanol production technology. The aim of this work was searching new butanol and butyric acid producer strains, their identification and studying the main steps of the selective strains biosynthesis. The objects of this study were microorganisms that had allocated from soils and sludges samples of Kiev’s lakes. Obtained cultures have been screened. Three strains were obtained as promising and identified as C. acetobutylicum, C. tyrobutylicum, C. butylicum. Selective medium have been developed and modified for the microorganisms. Producer’s features were investigated in order to maximize the accumulation of target metabolites.

  17. Isolation and biochemical and molecular characterization of Listeria monocytogenes in food

    International Nuclear Information System (INIS)

    Helel, Salma

    2008-01-01

    monocytogenes is a Gram-positive bacteria, saprophytic, non-spore. This is an extremely resistant seeds to environmental conditions outside, especially since the cold psychrotrophic. It can contaminate raw vegetables, cooked meals ready for consumption or foods to be stored in the refrigerator, such as cheese or meat. It is the bacteria responsible for listeriosis. It threatens first unborn children, infants, pregnant women, the elderly and people whose immune system is weakened. Strains of Listeria spp isolated from foods (seafood, meat, meat) were first identified at the stage of the genus by classical tests (Gram staining, catalase test, oxidase test and mobility) and stage of the test case by hemolysis, CAMP test and the gallery Api Listeria. Biochemical characterization allowed after a numerical analysis, to assign 100% of isolates to the genus Listeria. Molecular characterization was performed by PCR amplification of genes coding for protein p60 (iap), the listeriolysine O (hly), the Phosphatidylinositol Phospholipase C (PI-PLC plca) Phosphatidylcholine Phospholipase C (plcB). The result showed an amplification of the iap gene of 100% of the hly gene, plca, plcB of 31.81%. This characterization represents an identification of the collection on the genetic level and shows that 31.81% of isolates, is likely to express the genes responsible for virulence factors of L. monocytogenes, to produce listeriolysine O, phospholipase C and Lecithinase. The molecular identification was performed by microarray technique and identified isolates L. September monocytogenes (five original clinical isolates and two food-borne), fourteen L. innocua (of food) and a strain not identified by DNA chip.. (Author)

  18. Rapid molecular identification of human taeniid cestodes by pyrosequencing approach.

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    Tongjit Thanchomnang

    Full Text Available Taenia saginata, T. solium, and T. asiatica are causative agents of taeniasis in humans. The difficulty of morphological identification of human taeniids can lead to misdiagnosis or confusion. To overcome this problem, several molecular methods have been developed, but use of these tends to be time-consuming. Here, a rapid and high-throughput pyrosequencing approach was developed for the identification of three human taeniids originating from various countries. Primers targeting the mitochondrial cytochrome c oxidase subunit 1 (cox1 gene of the three Taenia species were designed. Variations in a 26-nucleotide target region were used for identification. The reproducibility and accuracy of the pyrosequencing technology was confirmed by Sanger sequencing. This technique will be a valuable tool to distinguish between sympatric human taeniids that occur in Thailand, Asia and Pacific countries. This method could potentially be used for the molecular identification of the taeniid species that might be associated with suspicious cysts and lesions, or cyst residues in humans or livestock at the slaughterhouse.

  19. Rapid molecular identification of human taeniid cestodes by pyrosequencing approach.

    Science.gov (United States)

    Thanchomnang, Tongjit; Tantrawatpan, Chairat; Intapan, Pewpan M; Sanpool, Oranuch; Janwan, Penchom; Lulitanond, Viraphong; Tourtip, Somjintana; Yamasaki, Hiroshi; Maleewong, Wanchai

    2014-01-01

    Taenia saginata, T. solium, and T. asiatica are causative agents of taeniasis in humans. The difficulty of morphological identification of human taeniids can lead to misdiagnosis or confusion. To overcome this problem, several molecular methods have been developed, but use of these tends to be time-consuming. Here, a rapid and high-throughput pyrosequencing approach was developed for the identification of three human taeniids originating from various countries. Primers targeting the mitochondrial cytochrome c oxidase subunit 1 (cox1) gene of the three Taenia species were designed. Variations in a 26-nucleotide target region were used for identification. The reproducibility and accuracy of the pyrosequencing technology was confirmed by Sanger sequencing. This technique will be a valuable tool to distinguish between sympatric human taeniids that occur in Thailand, Asia and Pacific countries. This method could potentially be used for the molecular identification of the taeniid species that might be associated with suspicious cysts and lesions, or cyst residues in humans or livestock at the slaughterhouse.

  20. Isolation and identification of bovine Brucella isolates from Pakistan by biochemical tests and PCR.

    Science.gov (United States)

    Ali, Shahzad; Ali, Qurban; Melzer, Falk; Khan, Iahtasham; Akhter, Shamim; Neubauer, Heinrich; Jamal, Syed M

    2014-01-01

    Brucellosis is endemic in bovines in Pakistan. The Brucella species and biovars involved, however, are unknown. The objectives of the present study were to isolate and characterize brucellae from seropositive milk samples, aborted fetuses, and vaginal swabs of cattle and buffaloes which had recently aborted. The seropositive milk samples, aborted fetuses, and vaginal swabs of cattle and buffaloes were collected from the Potohar Plateau, Pakistan. Isolation of brucellae was done on modified Farrell's serum dextrose agar. Isolates were characterized by conventional biotyping methods, while molecular typing was done by genus (B4/B5) and species-specific (Brucella abortus, Brucella melitensis, Brucella ovis, and Brucella suis) polymerase chain reaction (PCR). A total of 30 isolates were recovered from milk (n = 5), aborted fetuses (n = 13), and vaginal swabs (n = 12). Most isolates were from cattle (56.7 %). All of them were identified as B. abortus biovar 1 based on conventional biotyping methods and genus and species-specific PCR. This preliminary study provides the first report on the prevalence of B. abortus biovar 1 in cattle and buffaloes in Pakistan.

  1. Isolation, identification and antibiotic resistance of Campylobacter strains isolated from domestic and free-living pigeons.

    Science.gov (United States)

    Dudzic, A; Urban-Chmiel, R; Stępień-Pyśniak, D; Dec, M; Puchalski, A; Wernicki, A

    2016-04-01

    1. The aim of this study was to evaluate the occurrence of Campylobacter spp. in domestic and free-living pigeons and to evaluate the antibiotic resistance profiles. 2. The material consisted of cloacal swabs obtained from 108 homing pigeons and fresh faeces from 72 wild birds from Lublin and its vicinity. The identification of strains isolated on differential/selective media for Campylobacter spp. was carried out by MALDI-TOF and PCR. The susceptibility to antibiotics was evaluated by minimum inhibitory concentration (MIC) in Mueller-Hinton broth. 3. A total of 35 strains of Campylobacter spp. were isolated; 27 were identified as Campylobacter jejuni and 8 as Campylobacter coli. Over half of the isolates were resistant to erythromycin and streptomycin, 40% of strains were resistant to tetracycline and ampicillin and 37% isolates were resistant to amoxicillin. Resistance to two or more antibiotics was observed in all strains tested. 4. The results indicate that both domestic and free-living pigeons are reservoirs for bacteria of the genus Campylobacter, which are characterised by varied and growing resistance to commonly used antibiotics.

  2. Identification of lead-resistant endophytic bacteria isolated from rice

    International Nuclear Information System (INIS)

    Perez-Cordero, Alexander; Barraza-Roman, Zafiro; Martinez-Pacheco, Dalila

    2015-01-01

    Resistance of endophytic bacteria in vitro was evaluated at different lead concentrations. The tissue samples of commercial rice varieties at tillering stage were collected during the first half of 2013, in Monteria, Cordoba, Colombia. Each tissue was subjected to surface cleaning. Endophytic bacteria were isolated in agar R 2 A medium. The population density (CFU/g tissue) was determined from each tissue by direct counting of R 2 A medium surface. Morphotypes were classified by shape, color, size and appearance. A total of 168 morphotypes were isolated from root, tillers and leaf of different commercial varieties of rice. The lead resistance test is performed in vitro, The lead resistance test was performed in vitro, by the suspensions of endophytic bacteria in log phase and inoculation in minimal medium with five concentrations of lead as Pb (NO 3 ) 2 . The experiment was incubated at 32 degrees celsius and agitated at 150 rpm for five days. The measure of turbidimetry at 600 nm was conduced every hour afterstarting the test. Endophytic bacteria showed the ability to grow at concentrations of 100% of Pb as Pb (NO 3 ) 2 . The presence of Burkholderia cepacia and Pseudomonas putida, which showed resistance to differents lead concentration was confirmed as result of the identification with kit API20E. (author) [es

  3. Identification of lead- resistant endophytic bacteria isolated from rice.

    Directory of Open Access Journals (Sweden)

    Alexander Pérez-Cordero

    2015-06-01

    Full Text Available   The objective of this study was to evaluate in vitro the endophytic bacteria resistance to different lead concentrations. The sampling was undertaken in the first half of 2013, when tissue samples of commercial varieties of rice at tillering stage were collected in Montería, Cordoba, Colombia. Each tissue was subjected to surface cleaning. Endophytic bacteria in agar R2A medium were isolated. Population density (CFU/g tissue was determined from each tissue, by direct counting of R2A medium surface. morphotypes were classified by shape, color, size, and appearance. A total of 168 morphotypes were isolated from root, tillers, and leaf of different commercial varieties of rice. The lead resistance test was performed in vitro, to do that, suspensions of endophytic bacteria in log phase were prepared and inoculated in minimal medium with five concentrations of lead as Pb(NO32. The experiment was incubated at 32 °C and agitated at 150 rpm, for five days. Every hour afterstarting the test, turbidimetry measuring at 600 nm was conducted. Results showed the ability of endophytic bacteria to grow at concentrations of 100% of Pb as Pb(NO32. The results of the identification with kit API20E confirmed the presence of Burkholderia cepacia and Pseudomonas putida, which showed resistance to different lead concentrations.

  4. Physiological and Molecular Characterization of Atypical Isolates of Malassezia furfur▿

    Science.gov (United States)

    González, A.; Sierra, R.; Cárdenas, M. E.; Grajales, A.; Restrepo, S.; Cepero de García, M. C.; Celis, A.

    2009-01-01

    The species constituting the genus Malassezia are considered to be emergent opportunistic yeasts of great importance. Characterized as lipophilic yeasts, they are found in normal human skin flora and sometimes are associated with different dermatological pathologies. We have isolated seven Malassezia species strains that have a different Tween assimilation pattern from the one typically used to differentiate M. furfur, M. sympodialis, and M. slooffiae from other Malassezia species. In order to characterize these isolates of Malassezia spp., we studied their physiological features and conducted morphological and molecular characterization by PCR-restriction fragment length polymorphism and sequencing of the 26S and 5.8S ribosomal DNA-internal transcribed spacer 2 regions in three strains from healthy individuals, four clinical strains, and eight reference strains. The sequence analysis of the ribosomal region was based on the Blastn algorithm and revealed that the sequences of our isolates were homologous to M. furfur sequences. To support these findings, we carried out phylogenetic analyses to establish the relationship of the isolates to M. furfur and other reported species. All of our results confirm that all seven strains are M. furfur; the atypical assimilation of Tween 80 was found to be a new physiological pattern characteristic of some strains isolated in Colombia. PMID:18971363

  5. Physiological and molecular characterization of atypical isolates of Malassezia furfur.

    Science.gov (United States)

    González, A; Sierra, R; Cárdenas, M E; Grajales, A; Restrepo, S; Cepero de García, M C; Celis, A

    2009-01-01

    The species constituting the genus Malassezia are considered to be emergent opportunistic yeasts of great importance. Characterized as lipophilic yeasts, they are found in normal human skin flora and sometimes are associated with different dermatological pathologies. We have isolated seven Malassezia species strains that have a different Tween assimilation pattern from the one typically used to differentiate M. furfur, M. sympodialis, and M. slooffiae from other Malassezia species. In order to characterize these isolates of Malassezia spp., we studied their physiological features and conducted morphological and molecular characterization by PCR-restriction fragment length polymorphism and sequencing of the 26S and 5.8S ribosomal DNA-internal transcribed spacer 2 regions in three strains from healthy individuals, four clinical strains, and eight reference strains. The sequence analysis of the ribosomal region was based on the Blastn algorithm and revealed that the sequences of our isolates were homologous to M. furfur sequences. To support these findings, we carried out phylogenetic analyses to establish the relationship of the isolates to M. furfur and other reported species. All of our results confirm that all seven strains are M. furfur; the atypical assimilation of Tween 80 was found to be a new physiological pattern characteristic of some strains isolated in Colombia.

  6. Molecular characterisation of Mycobacterium caprae strains isolated in Poland.

    Science.gov (United States)

    Krajewska-Wędzina, Monika; Kozińska, Monika; Orłowska, Blanka; Weiner, Marcin; Szulowski, Krzysztof; Augustynowicz-Kopeć, Ewa; Anusz, Krzysztof; Smith, Noel H

    2018-03-10

    Bovine tuberculosis (bovine TB, bTB) is caused by bovine bacilli: Mycobacterium bovis and M caprae The studies conducted in Poland, in the National Bovine Tuberculosis Reference Laboratory in the Department of Microbiology of the National Veterinary Research Institute in Pulawy, show that animal tuberculosis in Poland is also caused by M caprae We here describe the identification and genotypic assessment of 52 isolates of M caprae obtained from Polish cattle and wild animals over the last five years. We show that strains isolated from bison have significant genotypic diversity and are distinct compared with the genotypes of strains isolated from cattle. Similarly, isolates from cattle herds can be highly genotypically variable. Formal designation of the members of the Mycobacterium tuberculosis complex is controversial in Poland; there is a gap in veterinary legislation with regard to bTB and no explicit mention of M caprae causing tuberculosis in animal. © British Veterinary Association (unless otherwise stated in the text of the article) 2018. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

  7. [The random amplified polymorphic DNA identification of 9 Taenia saginata isolates from four provinces].

    Science.gov (United States)

    Zhang, Ke; Yang, Ming; Bao, Huai-En

    2006-12-01

    To make molecular identification for 9 isolates of Taenia saginata from 4 provinces. Genomic DNA was extracted from the segments of adult tapeworms collected from Taoyuan of Taiwan (TW1), Duyun of Guizhou (DY1, DY2), Congjiang of Guizhou (CJ1, CJ2, CJ3, CJ4), Dali of Yunnan (DL1) and Wushi of Xinjiang (XJ1) respectively. PCRs were carried out with 13 random primers. A phylogenetic tree of different geographical strains was constructed. 331 DNA fragments were amplified. The number of DNA fragments amplified by single primer was between 3 and 28. The average number of amplified DNA fragments by the 13 primers was 14.15. The average number of fragments from the 9 isolates of T. saginata was 14.08. Phylogenetic tree revealed that there were two branches in the tree, DY1, DY2, DL1 and TW1 occupied one branch, while CJ1, CJ2, CJ3, CJ4 and XJ1 occupied the other one. By the RAPD analysis, the isolates DY1, DY2, DL1 and TW1 belong to Taenia saginata asiatica, and the isolates CJ1, CJ2, CJ3, CJ4 and XJ1 belong to T. saginata saginata.

  8. THE CHARACTERIZATION OF SIMPLISIA, ISOLATION AND IDENTIFICATION OF CHEMICAL CONSTITUENS FROM THALLUS Turbinaria decurrens Bory

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    Irma Sari

    2015-09-01

    Full Text Available The characterization of simplisia, phytochemical screening, extraction, isolation and identification of chemical constituens  from thallus Turbinaria decurrens Bory have been carried out. The examination of simplisia characteristics gave the water soluble extract the value of 10.59%, ethanol soluble extract valued at 0.93%, total ash valued at 15.64%, the acid insoluble ash value 0.79% and the water content valued at 8.66%. The result of phytochemical screening showed that there was triterpens/steroids present. The extraction process was carried out by percolation and then was separated by liquid vacum column chromatography. Then by preparative thin layer chromatography isolate B1 was obtained. The analysis of isolate B1 by infra red spectrophotometry showed hydroxyl, aliphatic C-H bond, C=O bond, double bond of C=C, C=O bond, C=H bond of CH3 and CH2, were present. Ultra violet spectrophotometry exhibited a maximum absorption at 242 nm and mass spectrometric fragmentation pattern exhibited that the molecular weight of isolate B1 was 394, it was suspected ergosta -4,7,22 –trien -3 one.

  9. An insight into the isolation, enumeration, and molecular detection of Listeria monocytogenes in food

    Science.gov (United States)

    Law, Jodi Woan-Fei; Ab Mutalib, Nurul-Syakima; Chan, Kok-Gan; Lee, Learn-Han

    2015-01-01

    Listeria monocytogenes, a foodborne pathogen that can cause listeriosis through the consumption of food contaminated with this pathogen. The ability of L. monocytogenes to survive in extreme conditions and cause food contaminations have become a major concern. Hence, routine microbiological food testing is necessary to prevent food contamination and outbreaks of foodborne illness. This review provides insight into the methods for cultural detection, enumeration, and molecular identification of L. monocytogenes in various food samples. There are a number of enrichment and plating media that can be used for the isolation of L. monocytogenes from food samples. Enrichment media such as buffered Listeria enrichment broth, Fraser broth, and University of Vermont Medium (UVM) Listeria enrichment broth are recommended by regulatory agencies such as Food and Drug Administration-bacteriological and analytical method (FDA-BAM), US Department of Agriculture-Food and Safety (USDA-FSIS), and International Organization for Standardization (ISO). Many plating media are available for the isolation of L. monocytogenes, for instance, polymyxin acriflavin lithium-chloride ceftazidime aesculin mannitol, Oxford, and other chromogenic media. Besides, reference methods like FDA-BAM, ISO 11290 method, and USDA-FSIS method are usually applied for the cultural detection or enumeration of L. monocytogenes. most probable number technique is applied for the enumeration of L. monocytogenes in the case of low level contamination. Molecular methods including polymerase chain reaction, multiplex polymerase chain reaction, real-time/quantitative polymerase chain reaction, nucleic acid sequence-based amplification, loop-mediated isothermal amplification, DNA microarray, and next generation sequencing technology for the detection and identification of L. monocytogenes are discussed in this review. Overall, molecular methods are rapid, sensitive, specific, time- and labor-saving. In future, there are

  10. An insight into the isolation, enumeration, and molecular detection of Listeria monocytogenes in food.

    Science.gov (United States)

    Law, Jodi Woan-Fei; Ab Mutalib, Nurul-Syakima; Chan, Kok-Gan; Lee, Learn-Han

    2015-01-01

    Listeria monocytogenes, a foodborne pathogen that can cause listeriosis through the consumption of food contaminated with this pathogen. The ability of L. monocytogenes to survive in extreme conditions and cause food contaminations have become a major concern. Hence, routine microbiological food testing is necessary to prevent food contamination and outbreaks of foodborne illness. This review provides insight into the methods for cultural detection, enumeration, and molecular identification of L. monocytogenes in various food samples. There are a number of enrichment and plating media that can be used for the isolation of L. monocytogenes from food samples. Enrichment media such as buffered Listeria enrichment broth, Fraser broth, and University of Vermont Medium (UVM) Listeria enrichment broth are recommended by regulatory agencies such as Food and Drug Administration-bacteriological and analytical method (FDA-BAM), US Department of Agriculture-Food and Safety (USDA-FSIS), and International Organization for Standardization (ISO). Many plating media are available for the isolation of L. monocytogenes, for instance, polymyxin acriflavin lithium-chloride ceftazidime aesculin mannitol, Oxford, and other chromogenic media. Besides, reference methods like FDA-BAM, ISO 11290 method, and USDA-FSIS method are usually applied for the cultural detection or enumeration of L. monocytogenes. most probable number technique is applied for the enumeration of L. monocytogenes in the case of low level contamination. Molecular methods including polymerase chain reaction, multiplex polymerase chain reaction, real-time/quantitative polymerase chain reaction, nucleic acid sequence-based amplification, loop-mediated isothermal amplification, DNA microarray, and next generation sequencing technology for the detection and identification of L. monocytogenes are discussed in this review. Overall, molecular methods are rapid, sensitive, specific, time- and labor-saving. In future, there are

  11. An insight into the isolation, enumeration and molecular detection of Listeria monocytogenes in food

    Directory of Open Access Journals (Sweden)

    Jodi Woan-Fei Law

    2015-11-01

    Full Text Available Listeria monocytogenes, a foodborne pathogen that can cause listeriosis through the consumption of food contaminated with this pathogen. The ability of L. monocytogenes to survive in extreme conditions and cause food contaminations have become a major concern. Hence, routine microbiological food testing is necessary to prevent food contamination and outbreaks of foodborne illness. This review provides insight into the methods for cultural detection, enumeration and molecular identification of L. monocytogenes in various food samples. There are a number of enrichment and plating media that can be used for the isolation of L. monocytogenes from food samples. Enrichment media such as buffered Listeria Enrichment Broth (BLEB, Fraser broth and University of Vermont Medium (UVM Listeria enrichment broth are recommended by regulatory agencies such as FDA-BAM, USDA-FSIS and ISO. Many plating media are available for the isolation of L. monocytogenes, for instance, PALCAM, Oxford and other chromogenic media. Besides, reference methods like FDA-BAM, ISO 11290 method and USDA-FSIS method are usually applied for the cultural detection or enumeration of L. monocytogenes. MPN technique is applied for the enumeration of L. monocytogenes in the case of low level contamination. Molecular methods including polymerase chain reaction (PCR, multiplex polymerase chain reaction (mPCR, real-time/quantitative polymerase chain reaction (qPCR, nucleic acid sequence-based amplification (NASBA, loop-mediated isothermal amplification (LAMP, DNA microarray and Next Generation Sequencing (NGS technology for the detection and identification of L. monocytogenes are discussed in this review. Overall, molecular methods are rapid, sensitive, specific, time- and labour-saving. In future, there are chances for the development of new techniques for the detection and identification of foodborne with improved features.

  12. Isolation and molecular characterization of Acanthamoeba and Balamuthia mandrillaris from combination shower units in Costa Rica.

    Science.gov (United States)

    Retana-Moreira, Lissette; Abrahams-Sandí, Elizabeth; Cabello-Vílchez, Alfonso Martín; Reyes-Batlle, María; Valladares, Basilio; Martínez-Carretero, Enrique; Piñero, José E; Lorenzo-Morales, Jacob

    2014-11-01

    Free living amoebae (FLA) are ubiquitous protozoa, which may behave as parasites under certain conditions. Four genera are recognized as causal agents of infections in humans and animals: Naegleria, Sappinia, Acanthamoeba and Balamuthia. This work determines the presence of FLA in combination shower units and employs molecular biology for the characterization of isolates. The morphological analysis and partial sequencing of the 18S rDNA gene revealed the presence of Acanthamoeba genotype T4 in 30% of the units sampled. In addition to Acanthamoeba cysts, trophozoites with morphological characteristics similar to Balamuthia were identified. PCR assay using the mitochondrial 16S rRNA gene as a target confirmed the identification of the amoeba as Balamuthia mandrillaris. Up to date, this is the first report of the isolation of B. mandrillaris in Central America and the fifth report worldwide.

  13. Molecular characterization of Giardia duodenalis isolated from Semai Pahang Orang Asli (Peninsular Malaysia aborigines).

    Science.gov (United States)

    Mahdy, A K Mohammed; Surin, Johari; Mohd-Adnan, A; Wan, K-L; Lim, Y A L

    2009-09-01

    This study was conducted to determine the genotypes of Giardia duodenalis isolated from human faecal samples at Pos Betau, Pahang, Malaysia. Faecal specimens were collected and examined for G. duodenalis cysts using Trichrome staining techniques. Molecular identification was carried out by the amplification of a region of the small subunit of the nuclear ribosomal RNA (SSU rRNA) gene using nested PCR and subsequent sequencing. The sequences from 15 isolates from G. duodenalis were subjected to phylogenetic analysis (including appropriate outgroups) using the neighbor-joining and maximum parsimony methods. The trees identified G. duodenalis assemblages A and B, with a predominance of assemblage B. The predominance of anthroponotic genotypes indicates the possibility of anthroponotic transmission of these protozoa in this Semai Pahang Orang Asli community.

  14. Identification of Turicella otitidis isolated from a patient with otorrhea associated with surgery: differentiation from Corynebacterium afermentans and Corynebacterium auris.

    Science.gov (United States)

    Renaud, F N; Grégory, A; Barreau, C; Aubel, D; Freney, J

    1996-01-01

    Turicella otitidis is a newly described coryneform bacterium isolated from middle ear fluids. We report here on the diagnosis of a strain isolated from otorrhea. The API Coryne system (bioMérieux, Marcy I'Etoile, France) used alone failed to differentiate T. otitidis, Corynebacterium afermentans, and Corynebacterium auris (ANF group). Biochemical tests such as DNase, enzymatic reactions (API ZYM; bioMérieux), and carbon substrate assimilation tests (Biotype 100; bioMérieux) allow presumptive identification. However, only chemotaxonomy and molecular biology can achieve unequivocal differentiation among these three species. PMID:8880538

  15. Molecular typing of Burkholderia cepacia complex isolated from patients attending an Italian Cystic Fibrosis Centre.

    Science.gov (United States)

    Teri, Antonio; Sottotetti, Samantha; Biffi, Arianna; Girelli, Daniela; D'Accico, Monica; Arghittu, Milena; Colombo, Carla; Corti, Fabiola; Pizzamiglio, Giovanna; Cariani, Lisa

    2018-03-02

    Bacteria from the Burkholderia cepacia complex (Bcc) are capable of causing severe infections in patients with cystic fibrosis (CF). Bcc infection is often extremely difficult to treat due to its intrinsic resistance to multiple antibiotics. In addition, it seems to speed up the decline of lung function and is considered a contraindication for lung transplantation in CF. This study investigates the species of the Bcc strains recovered from chronically infected CF subjects by means of: isolation, identification methods and complete recA nucleotide sequences of 151 samples. Molecular typing showed that B. cenocepacia III is the dominant strain found in the group of subjects being treated at the Milan CF Centre (Italy) and that the infection is chronically maintained by the same species. Defining species by means of molecular analysis yields important information for the clinician in order to establish the most appropriate therapy and implement correct measures for prevention of transmission among CF subjects.

  16. Classical and Molecular Identification of Thermotolerant Campylobacters from Poultry Meat

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    Tina Zorman

    2002-01-01

    Full Text Available Poultry meat samples from Slovenian retail market were examined for the presence of thermotolerant campylobacters. The isolates were identified by phenotypic and genotypic methods. ISO 10272 recommendations were followed for phenotypic identification. Different PCR assays, targeting species specific DNA regions in C. jejuni and C. coli, were checked for their applicability in identification. High degree of tested samples was positive (27/33, with significant proportion of C. coli (32 % among identified strains. High percentage of C. jejuni strains (54 % were hippurate negative. Phenotypic identification was therefore found to be inconvenient because of the presence of the strains with atypical phenotype and possible misinterpretation of test results. Multiplex PCR, targeting hippuricase gene in C. jejuni and species specific region in C. coli, was found to be an efficient method that allowed fast, simple and accurate identification of C. jejuni and C. coli. FlaA PCR is a reliable method to identify the group C. jejuni/C. coli, but it does not differentiate between the two species. CdtB PCR is inconvenient because of many false negative and some false positive results.

  17. [Morphological and molecular characterization of isolates of Macrophomina phaseolina associated with sugarcane in Mexico].

    Science.gov (United States)

    Leyva-Mir, Santos G; Velázquez-Martínez, Guadalupe C; Tlapal-Bolaños, Bertha; Tovar-Pedraza, Juan M; Rosas-Saito, Greta H; Alvarado-Gómez, Omar G

    2015-01-01

    Charcoal rot caused by Macrophomina phaseolina is an important disease of sugarcane in Mexico. This study was carried out to characterize isolates of M. phaseolina obtained from sugarcane by the combination of morphological and molecular analyses. The morphological characterization of 10 isolates was performed using scanning electron microscopy and light microscopy. To confirm the morphological identification, rDNA from two representative isolates was extracted, and the internal transcribed spacer (ITS) region was amplified by polymerase chain reaction and sequenced using specific primers MpKF1 and MpKR1. Based on their morphological characteristics, all isolates were identified as M. phaseolina. Moreover, the analysis of two ITS sequences showed 100% similarity with the M. phaseolina sequences deposited in the GenBank. To our knowledge, this is the first study in the world aimed at characterizing isolates of M. phaseolina obtained from sugarcane. Copyright © 2014 Asociación Argentina de Microbiología. Publicado por Elsevier España, S.L.U. All rights reserved.

  18. Molecular characterization of Vibrio cholerae isolates from cholera outbreaks in North India.

    Science.gov (United States)

    Kingston, Joseph J; Zachariah, Kuruvilla; Tuteja, Urmil; Kumar, Sanjay; Batra, Harsh Vardhan

    2009-02-01

    Vibrio cholerae isolates recovered from cholera outbreaks in Bhind district of Madhya Pradesh and Delhi, Northern India were characterized. The O1 serogroup isolates from Bhind outbreak were of Inaba serotype whereas both Ogawa and Inaba serotypes were recovered from Delhi. PCR analysis revealed that only O1 serogroup V. cholerae isolates carried the virulence-associated genes like ctxA, tcpA, ace, and zot. Molecular typing by repetitive sequence based ERIC, VCR1, and VC1 PCR's revealed similar DNA profile for both Inaba and Ogawa serotypes. A discrete VC1-PCR band identified among the El Tor strains had greater similarity (>97%) to the V. cholerae genome sequence and therefore has the potential to be used as a marker for the identification of the V. cholerae strains. Non-O1 strains recovered from Bhind region differed among themselves as well as from that of the O1 isolates. All the O1 serogroup isolates possessed SXT element and were uniformly resistant to the antibiotics nalidixic acid, polymyxin-B, furazolidone, cloxacilin, trimethoprim-sulfamethaxazole, and vibriostatic agent 0129. Inaba strains from both Delhi and Bhind differed from Ogawa strains by their resistance to streptomycin despite sharing similar DNA patterns in all the three rep-PCRs. Though Delhi and Bhind are separate geographical regions in Northern India, Inaba strains from both these places appear to be closely related owing to their similarity in antibiogram and genetic profile.

  19. Molecular epidemiology of clinical isolates of Pseudomonas aeruginosa isolated from horses in Ireland

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    Tazumi A

    2009-07-01

    Full Text Available Abstract Clinical isolates (n = 63 of Pseudomonas aeruginosa obtained from various sites in 63 horses were compared using ERIC2 RAPD PCR to determine their genetic relatedness. Resulting banding patterns (n = 24 genotypes showed a high degree of genetic heterogeneity amongst all isolates examined, indicating a relative non-clonal relationship between isolates from these patients, employing this genotyping technique. This study characterised 63 clinical isolates into 24 distinct genotypes, with the largest cluster (genotype E accounting for 10/63 (15.9% of the isolates. ERIC2 RAPD PCR proved to be a highly discriminatory molecular typing tool of P. aeruginosa in isolates recovered from horses. With the adoption of several controls to aid reproducibility, this technique may be useful as an alternative to PFGE, particularly in epidemiological investigations of outbreaks where speed may be a significant parameter. This is the first report of clonal heterogeneity amongst P. aeruginosa from horses and demonstrated that ERIC RAPD PCR is a rapid method for the examination of this species in horses, which may be useful in outbreak analysis.

  20. Rapid Isolation and Molecular Detection of Streptomycin-Producing Streptomycetes

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    M Motovali-bashi

    2006-07-01

    Full Text Available Introduction: Streptomyces species are mycelial, aerobic gram-positive bacteria that are isolated from soil and produce a diverse range of antibiotics. Streptomyces griseus produces the antibiotic, streptomycin and forms spores even in a liquid culture. The gene cluster for the production of Streptomycin antibiotic contains strR gene that encodes StrR, a pathway-specific regulator. Then, this pathway-specific regulator induces transcription of other streptomycin production genes in the gene cluster. The overall aim of this work was rapid isolation and molecular detection of streptomycin-producing Streptomycetes, especially S. griseus, from Iranian soils in order to manipulate them for increased production of streptomycin. Methods: This research used new initiative half-specific medium for isolation of Streptomycetes from natural environments, called FZmsn. The fifty colonies of Streptomyces strains grown on the surface of FZmsn medium isolated from environmental samples were defined on the basis of their morphological characteristics and light microscope studies. A set of primers was designed to detect strR by OLIGO software. Results: In colony-PCR reactions followed by gel electrophoresis, 6 colonies from Streptomyces strains colonies were detected as S. griseus colonies. Conclusion: These native Streptomyces strains will be used for genetic manipulation of S. griseus in order to increase production levels of streptomycin.

  1. Isolation, identification and molecular docking as cyclooxygenase (COX) inhibitors of the main constituents of Matricaria chamomilla L. extract and its synergistic interaction with diclofenac on nociception and gastric damage in rats.

    Science.gov (United States)

    Ortiz, Mario I; Fernández-Martínez, Eduardo; Soria-Jasso, Luis Enrique; Lucas-Gómez, Isaac; Villagómez-Ibarra, Roberto; González-García, Martha P; Castañeda-Hernández, Gilberto; Salinas-Caballero, Mireya

    2016-03-01

    Chamomile (Matricaria chamomilla L., Asteraceae) is a medicinal plant widely used as remedy for pain and gastric disorders. The association of non-steroidal anti-inflammatory drugs (NSAIDs) with medicinal plant extracts may increase its antinociceptive activity, permit the use of lower doses and limit side effects. The aim was to isolate and identify the main chemical constituents of Matricaria chamomilla ethanolic extract (MCE) as well as to explore their activity as cyclooxygenase (COX) inhibitors in silico; besides, to examine the interaction between MCE and diclofenac on nociception in the formalin test by isobolographic analysis, and to determine the level of gastric injury in rats. Three terpenoids, α-bisabolol, bisabolol oxide A, and guaiazulene, were isolated and identified by (1)H NMR. Docking simulation predicted COX inhibitory activity for those terpenoids. Diclofenac, MCE, or their combinations produced an antinociceptive effect. The sole administration of diclofenac and the highest combined dose diclofenac-MCE produced significant a gastric damage, but that effect was not seen with MCE alone. An isobologram was constructed and the derived theoretical ED35 for the antinociceptive effect was significantly different from the experimental ED35; hence, the interaction between diclofenac and MCE that mediates the antinociceptive effect is synergist. The MCE contains three major terpenoids with plausible COX inhibitory activity in silico, but α-bisabolol showed the highest affinity. Data suggest that the diclofenac-MCE combination can interact at the systemic level in a synergic manner and may have therapeutic advantages for the clinical treatment of inflammatory pain. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

  2. Principal methods for isolation and identification of soil microbial communities.

    Science.gov (United States)

    Stefanis, Christos; Alexopoulos, Athanasios; Voidarou, Chrissa; Vavias, Stavros; Bezirtzoglou, Eugenia

    2013-01-01

    Soil microbial populations play crucial role in soil properties and influence below-ground ecosystem processes. Microbial composition and functioning changes the soil quality through decomposition of organic matter, recycling of nutrients, and biological control of parasites of plants. Moreover, the discovery that soil microbes may translate into benefits for biotechnology, management of agricultural, forest, and natural ecosystems, biodegradation of pollutants, and waste treatment systems maximized the need of scientists for the isolation and their characterization. Operations such as the production of antibiotics and enzymic activities from microorganisms of soil constitute objectives of industry in her effort to cope with the increase of population of earth and disturbance of environment and may ameliorate the effects of global climate change. In the past decades, new biochemical and molecular techniques have been developed in our effort to identify and classify soil bacteria. The goal of measuring the soil microbial diversity is difficult because of the limited knowledge about bacteria species and classification through families and orders. Molecular techniques extend our knowledge about microbial diversity and help the taxonomy of species. Measuring and monitoring soil microbial communities can lead us to better understanding of their composition and function in many ecosystem processes.

  3. Molecular and serological characterization of the first Leptospira santarosai strain isolated from a dog.

    Science.gov (United States)

    Miotto, Bruno Alonso; Moreno, Luisa Zanolli; Guilloux, Aline Gil Alves; Sousa, Gisele Oliveira de; Loureiro, Ana Paula; Moreno, Andrea Micke; Lilenbaum, Walter; Vasconcellos, Silvio Arruda; Heinemann, Marcos Bryan; Hagiwara, Mitika Kuribayashi

    2016-10-01

    Leptospirosis is a zoonotic disease of global importance caused by pathogenic Leptospira species. Dogs can become asymptomatically infected, acting like reservoir hosts for pathogenic Leptospira, notably Leptospira interrogans serovar Canicola. Identification of such individuals and characterization of leptospires involved in chronic infections may unravel the role of dogs in the epidemiology of particular leptospiral strains. The aim of the present work was to describe the first Leptospira santarosai strain isolated from a dog. The dog was kept in a public shelter in São Paulo city, Brazil, and presented asymptomatic urinary shedding detected by PCR. Prospective evaluation was performed to fully characterize its chronic carrier state. The dog did not present anti-Leptospira titles or clinical/laboratorial abnormalities during the evaluations; nevertheless long-term urinary shedding was confirmed by PCR and leptospires were recovered from two occasions. The isolated strain was molecularly characterized by partial 16S rRNA and secY gene sequencing and MLST analysis. Serogroup identification was performed using polyclonal antibodies. The strain was identified as Leptospira santarosai, serogroup Sejroe. This is the first evidence in the literature of the isolation of L. santarosai in dogs. Our findings show that dogs can persistently harbor leptospires other than L. interrogans. Copyright © 2016 Elsevier B.V. All rights reserved.

  4. Molecular detection of an atypical, highly resistant, clonal Pseudomonas aeruginosa isolate in cystic fibrosis patients.

    LENUS (Irish Health Repository)

    Keating, Deirdre

    2013-03-01

    The identification of Pseudomonas aeruginosa (P. aeruginosa) isolates in sputum from cystic fibrosis (CF) patients can be challenging due to the multitude of phenotypic changes isolates undergo during adaptation to the microenvironment of the CF lung.

  5. Soy protein isolate molecular level contributions to bulk adhesive properties

    Science.gov (United States)

    Shera, Jeanne Norton

    Increasing environmental awareness and the recognized health hazards of formaldehyde-based resins has prompted a strong demand for environmentally-responsible adhesives for wood composites. Soy protein-based adhesives have been shown to be commercially viable with 90-day shelf stability and composite physical properties comparable to those of commercial formaldehyde-based particleboards. The main research focus is to isolate and characterize the molecular level features in soy protein isolate responsible for providing mechanical properties, storage stability, and water resistance during adhesive formulation, processing, and wood composite fabrication. Commercial composite board will be reviewed to enhance our understanding of the individual components and processes required for particleboard production. The levels of protein structure will be defined and an overview of current bio-based technology will be presented. In the process, the logic for utilizing soy protein as a sole binder in the adhesive will be reinforced. Variables such as adhesive components, pH, divalent ions, blend aging, protein molecular weight, formulation solids content, and soy protein functionalization will relate the bulk properties of soy protein adhesives to the molecular configuration of the soybean protein. This work has demonstrated that when intermolecular beta-sheet interactions and protein long-range order is disrupted, viscosity and mechanical properties decrease. Storage stability can be maintained through the stabilization of intermolecular beta-sheet interactions. When molecular weight is reduced through enzymatic digestion, long-range order is disrupted and viscosity and mechanical properties decrease accordingly. Processibility and physical properties must be balanced to increase solids while maintaining low viscosity, desirable mechanical properties, and adequate storage stability. The structure of the soybean protein must be related to the particleboard bulk mechanical

  6. Rapid and Accurate Molecular Identification of the Emerging Multidrug-Resistant Pathogen Candida auris.

    Science.gov (United States)

    Kordalewska, Milena; Zhao, Yanan; Lockhart, Shawn R; Chowdhary, Anuradha; Berrio, Indira; Perlin, David S

    2017-08-01

    Candida auris is an emerging multidrug-resistant fungal pathogen causing nosocomial and invasive infections associated with high mortality. C. auris is commonly misidentified as several different yeast species by commercially available phenotypic identification platforms. Thus, there is an urgent need for a reliable diagnostic method. In this paper, we present fast, robust, easy-to-perform and interpret PCR and real-time PCR assays to identify C. auris and related species: Candida duobushaemulonii , Candida haemulonii , and Candida lusitaniae Targeting rDNA region nucleotide sequences, primers specific for C. auris only or C. auris and related species were designed. A panel of 140 clinical fungal isolates was used in both PCR and real-time PCR assays followed by electrophoresis or melting temperature analysis, respectively. The identification results from the assays were 100% concordant with DNA sequencing results. These molecular assays overcome the deficiencies of existing phenotypic tests to identify C. auris and related species. Copyright © 2017 Kordalewska et al.

  7. Morphological and Molecular Identification of Colletotrichum acutatum from Tomato Fruit

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    Svetlana Živković

    2010-01-01

    Full Text Available Colletotrichum gloeosporioides, Colletotrichum acutatum, Colletotrichum coccodes, and Colletotrichum dematium are the four main species of Colletotrichum that cause tomato anthracnose. In Serbia, the occurrence of anthracnose on tomato fruit has been recorded during the last several years. Typical fruit symptoms include dark, sunken, and circular lesion with orange conidial masses. Pathogen isolates were obtained from a diseased tomato fruits, on PDA medium forming a white to gray colonies. The cultures developed black acervuli around the center of the colony. Conidia were hyaline, aseptate, and fusiform or rarely cylindrical. Appressoria were smooth, simple, clavate to ovate, and variedfrom light to dark brown. Pathogenicity tests with representative isolates were conducted on symptomless, detached tomato fruits. All tested isolates caused anthracnose lesions on tomato fruit after 7 days of incubation. Koch’s postulates were fulfilled by reisolationfrom inoculated tomato fruits. PCR analysis (using species-specific primer pair, CaInt2/ITS4 of genomic DNA from tomato isolates resulted in an amplification product of 490 bp, specific for C. acutatum, further confirming the identity of the pathogen. Based onmorphological and molecular characteristics, the isolates from tomato fruit were determined as C. acutatum.

  8. Isolation and Identification of L-asparaginase producing Erwinia strains which isolated from Potato Farms

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    Arastoo Badoei-Dalfard

    2016-09-01

    Full Text Available Introduction: L-Asparaginase can be effectively used for the treatment of lymphoblastic leukemia. The rapid growth of cancer cells are needed for L-asparagine abundant storage. L-asparaginase catalyzes the hydrolysis of L-asparagine into L-aspartic acid and ammonia. The purpose of this study was to isolate and identify the L-asparaginase producing Erwinia strains from the potato farms of Jiroft. Materials and methods: Pectolytic Erwinia species isolated from crumbling potato in M9 medium. The desired L-asparaginase producing bacteria were isolated based on the color changes. Biochemical-microbial and the plant pathogenicity tests of these strains were also investigated with potato and geranium. The L-asparaginase production and molecular detection of these Erwinia strains were also investigated. Results: In this study, L-asparaginase producing Erwinia was isolated on the CVP and M9 mediums. The inoculation of Erwinia strains on the potato and geranium plants showed that Er8 and Er11 species have the ability to cause plant pathogenicity. Results showed that the maximum pathogenicity of Er8 and Er11 was observed after 48 and 15 h of inoculation in potato and geranium plants, respectively. 16S rDNA sequencing and phylogenetic analyses exhibited that Er8 and Er11 strains were similar to Erwinia chrysanthemi with 98% homology. Discussion and conclusion: Because of several applications of the Erwinia L-asparaginase in various fields, isolated Erwinia and their L-asparaginase can be suitable for applied utilization.

  9. Molecular identification of Streptococcus equi subsp. equi and Streptococcus equi subsp. zooepidemicus in nasal swabs samples from horses suffering respiratory infections in Iran.

    Science.gov (United States)

    Jannatabadi, A A; Mohammadi, G R; Rad, M; Maleki, M

    2008-02-01

    The objective of this study was to evaluate the existence of Streptococcus equi subsp. equi and Streptococcus equi subsp. zooepidemicus as probable agents associated with naturally occurring infection of the equine upper respiratory disease in Mashhad area. Nasal swabs samples from thirty horses with upper respiratory tract infections were collected. The bacteria isolated and identified were Streptococcus equi subsp. equi (1 isolate), Streptococcus equi subsp. zooepidemicus (25 isolates), Pasteurella sp. (11 isolates), Staphylococcus sp. (17 isolates), Bacillus sp. (4 isolates), Pseudomonas sp. (4 isolates), Proteus sp. (1 isolate), Neisseria sp. (1 isolate) and E. coli (1 isolate). All 25 isolates of Streptococcus equi subsp. zooepidemicus and the isolate of Streptococcus equi subsp. equi were characterized by biochemical tests and molecular techniques. For molecular identification of the subspecies S. equi and S. zooepidemicus two genomic region SeM and sodA were amplified. This study is the first report of molecular identification of Streptococcus equi subsp. equi and Streptococcus equi subsp. zooepidemicus in Iran.

  10. Molecular identification of house dust mites and storage mites.

    Science.gov (United States)

    Wong, Shew Fung; Chong, Ai Ling; Mak, Joon Wah; Tan, Jessie; Ling, Suk Jiun; Ho, Tze Ming

    2011-10-01

    Mites are known causes of allergic diseases. Currently, identification of mites based on morphology is difficult if only one mite is isolated from a (dust) sample, or when only one gender is found, or when the specimen is not intact especially with the loss of the legs. The purpose of this study was to use polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) of the ITS2 gene, to complement the morphological data for the identification of mites to the species level. For this, six species were cultured: Dermatophagoides pteronyssinus, D. farinae, Blomia tropicalis, Tyrophagus putrescentiae, Aleuroglyphus ovatus and Glycycometus malaysiensis. Genomic DNA of the mites was extracted, quantified, amplified and digested individually with restriction enzymes. Hinf I and Ple I differentiated the restriction patterns of D. pteronyssinus and D. farinae. Bfa I and Alu I enzymes differentiated B. tropicalis and G. malaysiensis. Ple I enzyme was useful for the differentiation between T. putrescentiae and A. ovatus. Bfa I was useful for the differentiation of G. malaysiensis from the rest of the species. In conclusion, different species of mites can be differentiated using PCR-RFLP of ITS2 region. With the established PCR-RFLP method in this study, identification of these mites to the species level is possible even if complete and intact adult specimens of both sexes are not available. As no study to date has reported PCR-RFLP method for the identification of domestic mites, the established method should be validated for the identification of other species of mites that were not included in this study.

  11. Molecular epidemiology of pneumococcal isolates from children in China

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    Li-Hua Kang

    2016-04-01

    Full Text Available Objectives: To investigate the molecular epidemiology of pneumococcal isolates in Chongqing, China. Methods: In this cross-sectional study, 51 invasive Streptococcus pneumoniae (S. pneumoniae strains were from children with invasive pneumococcal disease (IPD and 32 carriage strains from healthy children from January 2010 to December 2013 at the Children’s Hospital of Chongqing Medical University, Chongqing, China. Multilocus sequence typing was used to identify the sequence types (STs. Capsular serotypes were determined by multiplex polymerase chain reaction. Drug susceptibility and resistance was determined by minimum inhibitory concentrations. Results: In this study, 11 serotypes were identified among the 83 S. pneumoniae clinical isolates tested. Prevalent serotypes were 19A (20.4%, 6A/B (20.4%, 19F (15.7%, 14 (14.5%, and 23F (10.8%. Serotype 19F was the most frequent carriage strain, and serotype 19A was the most frequent invasive strain. The ST983 was the most prevalent ST for carriage strains, and ST320 was the most prevalent ST for invasive strains. For gene analysis, psaA (99.5% and piaA (98.6% were present and much conserved in all pneumococci tested. The cps2A and pcsB genes were more frequent in invasive isolates than carriage strains. Antimicrobial resistance rates of invasive pneumococcal isolates to erythromycin, penicillin, meropenem, cefotaxime, and clindamycin were higher than the carriage isolates from children. Conclusion: Our epidemiological evidence shows that 19A, 6A/B, 19F, 14, and 23F remain the most prevalent serotypes, which can be targeted by PCV13. Genotypes and drug resistance varied between carriage and invasive strains. The PsaA and PiaA may be good protein vaccine candidates.

  12. Multilocus microsatellite markers for molecular typing of Candida tropicalis isolates.

    Science.gov (United States)

    Wu, Yuan; Zhou, Hai-jian; Che, Jie; Li, Wen-ge; Bian, Fu-ning; Yu, Shuan-bao; Zhang, Li-juan; Lu, Jinxing

    2014-11-20

    Candida tropicalis is considered to be the leading pathogen causing nosocomial fungemia and hepatosplenic fungal infections in patients with cancer, particularly those with leukemia. Microsatellite-based typing methods using sets of genetic markers have been developed and reported for population structure analysis of C. albicans, C. glabrata, and C. parapsilosis, but no studies have been published for genetic analysis of C. tropicalis. The objective of this study was to develop new microsatellite loci that have the ability to distinguish among C. tropicalis isolates. DNA sequences containing over 10 bi- or tri-nucleotide repeats were selected from the C. tropicalis genome database. Thirty PCR primers sets specific for the microsatellite loci were designed and tested using eight clinically independent isolates. According to the amplification efficiency, specificity, and observed polymorphisms, eight markers were selected for further population structure analysis and molecular typing. Sixty-five independent C. tropicalis isolates were genotyped using these 8 markers. Based on these analyses, six microsatellite loci were confirmed, although two loci were found to be with unstable flanking areas. The six polymorphic loci displayed 4-22 alleles and 7-27 genotypes. The discriminatory power of the six loci ranged from 0.70 to 0.95. Genotyping results obtained by microsatellite analysis were compared to PCR-fingerprinting and multi-locus sequence typing (MLST). The comparisons showed that microsatellite analysis and MLST had the similar discriminatory power for C. tropicalis, which were more powerful than PCR-fingerprinting. This is the first attempt to develop new microsatellite loci for C. tropicalis. These newly developed markers will be a valuable resource for the differentiation of C. tropicalis isolates. More C. tropicalis isolates will need to be sequenced and analyzed in order to fully show the potential of these newly developed microsatellite markers.

  13. Isolation and identification of Bacillus spp. from compost material, compost and mushroom casing soil active against Trichoderma spp.

    Directory of Open Access Journals (Sweden)

    Stanojević Olja

    2016-01-01

    Full Text Available The isolation of bacteria was carried out from samples of straw and chicken manure, compost at various stages of the composting process and casing soil used for growing button mushrooms. A preliminary screening of 108 bacterial isolates for antagonistic activity against Trichoderma aggressivum f. europaeum showed that 23 tested isolates inhibited mycelial growth of the pathogenic fungus. Further screening with four indicator isolates of fungi revealed that all 23 bacterial isolates inhibited the growth of T. aggressivum f. europaeum, T. harzianum and T. koningii, while only 13 isolates inhibited the growth of T. atroviride. T. aggressivum f. europaeum proved to be the most sensitive, with many bacterial isolates generating a high percentage of growth inhibition. Only two bacterial isolates (B-129 and B-268 were successful in inhibiting the growth of all 4 tested pathogens. All 23 bacterial isolates were characterized as Gram-positive and catalase-positive and were subjected to molecular identification based on the partial sequence, the hypervariant region of the 16S rDNA. It was shown that the obtained bacterial strains belong to Bacillus subtilis, B. amyloliquefaciens, B. licheniformis and B. pumilus species. [Projekat Ministarstva nauke Republike Srbije, br. 31043 i br. 173026

  14. Molecular characterization of the capsular antigens of Pasteurella multocida isolates using multiplex PCR

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    Khalid S. Al-Maary

    2017-02-01

    Full Text Available The use of molecular techniques for detection and characterization of the Pasteurella multocida is very important for rapid and specific detection and characterization of the organism. During the period from 15th February, 2014 to 15th April, 2015, 425 nasopharyngeal swabs and 175 lung and spleen samples were collected and examined by conventional methods, 80 strains (18.82% of P. multocida were isolated from the calves, sheep and goat with respiratory manifestation. Meanwhile, 77 strains (44% were isolated from emergency slaughtered animals. All the recovered strains were positive for specific PCR for detection of P. multocida strains previously identified as P. multocida by standard microbiological techniques. Multiplex PCR for molecular typing of the capsular antigens of the recovered P. multocida revealed positive amplification of 1044 bp fragments specific to the capsular antigen type A with 105 strains (66.88%, and amplification 511 bp fragments of the capsular antigen type E with 52 strain (33.12% and absence of B, D and F antigens. Multiplex PCR for molecular typing of the capsular antigens of P. multocida can be used as a simple, sensitive, rapid, reliable technique instead of the serological techniques for identification of the capsular antigens of P. multocida

  15. Identification and Characterization of Imipenem-Resistant Klebsiella pneumoniae and Susceptible Klebsiella variicola Isolates Obtained from the Same Patient.

    Science.gov (United States)

    Garza-Ramos, Ulises; Moreno-Dominguez, Stephania; Hernández-Castro, Rigoberto; Silva-Sanchez, Jesús; Barrios, Humberto; Reyna-Flores, Fernando; Sanchez-Perez, Alejandro; Carrillo-Casas, Erika M; Sanchez-León, María Carmen; Moncada-Barron, David

    2016-04-01

    Klebsiella variicola, a bacterium closely genetically related to Klebsiella pneumoniae, is commonly misidentified as K. pneumoniae by biochemical tests. To distinguish between the two bacteria, phylogenetic analysis of the rpoB gene and the identification of unique genes in both bacterial species by multiplex-polymerase chain reaction (PCR) provide the means to reliably identify and genotype K. variicola. In recent years, K. variicola has been described both as the cause of an intrahospital outbreak in a pediatric hospital, which resulted in sepsis in inpatients, and as a frequent cause of bloodstream infections. In the present study, K. pneumoniae and K. variicola were isolated from a unique patient displaying different antimicrobial susceptibility phenotypes and different genotypes of virulence determinants. Eight clinical isolates were obtained at different time intervals; all during a 5-month period. The isolates were identified as K. pneumoniae by an automated identification system. The clinical (biochemical test) and molecular (multiplex-PCR and rpoB gene) characterization identified imipenem resistance in the first six K. pneumoniae ST258 isolates, which encode the SHV-12 cephalosporinase and KPC-3 carbapenemase genes. The two last remaining isolates corresponded to susceptible K. variicola. The bacterial species showed a specific profile of virulence-associated determinants, specifically the fimA, fimH, and ecpRAB fimbrial-encoding genes identified only in K. pneumoniae isolates. However, the entb (enterobactin), mrkD (fimbrial adhesin), uge (epimerase), ureA (urease), and wabG (transferase) genes were shared between both bacterial species. Recent studies attribute a higher mortality rate to K. variicola than to K. pneumonia. This work highlights the identification of K. pneumoniae and the closely related K. variicola isolated from the same patient. The value of distinguishing between these two bacterial species is in their clinical significance, their

  16. Molecular Characterisation of Salmonella enterica Serovar Typhi Isolated from Typhoidial Humans

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    Arunava Das

    2012-09-01

    Full Text Available Aims: Salmonella enterica serovar Typhi is the major causative agent for typhoidial fever around the globe among human population reported till date. Present research work was carried out for detection and molecular characterisation of Salmonella enterica serovar Typhi isolated from humans with Typhoidial fever by biochemical, phenotypical and virulence gene based polymerase chain reaction (PCR techniques. The isolated strains were also investigated for antibiotic susceptibility patterns as a control measure. Methodology and Results: A total of 16 clinical samples were collected from the same numbers of patients (7 males and 9 females from Coimbatore, Erode and Salem districts of Tamil Nadu and were processed via broth enrichment methods for isolation and identification of the causative agent S. enterica serovar Typhi. Microbiological and biochemical investigations revealed the presence of S. Typhi from 16 samples. The biotyping of the isolates showed that all the isolates belonged to biotype IV. The PCR analysis confirmed the presence of invA (Invasion gene, 244bp, tyv (Tyveloseepimerase gene, 615 bp, fliC-d (Phage-1 flagellin gene for d-antigen, 750 bp and viaB (Vi antigen gene, 439bp in all 16 clinical samples. The antibiotic susceptibility test that was carried out among the isolates against 12 antimicrobial agents, showed 100 % resistance to only ampicillin and 100 % sensitivity to carbenicillin, chloramphenicol, clindamycin, gentamycin, kanamycin and tetracycline.Conclusion, significance and impact of study: This study confirmed the association of virulent strains of S. enterica serovar Typhi from Typhoidial fever among human population and suggested that PCR based diagnostic could be very useful for the rapid detection of S. Typhi isolates. Present study emphasized the use of antibiotic like chloramphenicol or in combination with other antibiotics for the effective control of S. Typhi.

  17. Molecular epidemiology of Mycobacterium tuberculosis clinical isolates in Southwest Ireland.

    LENUS (Irish Health Repository)

    Ojo, Olabisi O

    2010-10-01

    Tuberculosis has had significant effects on Ireland over the past two centuries, causing persistently higher morbidity and mortality than in neighbouring countries until the last decade. This study describes the results of genotyping and drug susceptibility testing of 171 strains of Mycobacterium tuberculosis complex isolated between January 2004 and December 2006 in a region of Ireland centred on the city of Cork. Spoligotype comparisons were made with the SpolDB4 database and clustered 130 strains in 23 groups, forty-one strains showed unique Spoligotyping patterns. The commonest spoligotypes detected were ST0137 (X2) (16.9%), and ST0351 (15.8%) (\\'U\\' clade). The major spoligotype clades were X (26.2%), U (19.3%), T (15.2%), Beijing (5.9%), Haarlem (4.7%), LAM (4.1%), BOVIS (1.75%), with 12.9% unassigned strains. A 24-locus VNTR genotyping produced 15 clusters containing 49 isolates, with high discrimination index (HGDI>0.99). A combination of Spoligotyping and VNTR reduced the number of clustered isolates to 47 in 15 clusters (27.5%). This study identified ST351 as common among Irish nationals, and found a low rate of drug resistance with little evidence of transmission of drug resistant strains. Strain clustering was significantly associated with age under 55 years and Irish nationality. Only strains of Euro-American lineage formed clusters. Molecular typing did not completely coincide with the results of contact investigations.

  18. Molecular identification of Taenia specimens after long-term preservation in formalin.

    Science.gov (United States)

    Jeon, Hyeong-Kyu; Kim, Kyu-Heon; Eom, Keeseon S

    2011-06-01

    The majority of Taenia tapeworm specimens in the museum collections are usually kept in a formalin fixative for permanent preservation mainly for use in morphological examinations. This study aims to improve Taenia tapeworm identification even of one preserved in formalin for a maximum of 81 years. Taenia tapeworms were collected by the parasite collection unit of the Swiss Natural History Museum and from units in Indonesia, Japan and Korea. A small amount of formalin-fixed tissue (100 mg) was crushed in liquid nitrogen and then soaked in a Tris-EDTA buffer for 3-5h. The sample was then digested in SDS and proteinase K (20 mg/ml) for 3-5h at 56 °C. After the addition of proteinase K (20mg/ml), SDS and hexadecyl-trimethyl-ammonium bromide (CTAB), incubation was continued for another 3h at 65 °C. A maximum yield of genomic DNA was obtained from this additional step and the quality of genomic DNA obtained with this extraction method seemed to be independent of the duration of storage time in the formalin fixative. The molecular identification of Taenia tapeworms was performed by using PCR and DNA sequences corresponding to position 80-428 of cox1 gene. T. asiatica was detected in the isolates of Indonesia, Japan and Korea. Improvements in the genomic DNA extraction method from formalin fixed museum collections will help in the molecular identification of parasites. Copyright © 2010 Elsevier Ireland Ltd. All rights reserved.

  19. Numerical Modelling of Rubber Vibration Isolators: identification of material parameters

    NARCIS (Netherlands)

    Beijers, C.A.J.; Noordman, Bram; de Boer, Andries; Ivanov, N.I.; Crocker, M.J.

    2004-01-01

    Rubber vibration isolators are used for vibration isolation of engines at high frequencies. To make a good prediction regarding the characteristics of a vibration isolator in the design process, numerical models can be used. However, for a reliable prediction of the dynamic behavior of the isolator,

  20. Isolation and molecular characterization of free-living amoebae from different water sources in Italy.

    Science.gov (United States)

    Montalbano Di Filippo, Margherita; Santoro, Maristella; Lovreglio, Piero; Monno, Rosa; Capolongo, Carmen; Calia, Carla; Fumarola, Luciana; D'Alfonso, Rossella; Berrilli, Federica; Di Cave, David

    2015-03-24

    Free-living amoebae (FLA) are protozoa ubiquitous in Nature, isolated from a variety of environments worldwide. In addition to their natural distribution, some species have been found to be pathogenic to humans. In the present study a survey was conducted in order to evaluate the presence and to characterize at molecular level the isolates of amoebic organisms collected from different water sources in Italy. A total of 160 water samples were analyzed by culture and microscopic examination. FLA were found in 46 (28.7%) of the investigated water samples. Groundwater, well waters, and ornamental fountain waters were the sources with higher prevalence rates (85.7%, 50.0%, and 45.9%, respectively). Identification of FLA species/genotypes, based on the 18S rDNA regions, allowed to identify 18 (39.1%) Acanthamoeba isolates (genotypes T4 and T15) and 21 (45.6%) Vermamoeba vermiformis isolates. Other FLA species, including Vahlkampfia sp. and Naegleria spp., previously reported in Italy, were not recovered. The occurrence of potentially pathogenic free-living amoebae in habitats related to human population, as reported in the present study, supports the relevance of FLA as a potential health threat to humans.

  1. Molecular Typing of Hospital-Acquired Staphylococcus aureus Isolated from Isfahan, Iran

    Science.gov (United States)

    Havaei, Seyed Asghar; Ghanbari, Fahimeh; Rastegari, Ali Asghar; Khademi, Farzad; Hosseini, Nafiseh; Ebrahimzadeh Namvar, Amirmorteza; Vaez, Hamid; Havaei, Seyed Mehdi; Shahin, Mojtaba

    2014-01-01

    Background. Staphylococcus aureus (S. aureus) is one of the most common pathogens that cause hospital- and community-acquired infections in the world. The use of molecular typing methods is essential for determining the origin of the strains, their clonal relations, and also in epidemiological investigations. The purpose of this study was to determine the prevalence of antibiotic resistant S. aureus isolates and using spa, agr, and SCCmec typing to determine the dominant types in Iran. Material and Method. Fifty isolates of S. aureus were collected from January to May 2010. S. aureus identification was performed by biochemical tests. Disk diffusion method was employed to assess the sensitivity of S. aureus strains to antibiotics and then genetic analysis of bacteria was performed using SCCmec, agr, and spa typing. Results. S. aureus resistance to tetracycline, cefoxitin, clindamycin, ciprofloxacin, gentamicin, Cot: cotrimoxazole, levofloxacin, rifampin, and vancomycin were found to be 36%, 18%, 12%, 12%, 22%, 6%, 6%, and 0%, respectively. The results of this study showed that 16% of the isolates were resistant to methicillin (MRSA) and the majority of isolates were SSC mec type IV. In addition spa and agr typing revealed agr typeI and spa type t7688 to be the most predominant. Conclusion. In this study, spa typing showed 100% reliability and the t7688 spa type had a frequency of 26% compared to the frequency of 0.0% in the Ridom SpaServer. The frequency of t304 spa type was higher than the global average. PMID:27350987

  2. Isolation and Molecular Characterization of Acanthamoeba Strains from Dental Units in Costa Rica.

    Science.gov (United States)

    Retana-Moreira, Lissette; Abrahams-Sandí, Elizabeth; Castro-Artavia, Esteban; Fernández-Sánchez, Ana; Castro-Castillo, Alfredo; Reyes-Batlle, María; Lorenzo-Morales, Jacob

    2015-01-01

    Free-living amoebae are protozoa widely distributed in nature, which can be found in a variety of environments. Four genera are recognized as causal agents of infections in humans and animals: Acanthamoeba, Naegleria, Balamuthia, and Sappinia. In this study, the presence of Acanthamoeba in dental units was determined and the isolates obtained were molecularly characterized; osmotolerance and thermotolerance assays were also performed to evaluate multiplication under these conditions, frequently associated with pathogenicity. The morphological analysis and partial sequencing of the 18S rDNA gene revealed the presence of Acanthamoeba genotype T4 in 14% of the units sampled. Osmotolerance and thermotolerance tests were positive for more than 80% of the isolates. Up to date, this is the first study that reports the detection, identification, and genotyping of Acanthamoeba isolated from dental units in Costa Rica and even in Latin-America. Further assays to determine the potential pathogenicity of these Acanthamoeba isolates are underway. © 2015 The Author(s) Journal of Eukaryotic Microbiology © 2015 International Society of Protistologists.

  3. Isolation and Molecular Characterization of Free-Living Amoebae from Different Water Sources in Italy

    Directory of Open Access Journals (Sweden)

    Margherita Montalbano Di Filippo

    2015-03-01

    Full Text Available Free-living amoebae (FLA are protozoa ubiquitous in Nature, isolated from a variety of environments worldwide. In addition to their natural distribution, some species have been found to be pathogenic to humans. In the present study a survey was conducted in order to evaluate the presence and to characterize at molecular level the isolates of amoebic organisms collected from different water sources in Italy. A total of 160 water samples were analyzed by culture and microscopic examination. FLA were found in 46 (28.7% of the investigated water samples. Groundwater, well waters, and ornamental fountain waters were the sources with higher prevalence rates (85.7%, 50.0%, and 45.9%, respectively. Identification of FLA species/genotypes, based on the 18S rDNA regions, allowed to identify 18 (39.1% Acanthamoeba isolates (genotypes T4 and T15 and 21 (45.6% Vermamoeba vermiformis isolates. Other FLA species, including Vahlkampfia sp. and Naegleria spp., previously reported in Italy, were not recovered. The occurrence of potentially pathogenic free-living amoebae in habitats related to human population, as reported in the present study, supports the relevance of FLA as a potential health threat to humans.

  4. Isolation and Identification of Phenol Degrader Bacteria from Sirjan Golgohar Mine Effluent

    Directory of Open Access Journals (Sweden)

    Mehdi Hassanshhian

    2016-03-01

    Full Text Available Phenol and phenolic compounds are highly toxic substances that are found as monoaromatic compounds in various industrial effluents from oil refineries, petrochemical plants, (coal mines, and phenol resin plants. Their discharge into the environment, especially in water resources, causes serious toxicity. Traditionally, physicochemical methods have been used for the removal of phenol and phenolic compounds. Nowadays, bioremediation is known to be the best method for phenol removal from wastewater. The objective of the present study was twofold: isolation and identification of phenol degrading bacteria in the effluent from Golgohar Mine in Sirjan. For this purpose, samples were collected from different sections at Golgohar Mine and its effluent. Phenol degrading bacteria were isolated via enrichment of the samples in the Bushnell Hass medium with phenol used as the only source of carbon and energy. The predominant phenol degrader bacteria were selected by measuring turbidity at 600 nm. The bacteria were subsequently identified by amplification of 16S rRNA with specific primers and PCR sequencing. In this study, 17 strains of phenol degrader bacteria were isolated in soil and wastewater samples collected from different zones of the mine. Screening methods confirmed that 4 strains exhibit a better capability for phenol degradation as evidenced by their capability to degrade 0.4 g/l of phenol. Molecular identification showed that these bacteria belong to the species Pesudomonas sp, Nitrratireductor sp., and Salegentibacter sp. The results also show that the effluent from Golgohar Mine in Sirjan contains many phenol degrading bacteria. The use of these bacteria in the treatment process may lead to a significant reduction in phenol pollution in the mineral effluent.

  5. Isolation, identification and characterization of Bacillus amyloliquefaciens BZ-6, a bacterial isolate for enhancing oil recovery from oily sludge.

    Science.gov (United States)

    Liu, Wuxing; Wang, Xiaobing; Wu, Longhua; Chen, Mengfang; Tu, Chen; Luo, Yongming; Christie, Peter

    2012-06-01

    Over 100 biosurfactant-producing microorganisms were isolated from oily sludge and petroleum-contaminated soil from Shengli oil field in north China. Sixteen of the bacterial isolates produced biosurfactants and reduced the surface tension of the growth medium from 71 to identification, isolate BZ-6 was identified as Bacillus amyloliquefaciens. The biosurfactant produced by isolate BZ-6 was purified and analyzed by high performance liquid chromatography-electrospray ionization tandem mass spectrometry. There were four ion peaks representing four different fengycin A homologues. Copyright © 2012. Published by Elsevier Ltd.

  6. Molecular toolbox for the identification of unknown genetically modified organisms.

    Science.gov (United States)

    Ruttink, Tom; Demeyer, Rolinde; Van Gulck, Elke; Van Droogenbroeck, Bart; Querci, Maddalena; Taverniers, Isabel; De Loose, Marc

    2010-03-01

    Competent laboratories monitor genetically modified organisms (GMOs) and products derived thereof in the food and feed chain in the framework of labeling and traceability legislation. In addition, screening is performed to detect the unauthorized presence of GMOs including asynchronously authorized GMOs or GMOs that are not officially registered for commercialization (unknown GMOs). Currently, unauthorized or unknown events are detected by screening blind samples for commonly used transgenic elements, such as p35S or t-nos. If (1) positive detection of such screening elements shows the presence of transgenic material and (2) all known GMOs are tested by event-specific methods but are not detected, then the presence of an unknown GMO is inferred. However, such evidence is indirect because it is based on negative observations and inconclusive because the procedure does not identify the causative event per se. In addition, detection of unknown events is hampered in products that also contain known authorized events. Here, we outline alternative approaches for analytical detection and GMO identification and develop new methods to complement the existing routine screening procedure. We developed a fluorescent anchor-polymerase chain reaction (PCR) method for the identification of the sequences flanking the p35S and t-nos screening elements. Thus, anchor-PCR fingerprinting allows the detection of unique discriminative signals per event. In addition, we established a collection of in silico calculated fingerprints of known events to support interpretation of experimentally generated anchor-PCR GM fingerprints of blind samples. Here, we first describe the molecular characterization of a novel GMO, which expresses recombinant human intrinsic factor in Arabidopsis thaliana. Next, we purposefully treated the novel GMO as a blind sample to simulate how the new methods lead to the molecular identification of a novel unknown event without prior knowledge of its transgene

  7. Isolation and Identification of miRNAs in Jatropha curcas

    Science.gov (United States)

    Wang, Chun Ming; Liu, Peng; Sun, Fei; Li, Lei; Liu, Peng; Ye, Jian; Yue, Gen Hua

    2012-01-01

    MicroRNAs (miRNAs) are small noncoding RNAs that play crucial regulatory roles by targeting mRNAs for silencing. To identify miRNAs in Jatropha curcas L, a bioenergy crop, cDNA clones from two small RNA libraries of leaves and seeds were sequenced and analyzed using bioinformatic tools. Fifty-two putative miRNAs were found from the two libraries, among them six were identical to known miRNAs and 46 were novel. Differential expression patterns of 15 miRNAs in root, stem, leave, fruit and seed were detected using quantitative real-time PCR. Ten miRNAs were highly expressed in fruit or seed, implying that they may be involved in seed development or fatty acids synthesis in seed. Moreover, 28 targets of the isolated miRNAs were predicted from a jatropha cDNA library database. The miRNA target genes were predicted to encode a broad range of proteins. Sixteen targets had clear BLASTX hits to the Uniprot database and were associated with genes belonging to the three major gene ontology categories of biological process, cellular component, and molecular function. Four targets were identified for JcumiR004. By silencing JcumiR004 primary miRNA, expressions of the four target genes were up-regulated and oil composition were modulated significantly, indicating diverse functions of JcumiR004. PMID:22419887

  8. Identification of Malassezia Species Isolated from Patients with Pityriasis Versicolor Using PCR-RFLP Method in Markazi Province, Central Iran.

    Science.gov (United States)

    Didehdar, Mojtaba; Mehbod, Amir Sayed Ali; Eslamirad, Zahra; Mosayebi, Mahdi; Hajihossein, Reza; Ghorbanzade, Behzad; Khazaei, Mahmoud Reza

    2014-05-01

    The lipophilic yeasts of Malassezia species are members of the normal skin microbial that are cause of pityriasis versicolor. Pityriasis versicolor is a common superficial fungal infection with world-wide distribution. The phenotypic methods for identification of Malassezia species usually are time consuming and unreliable to differentiate newly identified species. But DNA-based techniques rapidly and accurately identified Malassezia species. The purpose of this study was isolation and identification of Malassezia Species from patients with pityriasis versicolor by molecular methods in Markazi Province, Central Iran in 2012. Mycologic examinations including direct microscopy and culture were performed on clinical samples. DNA extraction was performed from colonies. The ITS1 region of rDNA from isolates of Malassezia species were amplified by PCR reaction. The PCR were digested by Cfo I enzyme. From 70 skin samples, were microscopically positive for Malassezia elements, 60 samples were grown on culture medium (85.7%). Using PCR-RFLP method, that was performed on 60 isolates, 37(61.6%) M. globosa, 14(23.3%) M. furfur, 5(8.4%) M. sympodialis and 4(6.7%) M. restrictawere identified. In one case was isolated M. globosa along with M. restricta. The PCR-RFLP method is a useful and reliable technique for identification of differentiation of Malas-sezia species.

  9. Isolation and Molecular Characterization of Brucella Isolates in Cattle Milk in Uganda

    Directory of Open Access Journals (Sweden)

    Denis Rwabiita Mugizi

    2015-01-01

    Full Text Available Brucellosis is endemic in livestock and humans in Uganda and its transmission involves a multitude of risk factors like consumption of milk from infected cattle. To shed new light on the epidemiology of brucellosis in Uganda the present study used phenotypic and molecular approaches to delineate the Brucella species, biovars, and genotypes shed in cattle milk. Brucella abortus without a biovar designation was isolated from eleven out of 207 milk samples from cattle in Uganda. These isolates had a genomic monomorphism at 16 variable number tandem repeat (VNTR loci and showed in turn high levels of genetic variation when compared with other African strains or other B. abortus biovars from other parts of the world. This study further highlights the usefulness of MLVA as an epidemiological tool for investigation of Brucella infections.

  10. Molecular Characterization of Yeast Strains Isolated from Different Sources by Restriction Fragment Length Polymorphism

    International Nuclear Information System (INIS)

    Ali, M. S.; Latif, Z.

    2016-01-01

    Various molecular techniques like analysis of the amplified rDNA internal transcribed spacers (ITS), intragenic spacers and total ITS region analysis by restriction fragment length polymorphism (RFLP) has been introduced for yeast identification but there are limited databases to identify yeast species on the basis of 5.8S rDNA. In this study, twenty nine yeast strains from various sources including spoiled fruits, vegetables, foodstuffs, and concentrated juices were characterized by PCR-RFLP. PCR-RFLP has been used to characterize yeasts present in different spoiled food samples after isolation of the yeasts. By using this technique, the isolated yeast strains were characterized by direct 5.8S-ITS rDNA region amplification. RFLP analysis was applied to each of the amplification products (varied from 400bp to 800bp) detected, and the corresponding yeast identifications were made according to each specific restriction patterns obtained after treatment with two endonucleases TaqI and HaeIII which yielded a specific banding pattern for each species. For further confirmation amplified products of eleven selected isolates were sequenced and blast on NCBI. Both RFLP and sequence analyses of the strains with accession nos. KF472163, KF472164, KF472165, KF472166, KF472167, KF472168, KF472169, KF472170, KF472171, KF472172, KF472173 gave significantly similar results. The isolates were found to belong five different yeast species including; Candida spp., Pichia spp., Kluyveromyces spp., Clavispora spp. and Hanseniaspora spp. This method provides a fast, easy, reliable and authentic way for determining yeast population present in different type of samples, as compared to traditional characterization technique. (author)

  11. Molecular identification of environmental bacteria in indoor air in the domestic home: description of a new species of Exiguobacterium.

    Science.gov (United States)

    Yuan, Ivan; Xu, Jiru; Millar, B Cherie; Dooley, James S G; Rooney, Paul J; Alexander, H Denis; Moore, John E

    2007-02-01

    The quality of indoor air in terms of its bioaerosol composition with microorganisms is important due to its potential aetiological role in development of conditions such as Sick Building Syndrome. Hence, laboratory identification of bacteriological components in any bioaerosol from buildings may help elucidate the role of such organisms in disease states, particularly allergy-related conditions. A molecular method was developed employing universal or "broad-range" eubacterial PCR to help identify environmental culturable bacteria from domestic household air. In a "proof of concept" experiment, 16S rDNA PCR was performed on a collection of bacterial isolates originating from indoor air in the domestic home. 16S rDNA PCR was performed using a set of universal primers to successfully generate an amplicon of approximately 1400 bp, which was sequenced to obtain each isolate's identity. Sequence analysis was able to identify 12/13 of the isolates, whereby the majority were Gram-positive (12/13). Nine different genera were identified from the 13 isolates examined, of which, 12/13 were Gram-positive, with the exception being Moraxella osloensis, which was Gram-negative, as well as a novel species of Exiguobacterium. The closest phylogenetic neighbour of the wildtype isolate to a named species within this genus was E. aestuarii (1364/1384 bases; 98.4% homology), followed by E. marinum (97.5%) and with E. acetylicum being the most distantly related of all the described species. On account of this divergence within the 16S rDNA gene operon of the unknown Exiguobacterium isolate, we believe this isolate to represent a novel species of Exiguobacterium, which we have tentatively named Exiguobacterium belfastensis. Although from this study, these organisms are usually unlikely to be clinically significant to healthy individuals with a competent immune system, we recommend that molecular identification methods are used, if considered necessary, as an adjunct to first line

  12. Isolation and Molecular Characterization of Circulating Melanoma Cells

    Directory of Open Access Journals (Sweden)

    Xi Luo

    2014-05-01

    Full Text Available Melanoma is an invasive malignancy with a high frequency of blood-borne metastases, but circulating tumor cells (CTCs have not been readily isolated. We adapted microfluidic CTC capture to a tamoxifen-driven B-RAF/PTEN mouse melanoma model. CTCs were detected in all tumor-bearing mice and rapidly declined after B-RAF inhibitor treatment. CTCs were shed early from localized tumors, and a short course of B-RAF inhibition following surgical resection was sufficient to dramatically suppress distant metastases. The large number of CTCs in melanoma-bearing mice enabled a comparison of RNA-sequencing profiles with matched primary tumors. A mouse melanoma CTC-derived signature correlated with invasiveness and cellular motility in human melanoma. CTCs were detected in smaller numbers in patients with metastatic melanoma and declined with successful B-RAF-targeted therapy. Together, the capture and molecular characterization of CTCs provide insight into the hematogenous spread of melanoma.

  13. [Identification of Campylobacter spp. isolates with phenotypic methods and multiplex polymerase chain reaction and their antibiotic susceptibilities].

    Science.gov (United States)

    Kayman, Tuba; Abay, Seçil; Hızlısoy, Harun

    2013-04-01

    The aims of this study were to detect the frequency of isolation of thermophilic Campylobacter spp. from acute gastroenteritis cases by phenotypic and molecular methods and to evaluate the antibiotic susceptibilities of the isolates. A total of 3287 stool samples obtained from diarrheal patients who were admitted to Kayseri Training and Research Hospital, Kayseri, Turkey, between March 2010 - March 2011 and sent to the microbiology laboratory, were included in the study. Cefoperazone, amphotericin B and teicoplanin (CAT) supplemented Campylobacter Blood-free Selective Medium (modified CCDA-Preston, Oxoid CM739, UK) was used for the isolation of Campylobacter spp. The media inoculated with stool samples were incubated at 42°C microaerobically for 72 to 96 hours. The genus and species level identifications of the thermophilic Campylobacter spp. isolates defined by phenotypic tests were carried out with multiplex polymerase chain reaction (mPCR). Antibiotic susceptibilities of the isolates were detected by disc diffusion method and the results were evaluated according to the CLSI guidelines. The thermophilic Campylobacter spp. were isolated from 5.4% (179/3287) of the patients' stool samples. Of Campylobacter positive cases 71% (127/179) were children and 58% (104/179) were male. The prevalence rate was estimated as 7.5% (127/1683) for children and 3.2% (52/1604) for adults. Of the isolates, 146 (82%) were identified as C.jejuni, 24 (13%) were C.coli, 6 (3%) were C.lari and 3 (2%) were C.upsaliensis with phenotypic tests. By using mPCR, 152 (85%) and 27 (15%) of 179 isolates were identified as C.jejuni and C.coli, respectively. Three of the six isolates identified as C.lari by the phenotypic methods were identified as C.jejuni and the remaining three as C.coli by mPCR. Phenotypically identified three C.upsaliensis isolates were shown to be C.jejuni (n= 2) and C.coli (n= 1). On the other hand one C.coli isolate was found to be C.jejuni by mPCR. The rates of resistance

  14. Molecular identification of Malaysian Chrysomya megacephala (Fabricius) and Chrysomya rufifacies (Macquart) using life stage specific mitochondrial DNA.

    Science.gov (United States)

    Kavitha, R; Tan, T C; Lee, H L; Nazni, W A; Sofian, A M

    2013-06-01

    DNA identification of blow fly species can be a very useful tool in forensic entomology. One of the potential benefits that mitochondrial DNA (mtDNA) has offered in the field of forensic entomology is species determination. Conventional identification methods have limitations for sibling and closely related species of blow fly and stage and quality of the specimen used. This could be overcome by DNA-based identification methods using mitochondrial DNA which does not demand intact or undamaged specimens. Mitochondrial DNA is usually isolated from whole blow fly and legs. Alternate sources for mitochondrial DNA isolation namely, egg, larva, puparium and empty puparium were explored in this study. The sequence of DNA obtained for each sample for every life cycle stage was 100% identical for a particular species, indicating that the egg, 1st instar, 2nd instar, 3rd instar, pupa, empty puparium and adult from the same species and obtained from same generation will exhibit similar DNA sequences. The present study also highlighted the usefulness of collecting all life cycle stages of blow fly during crime scene investigation with proper preservation and subsequent molecular analysis. Molecular identification provides a strong basis for species identification and will prove an invaluable contribution to forensic entomology as an investigative tool in Malaysia.

  15. Renal biopsy-driven molecular target identification in glomerular disease.

    Science.gov (United States)

    Lindenmeyer, Maja T; Kretzler, Matthias

    2017-08-01

    Chronic kidney disease has severe impacts on the patient and represents a major burden to the health care systems worldwide. Despite an increased knowledge of pathophysiological processes involved in kidney diseases, the progress in defining novel treatment strategies has been limited. One reason is the descriptive disease categorization used in nephrology based on clinical findings or histopathological categories irrespective of potential different molecular disease mechanisms. To accelerate progress toward a targeted treatment, a definition of human disease extending from phenotypic disease classification to mechanism-based disease definitions is needed. In recent years, we have witnessed a major transition in biomedical research from a single gene research to an information rich and collaborative science. Tissue-based analysis in renal disease allows to link structure to molecular function. In our review, we introduce the concept of precision medicine in nephrology, describe several large cohort studies established for molecular analysis of kidney diseases, and highlight examples of renal biopsy-driven target identification by integrative systems biology approaches. Furthermore, we give an outlook on how the new disease definitions can be used for patient stratification in clinical trial design. Finally, we introduce the concept of an informational commons of renal precision medicine for joint analyses of large-scale data sets in renal failure.

  16. COMPARATIVE ANALYSIS OF METHODS FOR IDENTIFICATION OF NONTUBERCULOUS MYCOBACTERIA ISOLATED FROM CLINICAL MATERIAL

    Directory of Open Access Journals (Sweden)

    A. V. Lyamin

    2017-01-01

    Full Text Available Recently there has been a significant increase in the incidence of mycobacteriosis, which is due to an increase in the proportion of immunosuppressed patients, the presence of these various comorbid conditions, as well as the improvement of diagnostic methods. Selecting the most accurate method of identification is extremely important in determining treatment strategy of patients. The aim of the study was to conduct a comparative analysis of modern methods of identification NTMB isolated from clinical specimens in 2015 in the Samara region. The work was carried out identification of 78 strains of microorganisms. Laboratory diagnosis was carried out using the DNA hybridization method and MALDI-ToF mass spectrometry. When microbial identification using MALDI-ToF mass spectrometry was isolated 16 strains (20.5% M. kansasii; 11 strains (14.1% M. avium and M. fortuitum; 9 strains (11.5% M. gordonae; strain 3 (3.8% M. peregrinum, M. szulgai, M. chimera intracellulare group, strain 2 (2.6% M. abscessus, M. septicum, M. paragordonae, M. senegalence, 1 strain (1.3% M. chelonae, M. frederiksbergense, M. monacense, M. lentiflavum. By using mass spectrometry, it was identified 15 types NTMB compared with 9 types — by DNA hybridization. Full match identification results was observed only in 45 (57.7% strains of divergent strains were found in 16 (20.5%. Most often when using the DNA hybridization method, discrepancy was detected in slow-growing cultures (9 strains with a predominance of microorganisms identified as M. gordonae. Among the representatives of fast-growing NTMB, seven investigations were identified in the identification, more often among representatives of the M. fortuitum and M. peregrinum groups. Particular attention should be paid to the identification of the M. kansasii strain by a molecular genetic method, which mass spectrometry was defined as M. bovis. Both cultures of M. tuberculosis complex, which were identified by MALDI

  17. Identification and Characterization of Yeast Isolates from Pharmaceutical Waste Water

    Directory of Open Access Journals (Sweden)

    Marjeta Recek

    2002-01-01

    Full Text Available In order to develop an efficient an system for waste water pretreatment, the isolation of indigenous population of microorganisms from pharmaceutical waste water was done. We obtained pure cultures of 16 yeast isolates that differed slightly in colony morphology. Ten out of 16 isolates efficiently reduced COD in pharmaceutical waste water. Initial physiological characterization failed to match the 10 yeast isolates to either Pichia anomala or Pichia ciferrii. Restriction analysis of rDNA (rDNA-RFLP using three different restriction enzymes: HaeIII, MspI and CfoI, showed identical patterns of the isolates and Pichia anomala type strain. Separation of chromosomal DNAs of yeast isolates by the pulsed field gel electrophoresis revealed that the 10 isolates could be grouped into 6 karyotypes. Growth characteristics of the 6 isolates with distinct karyotypes were then studied in batch cultivation in pharmaceutical waste water for 80 hours.

  18. Isolation, identification and application in lignin degradation of an ...

    African Journals Online (AJOL)

    This study was undertaken to isolate an ascomycete producing ligninolytic enzyme and characterize its lignin degradation capability. Among 20 isolates, GHJ-4 was isolated from decayed wood of Salix matsudana Koidz in Mount Tai, China, by different indicator compounds assay. The taxonomy of the fungi was ...

  19. Molecular approaches for the detection and identification of bifidobacteria and lactobacilli in the human gastrointestinal tract

    NARCIS (Netherlands)

    Satokari, R.M.; Vaughan, E.E.; Smidt, H.; Saarela, M.; Matto, J.; Vos, de W.M.

    2003-01-01

    In this review an overview of various molecular techniques and their application for the detection and identification of bifidobacteria and lactobacilli in the gastrointestinal (GI) tract is presented. The techniques include molecular typing techniques such as amplified ribosomal DNA restriction

  20. Identification of unusual Campylobacter-like isolates from poultry products as Helicobacter pullorum

    DEFF Research Database (Denmark)

    Atabay, H.I.; Corry, J.E.L.; On, Stephen L.W.

    1998-01-01

    Twenty-six unclassified Campylobacter-like strains previously isolated from 15 chicken carcasses and caecal contents, together with two more strains isolated from chicken faeces on a different occasion, were identified as Helicobacter pullorum using various phenotypic identification methods. API...... Campy identification kits and a 16-test identification scheme developed for campylobacters failed to identify these bacteria, or identified them as Campylobacter spp. Eighteen strains (including the two isolated on a different occasion) were chosen for examination using a more comprehensive......%) suggests that routine isolation and identification methods should be amended to enable a thorough evaluation of its role in human gastroenteritis and avian hepatitis. Some phenotypic characters useful in identifying poultry campylobacteria are presented which could be utilized, along with other technique...

  1. Molecular identification of Giardia lamblia isolates from adult human ...

    African Journals Online (AJOL)

    ARL

    2013-02-27

    Feb 27, 2013 ... Giardia lamblia is a flagellated protozoa that infects the intestinal tract of a wide range of mammalian hosts, including both wild and domestic animals as well as humans. ... Person-to-person, zoonotic, foodborne and waterborne transmissions may occur through the fecal-oral route after contact with the ...

  2. Molecular identification of Giardia lamblia isolates from adult human ...

    African Journals Online (AJOL)

    ... reflect environmental contamination of water resources. Therefore, it seems further studies are needed to clarify the route of infection in the study area. Keywords: Giardia lamblia, glutamate dehydrogenase (gdh), semi-nested polymerase chain reaction (PCR), PCR-RFLP, Ahvaz, Iran African Journal of Biotechnology Vol.

  3. Isolation and molecular identification of β-carotene producing strains ...

    African Journals Online (AJOL)

    Dunaliella salina and Dunaliella bardawil are unique species of the genus Dunaliella that produce large amounts of â-carotene when cultivated under appropriate conditions. These include high light intensity, high sodium chloride concentration, nitrate deficiency and extreme temperatures. Under these conditions, only D.

  4. Isolation and molecular identification of a Serratia strain from ...

    African Journals Online (AJOL)

    ONOS

    2010-04-05

    Apr 5, 2010 ... ribosomal RNA (16S rRNA) gene sequence. Key words: Serratia, tree shrew, 16S rRNA gene. INTRODUCTION. The genus Serratia belongs to the family Entero- bacteriaceae and consists of the recognized species: Serratia marcescens, Serratia liquefaciens, Serratia ficaria, Serratia rubidaea, Serratia ...

  5. Isolation and molecular identification of Vibrio spp. by sequencing of ...

    African Journals Online (AJOL)

    Out of the 93 cultured samples only 48 (51.6%) yielded colonies on Thiosulfate Citrate Bile Salt agar (TCBS) with culture characteristics of Vibrio spp. More than half (n=27) of processed seafood samples (n=46) yielded colonies on TCBS, while only 44.6% of samples of meat and meat products showed colonies on TCBS.

  6. Identification of Stenotrophomonas maltophilia strains isolated from environmental and clinical samples: a rapid and efficient procedure.

    Science.gov (United States)

    Pinot, C; Deredjian, A; Nazaret, S; Brothier, E; Cournoyer, B; Segonds, C; Favre-Bonté, S

    2011-11-01

    Aim of the study is to identify accurately Stenotrophomonas maltophilia isolates recovered from environmental and clinical samples. Recovery of Sten. maltophilia-like isolates from soil samples using the vancomycin, imipenem, amphotericin B (VIA) selective agar medium enabled distinction of various morphotype colonies. A set of soil and clinical isolates was tested for species identification using different methods. 16S rDNA analyses showed the dark green with a blue halo morphotype to be typical Sten. maltophilia strains. The API-20NE, Vitek-2 and Biolog phenotypic analyses typically used for the identification of clinical isolates did not perform well on these soil isolates. The species-specific PCR screening targeting Sten. maltophilia 23S rDNA and the multiplex smeD/ggpS PCR, differentiating Sten. maltophilia from Stenotrophomonas rhizophila, were tested for improvement of these identification schemes. The latter multiplex PCR identified all isolates tested in this study, whatever be their origin. Isolation on VIA medium and confirmation of Sten. maltophilia species membership by smeD PCR is proposed to identify environmental and clinical isolates of Sten. maltophilia. The proposed approach enables isolation and identification of Sten. maltophilia from different environments in an easy and rapid way. This approach will be useful to accurately manage studies on the abundance and distribution of Sten. maltophilia in hospital and nonhospital environments. © 2011 The Authors. Journal of Applied Microbiology © 2011 The Society for Applied Microbiology.

  7. First molecular identification of Entamoeba moshkovskii in Malaysia.

    Science.gov (United States)

    Anuar, Tengku Shahrul; Al-Mekhlafi, Hesham M; Ghani, Mohamed Kamel Abdul; Azreen, Siti Nor; Salleh, Fatmah Md; Ghazali, Nuraffini; Bernadus, Mekadina; Moktar, Norhayati

    2012-10-01

    Entamoeba moshkovskii and Entamoeba dispar are microscopically indistinguishable from the pathogenic species Entamoeba histolytica. Although sporadic cases of human infection with E. moshkovskii have been reported, the amoeba is still considered primarily as a free-living amoeba. A cross-sectional study was carried out among Orang Asli communities in 3 different states of Peninsular Malaysia. Fecal samples were examined by formalin-ether sedimentation and trichrome staining techniques and then single-round PCR assay was used to detect E. moshkovskii. Out of 500 fecal samples examined microscopically, 93 (18·6%) samples were positive for E. histolytica/E. dispar/E. moshkovskii complex cysts and/or trophozoites. PCR products were detected in 106 fecal samples. E. moshkovskii isolates were detected in 13 (12·3%) fecal samples. Of the 13 E. moshkovskii-positive samples, 5 were of single isolation of E. moshkovskii, 6 were also positive for E. dispar, and only 2 samples were positive for E. dispar and E. histolytica. Moreover, 3 E. moshkovskii-positive samples were collected from symptomatic individuals while the remaining 10 samples were from asymptomatic subjects. This is the first report on the identification of E. moshkovskii in Malaysia. Further studies are needed to confirm the pathogenicity of E. moshkovskii infection and determine the epidemiology among Orang Asli communities in Malaysia.

  8. Isolation and molecular characterization of polyvinyl chloride (PVC) plastic degrading fungal isolates.

    Science.gov (United States)

    Ali, Muhammad Ishtiaq; Ahmed, Safia; Robson, Geoff; Javed, Imran; Ali, Naeem; Atiq, Naima; Hameed, Abdul

    2014-01-01

    The recalcitrant nature of polyvinyl chloride creates serious environmental concerns during manufacturing and waste disposal. The present study was aimed to isolate and screen different soil fungi having potential to biodegrade PVC films. After 10 months of soil burial experiment, it was observed that a number of fungal strains were flourishing on PVC films. On morphological as well as on 18rRNA gene sequence and phylogenetic basis they were identified as Phanerochaete chrysosporium PV1, Lentinus tigrinus PV2, Aspergillus niger PV3, and Aspergillus sydowii PV4. The biodegradation ability of these fungal isolates was further checked in shake flask experiments by taking thin films of PVC (C source) in mineral salt medium. A significant change in color and surface deterioration of PVC films was confirmed through visual observation and Scanning electron microscopy. During shake flask experiments, P. chrysosporium PV1 produced maximum biomass of about 2.57 mg ml(-1) followed by A. niger PV3. P. chrysosporium PV1 showed significant reduction (178,292 Da(-1)) in Molecular weight of the PVC film than control (200,000 Da(-1)) by gel permeation chromatography. Furthermore more Fourier transform infrared spectroscopy and nuclear magnetic resonance also revealed structural changes in the PVC. It was concluded that isolated fungal strains have significant potential for biodegradation of PVC plastics. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  9. Morphological and molecular identification of Fusarium tricinctum and Fusarium acuminatum as causal agents of garlic bulbs rot in Serbia

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    Ignjatov Maja V.

    2017-01-01

    Full Text Available Garlic (Allium sativum L. is considered to be one of the oldest crops in the world. During 2016, infected garlic bulbs occurred in storages on several localities of the Province of Vojvodina. Symptomatic cloves showed typical rot symptoms such as softened and spongy areas covered with white fungal growth with deep lesions formed on the cloves which became dry over time. A total of 36 isolates of Fusarium species were obtained from diseased cloves of garlic. Colony morphology and microscopic properties of isolated Fusarium species were recorded from the cultures grown on PDA and CLA, respectively. Identification of two chosen isolates was performed by sequencing the EF-1α gene. The TEF sequence of isolate JBL12 showed 100% similarity with several F. tricinctum sequences and sequence of JBL539 showed 99% identity with several F. acuminatum sequences and they were deposited in the NCBI GenBank. Based on the results of the morphological and molecular identification, isolates JBL12 and JBL539 were identified as F. tricinctum and F. acuminatum, respectively, as new causal agents of garlic bulbs rot in Serbia. Specific primers were designed for the PCR identification of the F. tricinctum. [Project of the Serbian Ministry of Education, Science and Technological Development, Grant no. TR31030

  10. Molecular identification of Bigeyes (Perciformes, Priacanthidae) from Indian waters.

    Science.gov (United States)

    K K, Bineesh; A, Gopalakrishnan; J K, Jena; V S, Basheer; C, Mohitha; N, Vineesh; M, Joselet; Pillai, N G K

    2016-11-01

    Thirty-five individuals of six priacanthid fish species were sampled from different localities along the coast of India covering the Arabian Sea and Bay of Bengal. The partial sequence of 16S rRNA and cytochrome oxidase subunit I (COI) genes were analyzed for species identification and phylogenetic relationship among the Indian priacanthids (Priacanthus hamrur, P. prolixus, P. blochii, P. sagittarius, Cookeolus japonicus, and Pristigenys refulgens). The intraspecies genetic distance ranged from 0.000 to 0.002, while distances varied from 0.008 to 0.157 interspecies based on 16S sequences. Using COI data analysis, the intraspecies genetic distance ranged from 0.000 to 0.005, while interspecies distances varied from 0.009 to 0.108. Several sequences labeled Priacanthus hamrur in GenBank are shown to be P. prolixus. We also observed cryptic speciation in Heteropriacanthus cruentatus. Partial sequences of 16S rRNA and COI genes provided phylogenetic information to distinguish thirteen species of priacanthids, indicating the usefulness of molecular markers in species identification.

  11. Development of a novel, simple and rapid molecular identification system for clinical Candida species

    NARCIS (Netherlands)

    Deak, R.; Bodia, L.; Aarts, H.J.M.; Maraz, A.

    2004-01-01

    Identification of clinical yeast isolates causing candidiasis is routinely performed by commercial yeast identification systems based on biochemical, morphological and physiological tests. These systems require 3-5 days and the proportion of identifications that are incorrect is high. Our novel and

  12. MALDI-TOF mass spectrometry proteomic based identification of clinical bacterial isolates

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    Ashutosh Panda

    2014-01-01

    Full Text Available Background & objectives: Pathogenic bacteria often cause life threatening infections especially in immunocompromised individuals. Therefore, rapid and reliable species identification is essential for a successful treatment and disease management. We evaluated a rapid, proteomic based technique for identification of clinical bacterial isolates by protein profiling using matrix-assisted laser desorption-ionization time - of - flight mass spectrometry (MALDI-TOF MS. Methods: Freshly grown bacterial isolates were selected from culture plates. Ethanol/formic acid extraction procedure was carried out, followed by charging of MALDI target plate with the extract and overlaying with α-cyano-4 hydroxy-cinnamic acid matrix solution. Identification was performed using the MALDI BioTyper 1.1, software for microbial identification (Bruker Daltonik GmbH, Bremen, Germany. Results: A comparative analysis of 82 clinical bacterial isolates using MALDI -TOF MS and conventional techniques was carried out. Amongst the clinical isolates, the accuracy at the species level for clinical isolates was 98.78%. One out of 82 isolates was not in accordance with the conventional assays because MALDI-TOF MS established it as Streptococcus pneumoniae and conventional methods as Streptococcus viridans. Interpretation & conclusions: MALDI - TOF MS was found to be an accurate, rapid, cost-effective and robust system for identification of clinical bacterial isolates. This innovative approach holds promise for earlier therapeutic intervention leading to better patient care.

  13. Molecular identification and prevalence of malassezia species in pityriasis versicolor patients from kashan, iran.

    Science.gov (United States)

    Talaee, Rezvan; Katiraee, Farzad; Ghaderi, Maryam; Erami, Mahzad; Kazemi Alavi, Azam; Nazeri, Mehdi

    2014-08-01

    Malassezia species are lipophilic yeasts found on the skin surface of humans and other warm-blooded vertebrates. It is associated with various human diseases, especially pityriasis versicolor, which is a chronic superficial skin disorder. The aim of the present study was to identify Malassezia species isolated from patients' samples affected by pityriasis versicolor, using molecular methods in Kashan, Iran. A total of 140 subjects, suspected of having pityriasis versicolor from Kashan, were clinically diagnosed and then confirmed by direct microscopic examination. The scraped skin specimens were inoculated in modified Dixon's medium. DNA was extracted from the colonies and PCR amplification was carried out for the 26s rDNA region. PCR products were used to further restriction fragment length polymorphism by CfoI enzyme. Direct examination was positive in 93.3% of suspected pityriasis versicolor lesions. No statistically significant difference was observed in the frequency of Malassezia species between women and men. The highest prevalence of tinea versicolor was seen in patients 21-30 years-of-age. No difference could be seen in the frequency of Malassezia species depending on the age of the patients. In total, 65% of patients with pityriasis versicolor had hyperhidrosis. The most commonly isolated Malassezia species in the pityriasis versicolor lesions were; Malassezia globosa (66%), M. furfur (26%), M. restricta (3%), M. sympodialis (3%), and M. slooffiae (2%). Malassezia species were mainly isolated from the neck and chest. This study showed M. globosa to be the most common Malassezia species isolated from Malassezia skin disorders in Kashan, Iran. The PCR-RFLP method was useful in the rapid identification of the Malassezia species. By using these methods, the detection and identification of individual Malassezia species from clinical samples was substantially easier.

  14. Molecular Identification and Prevalence of Malassezia Species in Pityriasis Versicolor Patients From Kashan, Iran

    Science.gov (United States)

    Talaee, Rezvan; Katiraee, Farzad; Ghaderi, Maryam; Erami, Mahzad; Kazemi Alavi, Azam; Nazeri, Mehdi

    2014-01-01

    Background: Malassezia species are lipophilic yeasts found on the skin surface of humans and other warm-blooded vertebrates. It is associated with various human diseases, especially pityriasis versicolor, which is a chronic superficial skin disorder. Objectives: The aim of the present study was to identify Malassezia species isolated from patients’ samples affected by pityriasis versicolor, using molecular methods in Kashan, Iran. Patients and Methods: A total of 140 subjects, suspected of having pityriasis versicolor from Kashan, were clinically diagnosed and then confirmed by direct microscopic examination. The scraped skin specimens were inoculated in modified Dixon’s medium. DNA was extracted from the colonies and PCR amplification was carried out for the 26s rDNA region. PCR products were used to further restriction fragment length polymorphism by CfoI enzyme. Results: Direct examination was positive in 93.3% of suspected pityriasis versicolor lesions. No statistically significant difference was observed in the frequency of Malassezia species between women and men. The highest prevalence of tinea versicolor was seen in patients 21–30 years-of-age. No difference could be seen in the frequency of Malassezia species depending on the age of the patients. In total, 65% of patients with pityriasis versicolor had hyperhidrosis. The most commonly isolated Malassezia species in the pityriasis versicolor lesions were; Malassezia globosa (66%), M. furfur (26%), M. restricta (3%), M. sympodialis (3%), and M. slooffiae (2%). Malassezia species were mainly isolated from the neck and chest. Conclusions: This study showed M. globosa to be the most common Malassezia species isolated from Malassezia skin disorders in Kashan, Iran. The PCR-RFLP method was useful in the rapid identification of the Malassezia species. By using these methods, the detection and identification of individual Malassezia species from clinical samples was substantially easier. PMID:25485051

  15. Isolation and identification mould micoflora inhabiting plant leaf litter from Mount Lawu, Surakarta, Central Java

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    MUHAMMAD ILYAS

    2007-04-01

    Full Text Available A study on isolation and identification mould inhabiting plant leaf litter had been conducted. The objective of the study was to isolate and identify mould inhabiting plant leaf litter from Mount Lawu, Surakarta, Central Java. The mould isolation was based on washing and filtering with membrane isolation method. The result showed that 39 moulds generas with 55 species varians, one group identified in class level, and three groups of unidentified mould isolates had been isolated. Taxas distributions showed that there were endophyte and phytopatogen mould isolates had been isolated such as Fusarium, Pestalotiopsis, Phoma, and Coelomycetes. However, typical soil taxa and common saprobic fungi such as Aspergillus, Cunninghamella, Mucor, Paecilomyces, Penicillium, Rhizopus, and Trichoderma remain dominated the resulted isolates.

  16. Exploration, Isolation, and Identification of Carotenoid from Bacterial Symbiont of Sponge Callyspongia vaginalis

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    Iqna Kamila Abfa

    2017-06-01

    Full Text Available During the past two decades research on marine bacteria has highlighted the tremendous potential of symbiotic-microorganisms as a source of bioactive secondary. One of the potential of the bacterial symbionts is producing a natural pigment, and these organisms can be used as a sustainable source of natural pigments. Carotenoid is one of the most important pigments that has important roles in physiological and molecular processes of microorganisms, as well as for human health. The objective of this study is to analyze carotenoid pigments from marine bacterial symbionts from sponge and to identify bacterial symbionts that produce carotenoid pigments. Pigment analysis was performed by a UV-VIS spectrophotometer and High Performance Liquid Chromatography (HPLC. Molecular bacterial identification was performed based on 16S rDNA sequence. The isolation of bacterial symbionts from C. vaginalison Zobell 2216E medium resulted in one bacterium, CB-SP5, positively synthesized carotenoids. By reverse phase HPLC analysis, the carotenoid pigments in the bacterial symbionts were identified as diadinoxanthin, fucoxanthin, neoxanthin, dinoxanthin, anddiadinochrome. CB-SP5 shared the highest level of 16S rDNA gene sequence similarity with Psychrobacter celer (99%.   Keywords : carotenoid, sponge, bacterial symbiont, 16S rDNA.

  17. Alimentary Tract Bacteria Isolated and Identified with API-20E and Molecular Cloning Techniques from Australian Tropical Fruit Flies, Bactrocera cacuminata and B. tryoni

    OpenAIRE

    Thaochan, N.; Drew, R. A. I.; Hughes, J. M.; Vijaysegaran, S.; Chinajariyawong, A.

    2010-01-01

    Bacteria were isolated from the crop and midgut of field collected Bactrocera cacuminata (Hering) and Bactrocera tryoni (Froggatt) (Diptera: Tephritidae). Two methods were used, firstly isolation onto two types of bacteriological culture media (PYEA and TSA) and identification using the API-20E diagnostic kit, and secondly, analysis of samples using the 16S rRNA gene molecular diagnostic method. Using the API-20E method, 10 genera and 17 species of bacteria in the family Enterobacteriaceae we...

  18. Identification of Bifidobacterium Strains Isolated from Kashk-e Zard: A Traditional Iranian Fermented Cereal-Dairy Based Food

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    Mashak

    2016-09-01

    Full Text Available Objectives The genus Bifidobactrium enjoys considerable significance among the probiotic bacteria for having appropriately adapted to the human gastrointestinal tract. As the properties of Bifidobacteria are strain-oriented and niche-dependent, there is growing interest in studying the different sources of these probiotics. Kashk-e Zard, a traditional fermented food produced from wheat and yogurt through a two-week, two-step fermentation process, is rich in probiotics and is worthy of study in this regard. The present study aimed to identify Bifidobacterium spp. in Kashk-e Zard. Methods Twenty-three samples of Kashk-e Zard were collected and subjected to Bifidobacterium identification experiments. Polymerase chain reaction (PCR and sequencing methods were applied for bacterial identification. Results Twelve of the isolates obtained were G +, rod-shaped, and catalase-, whereas only three of them identified positive for fructose 6-phosphate phosphoketolase (F6PPK a Bifidobacterium specific test and mupirocin resistance. These three isolates were then considered for further identification using the 16SrDNA sequencing technique. Conclusions Although carbohydrate fermentation patterns specified these three isolates as B. infantis, B. bifidum, and B. longum, the molecular results did not confirm B. longum, which is still also controversial in the literature. Overall, our results demonstrated that Kashk-e Zard is a rich potential source of probiotic bacteria and further investigations should be undertaken.

  19. Strategy for identification & characterization of Bartonella henselae with conventional & molecular methods

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    Kavita Diddi

    2013-01-01

    Full Text Available Background & objectives: Bartonella henselae is a fastidious gram-negative bacterium usually causing self limiting infections in immunocompetent individuals but often causes potentially life threatening infection, such as bacillary angiomatosis in immunocompromised patients. Both diagnosis of infections and research into molecular mechanisms of pathogenesis have been hindered by lack of appropriate and reliable diagnostic techniques. We undertook this study to standardize methods to characterize B. henselae in clinical samples to diagnose Bartonella infection correctly. Methods: B. henselae ATCC 49882 strain was procured from American type culture collection, USA. This strain was revived and maintained in the laboratory, and identification and characterization of this strain was done by conventional and molecular techniques, which included culture on various media, staining by different methods including electron microscopy, biochemical analysis by conventional methods and API, polymerase chain reaction (PCR for amplification of citrate synthase gene followed by restriction fragment length polymorphism (RFLP. Results: This organism was biochemically inert due to slow growth and generated unique identification code with API. The amplification of the citrate-synthase gene with primers yielded a 381 bp product followed by specific RFLP profile for B. henselae. Interpretation & conclusions: Bartonella is fastidious and fragile organism and should be handled carefully. Extra effort and careful observation are required to isolate and characterize this organism.

  20. An investigation on non-invasive fungal sinusitis; Molecular identification of etiologic agents

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    Abdolrasoul Mohammadi

    2017-01-01

    Full Text Available Background: Fungal sinusitis is increasing worldwide in the past two decades. It is divided into two types including invasive and noninvasive. Noninvasive types contain allergic fungal sinusitis (AFS and fungus ball. AFS is a hypersensitivity reaction to fungal allergens in the mucosa of the sinonasal tract in atopic individuals. The fungus ball is a different type of noninvasive fungal rhinosinusitis which is delineated as an accumulation of debris and fungal elements inside a paranasal sinus. Fungal sinusitis caused by various fungi such as Aspergillus species, Penicillium, Mucor, Rhizopus, and phaeohyphomycetes. The aim of the present study is to identify fungal species isolated from noninvasive fungal sinusitis by molecular methods. Materials and Methods: During 2015–2016, a total of 100 suspected patients were examined for fungal sinusitis. Functional endoscopic sinus surgery was performed using the Messerklinger technique. Clinical samples were identified by phenotypic and molecular methods. Polymerase chain reaction (PCR sequencing of ITS1-5.8S-ITS2 region and PCR-restriction fragment length polymorphism with Msp I restriction enzyme was performed for molecular identification of molds and yeasts, respectively. Results: Twenty-seven out of 100 suspected cases (27% had fungal sinusitis. Nasal congestion (59% and headache (19% were the most common clinical signs among patients. Fifteen patients (55.5% were male and 12 patients (44.5% were female. Aspergillus flavus was the most prevalent fungal species (26%, followed by Penicillium chrysogenum (18.5% and Candida glabrata species complex (15%. Conclusion: Since clinical manifestations, computed tomography scan, endoscopy, and histopathological findings are very nonspecific in AFS and fungus ball; therefore, molecular investigations are compulsory for precise identification of etiologic agents and appropriate management of these fungal infections.

  1. MOLECULAR IDENTIFICATION OF SARCOPTES SCABIEI VAR. CUNICULI FROM SURABAYA AND MALANG REGIONS OF EAST JAVA

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    Kurnia Desiandura

    2017-12-01

    Full Text Available Scabies is a zoonotic skin disease caused by Sarcoptes scabiei mites. As an emerging/re-emerging parasitic disease, scabies represents a significant global threat to both human and animal health. Numerous cases of scabies in Indonesia have been reported, which support research on the prevalence of S. scabiei. However, most such studies have involved conventional morphological studies, with limited molecular diagnostic studies. The purpose of the present study was the genetic characterization of S. scabiei var. cuniculi in domestic rabbits to generate baseline genotypic data. S. scabiei var. cuniculi was isolated and identified from scabies-infected rabbits from the Surabaya and Malang regions of East Java. Molecular identification was performed using Polymerase Chain Reaction (PCR using specific primers targeting the COX1 gene. We performed COX1 PCR using rabbit isolates of S. scabiei from Indonesia. To the best of our knowledge, no such study had been reported previously. This study was performed in the Laboratory of Veterinary Parasitology, Faculty of Veterinary Medicine and the Tropical Disease Diagnostic Center Laboratory, Universitas Airlangga. The results with agarose gel electrophoresis revealed a 289 bp PCR product amplified from the DNA of S. scabiei isolates from both Surabaya and Malang in accordance with the expected COX1 amplicon size, that indicated a single band 289 bp in length, demonstrating specific detection of S. scabiei var. cuniculi from Surabaya and Malang using COX1 primers. The results were consistent with the calculated amplicon size based on primer positions within the COX1 locus, with the forward primer spanning nucleotides 61–94, and the reverse primer spanning nucleotides 331–350 ( 350 − 61 = 289 bp. PCR genotyping of the isolates yielded an identical nucleotide length of 289 bp. Further studies are required to sequence the amplified fragments for homology assessment.

  2. [Molecular characterization of Echinococcus granulosus isolates obtained from different hosts].

    Science.gov (United States)

    Erdoğan, Emrah; Özkan, Bora; Mutlu, Fatih; Karaca, Serkan; Şahin, İzzet

    2017-01-01

    Echinococcus granulosus is a parasite that can be seen throughout the world. So far, five species of genus Echinococcus have been identified as parasite in people: E.granulosus, E.multilocularis, E.vogeli, E.oligarthrus, E.shiquicus. Larval (metacestod) form of parasite settles in internal organs of hoofed animals (cattle, goats, pigs, horses, sheep, etc.) and human; the adult form is found in small intestine of final host, canine. Disease caused by parasite called as "Cystic echinococcosis" (CE) is an important health problem and causes economic losses in many countries including our country that livestock is common. Infective eggs cause infections in intermediate hosts by taking oral way and rarely inhalation. Received egg opens in the stomach and intestines of intermediate host and oncosphere is released. Oncosphere quickly reaches the lamina propria of the villus epithelium by its histolytic enzymes and hooks. It usually transported from here to the liver and lungs, less frequently, muscle, brain, spleen, kidney and to other organs through the veins. By molecular studies, five species have been validated taxonomically and 10 different variants or strains of E.granulosus have been identified. Host and developmental differences between strains may negatively affect control studies and fight against the parasite. This study aimed to determinate E.granulosus strains obtained from cyst material of different intermediate hosts from different regions of Turkey by molecular methods. In the study, 25 human, 8 cattle, 6 sheep and 2 goat cysts material has been collected. Total genomic DNA was isolated from protoscoleces in cyst fluid and analyzed by PCR with COX-1 (L) and COX-1 (S) genes specific primers. DNA sequence analysis for each PCR product has been made. DNA sequence analysis results evaluated phylogenetically by MEGA analyze and BLAST software. As a result of this study, all isolates were identified as E.granulosus sensu stricto (G1) by DNA sequence analysis. CE

  3. MALDI-TOF MS enables the rapid identification of the major molecular types within the Cryptococcus neoformans/C. gattii species complex.

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    Carolina Firacative

    Full Text Available BACKGROUND: The Cryptococcus neoformans/C. gattii species complex comprises two sibling species that are divided into eight major molecular types, C. neoformans VNI to VNIV and C. gattii VGI to VGIV. These genotypes differ in host range, epidemiology, virulence, antifungal susceptibility and geographic distribution. The currently used phenotypic and molecular identification methods for the species/molecular types are time consuming and expensive. As Matrix-Assisted Laser Desorption Ionization-Time-of-Flight Mass Spectrometry (MALDI-TOF MS offers an effective alternative for the rapid identification of microorganisms, the objective of this study was to examine its potential for the identification of C. neoformans and C. gattii strains at the intra- and inter-species level. METHODOLOGY: Protein extracts obtained via the formic acid extraction method of 164 C. neoformans/C. gattii isolates, including four inter-species hybrids, were studied. RESULTS: The obtained mass spectra correctly identified 100% of all studied isolates, grouped each isolate according to the currently recognized species, C. neoformans and C. gattii, and detected potential hybrids. In addition, all isolates were clearly separated according to their major molecular type, generating greater spectral differences among the C. neoformans molecular types than the C. gattii molecular types, most likely reflecting a closer phylogenetic relationship between the latter. The number of colonies used and the incubation length did not affect the results. No spectra were obtained from intact yeast cells. An extended validated spectral library containing spectra of all eight major molecular types was established. CONCLUSIONS: MALDI-TOF MS is a rapid identification tool for the correct recognition of the two currently recognized human pathogenic Cryptococcus species and offers a simple method for the separation of the eight major molecular types and the detection of hybrid strains within this

  4. Identification of Lactic Acid Bacteria isolated from Opaque beer ...

    African Journals Online (AJOL)

    A study was carried out to identify lactic acid bacteria (LAB) isolated from chibuku that would be later assessed for potential as starter cultures. Thirty-eight isolates were Gram stained and the 20, which were Gram positive, were identified to genus level using morphological, physiological and biochemical tests. Five genera ...

  5. Identification of Candida strains isolated from Tanzanian pregnant ...

    African Journals Online (AJOL)

    Objective: To identify Candida strains isolated from Tanzanian women (13 to 45 years) with vaginal candidiasis. Design: A cross-sectional study. Setting: Antenatal clinic in llala district hospital in Dar es Salaam, Tanzania from March 1998 to December 2000. Results: The identities of the 272 isolates tested with API Candida ...

  6. Isolation and Identification of Spoilage Fungi Associated With Rice ...

    African Journals Online (AJOL)

    The spoilage fungi isolated were Aspergillus species, Rhizopus, Penicilluim, Fusarium, Eurotium, Mucor, Geotrichum, Alternaria, Cladosporium and Actinomyces species. The predominant spoilage fungi in the grains were Aspergillus species. The populations of some spoilage fungi isolated from the grains were not high ...

  7. Screening and identification of lactic acid bacteria isolated from ...

    African Journals Online (AJOL)

    The lactic acid bacteria (LAB) isolated from sorghum (Sorghum bicolor. L.) silage were identified during different periods of evolution of sorghum silage in west Algeria. Morphological, physiological, biochemical and technological techniques were used to characterize lactic acid bacteria isolates. A total number of 27 ...

  8. Isolation and identification of fungal flora associated with groundnut ...

    African Journals Online (AJOL)

    A total of 25 colonies were isolated from all the samples from which 6 fungal species were identified, namely Mucor, Aspergillus, Rhizophus, Curvularia, Pencillium and Fusarium spp. Of these, Mucor and Rhizopus were most prevalent having been isolated from the three storage facilities studied. Curvularia and Penicillium ...

  9. Isolation, identification and screening of potential cellulase-free ...

    African Journals Online (AJOL)

    In order to isolate cellulase-free xylanase producing fungi, screening and isolation was done using composting soil as microbial source. Eight fungal species were selected for further study based on clearing zones formation on agar media containing covalently linked xylan with dye cibacron brilliant red-3BA. Both solid ...

  10. Isolation, identification and screening of potential cellulase-free ...

    African Journals Online (AJOL)

    In order to isolate cellulase-free xylanase-producing fungi, screening and isolation of fungi was done using decaying wood, agricultural wastes and other lignocellulosic wastes as microbial source. Thirty (30) fungal species were selected for further analysis based upon clearing zones formation on xylan enriched agar ...

  11. Identification of Campylobacter pyloridis isolates by restriction endonuclease DNA analysis

    NARCIS (Netherlands)

    Langenberg, W.; Rauws, E. A.; Widjojokusumo, A.; Tytgat, G. N.; Zanen, H. C.

    1986-01-01

    Campylobacter pyloridis isolates recovered from gastric biopsy specimens of 16 patients were examined by restriction endonuclease DNA analysis with HindIII. For 8 of these 16 patients two different isolates were compared to study the persistence of the colonizing strains and the stability of their

  12. Isolation and identification of phenol degrading bacteria from Lake ...

    African Journals Online (AJOL)

    The aim of this study is to isolate and identify phenol degrading bacteria from Lake Parishan and to assay their kinetic growth. Sixty samples of water and sedimentation of different area of Lake Parishan were collected. In order to isolate phenol degrading bacteria, samples were cultured on salt base phenol broth media.

  13. Isolation, Identification, and Characterization of Cadmium Resistant Pseudomonas sp. M3 from Industrial Wastewater

    OpenAIRE

    Syed Zaghum Abbas; Mohd Rafatullah; Norli Ismail; Japareng Lalung

    2014-01-01

    The present study deals with the isolation, identification, and characterization of the cadmium resistant bacteria from wastewater collected from industrial area of Penang, Malaysia. The isolate was selected based on high level of the cadmium and antibiotic resistances. On the basis of morphological, biochemical characteristics, 16S rDNA gene sequencing and phylogeny analysis revealed that the strain RZCd1 was authentically identified as Pseudomonas sp. M3. The industrial isolate showed more ...

  14. Use of PCR-restriction fragment length polymorphism analysis for identification of yeast species isolated from bovine intramammary infection.

    Science.gov (United States)

    Fadda, M E; Pisano, M B; Scaccabarozzi, L; Mossa, V; Deplano, M; Moroni, P; Liciardi, M; Cosentino, S

    2013-01-01

    This study reports a rapid PCR-based technique using a one-enzyme RFLP for discrimination of yeasts isolated from bovine clinical and subclinical mastitis milk samples. We analyzed a total of 1,486 milk samples collected over 1 yr in south Sardinia and northern Italy, and 142 yeast strains were preliminarily grouped based on their cultural morphology and physiological characteristics. Assimilation tests were conducted using the identification kit API ID 32C and APILAB Plus software (bioMérieux, Marcy l'Etoile, France). For PCR-RFLP analysis, the 18S-ITS1-5.8S ribosomal(r)DNA region was amplified and then digested with HaeIII, and dendrogram analysis of RFLP fragments was carried out. Furthermore, within each of the groups identified by the API or PCR-RFLP methods, the identification of isolates was confirmed by sequencing of the D1/D2 region using an ABI Prism 310 automatic sequencer (Applied Biosystems, Foster City, CA). The combined phenotypic and molecular approach enabled the identification of 17 yeast species belonging to the genera Candida (47.9%), Cryptococcus (21.1%), Trichosporon (19.7%), Geotrichum (7.1%), and Rhodotorula (4.2%). All Candida species were correctly identified by the API test and their identification confirmed by sequencing. All strains identified with the API system as Geotrichum candidum, Cryptococcus uniguttulatus, and Rhodotorula glutinis also produced characteristic restriction patterns and were confirmed as Galactomyces geotrichum (a teleomorph of G. candidum), Filobasidium uniguttulatum (teleomorph of Crypt. uniguttulatus), and R. glutinis, respectively, by D1/D2 rDNA sequencing. With regard to the genus Trichosporon, preliminary identification by API was problematic, whereas the RFLP technique used in this study gave characteristic restriction profiles for each species. Moreover, sequencing of the D1/D2 region allowed not only successful identification of Trichosporon gracile where API could not, but also correct identification of

  15. A grass molecular identification system for forensic botany: a critical evaluation of the strengths and limitations.

    Science.gov (United States)

    Ward, Jodie; Gilmore, Simon R; Robertson, James; Peakall, Rod

    2009-11-01

    Plant material is frequently encountered in criminal investigations but often overlooked as potential evidence. We designed a DNA-based molecular identification system for 100 Australian grasses that consisted of a series of polymerase chain reaction assays that enabled the progressive identification of grasses to different taxonomic levels. The identification system was based on DNA sequence variation at four chloroplast and two mitochondrial loci. Seventeen informative indels and 68 single-nucleotide polymorphisms were utilized as molecular markers for subfamily to species-level identification. To identify an unknown sample to subfamily level required a minimum of four markers or nine markers for species identification. The accuracy of the system was confirmed by blind tests. We have demonstrated "proof of concept" of a molecular identification system for trace botanical samples. Our evaluation suggests that the adoption of a system that combines this approach with DNA sequencing could assist the morphological identification of grasses found as forensic evidence.

  16. Pigments Characterization and Molecular Identification of Bacterial Symbionts of Brown Algae Padinasp. Collected from Karimunjawa Island

    Directory of Open Access Journals (Sweden)

    Damar Bayu Murti

    2016-06-01

    Full Text Available The search for carotenoids in nature has been extensively studied because of their applications in foods. One treasure of the biopigment source is symbiotic-microorganisms with marine biota. The advantages of symbiont bacteria are easy to culture and sensitize pigments. The use of symbiont bacteria helps to conserve fish, coral reefs, seagrass, and seaweed. Therefore, the bacteria keeps their existence in their ecosystems. In this study, bacterial symbionts were successfully isolated from brown algae Padina sp. The bacterial symbionts had yellow pigment associated with carotenoids. The pigments were characterized using High Performance Liquid Chromatography (HPLC with a Photo Diode Array (PDA detector. The carotenoid pigments in the bacterial symbionts were identified as dinoxanthin, lutein and neoxanthin. Molecular identification by using a 16S rRNA gene sequence method, reveals that the bacterial symbionts were closely related to Bacillus marisflavi with a homology of 99%. Keywords :carotenoid pigments, brown algae, Padina, bacterial symbionts, 16S rRNA

  17. Identification of new IS711 insertion sites in Brucella abortus field isolates

    Directory of Open Access Journals (Sweden)

    Moriyón Ignacio

    2011-08-01

    Full Text Available Abstract Background Brucellosis is a zoonosis caused by Brucella spp., a group of highly homogeneous bacteria. The insertion sequence IS711 is characteristic of these bacteria, and occurs in variable numbers and positions, but always constant within a given species. This species-associated polymorphism is used in molecular typing and identification. Field isolates of B. abortus, the most common species infecting cattle, typically carry seven IS711 copies (one truncated. Thus far, IS711 transposition has only been shown in vitro and only for B. ovis and B. pinnipedialis, two species carrying a high number of IS711 copies, but never in other Brucella species, neither in vitro nor in field strains. Results We found several B. abortus strains isolated from milk and aborted fetuses that carried additional IS711 copies in two hitherto undescribed insertion sites: one in an intergenic region near to the 3' end of a putative lactate permease gene and the other interrupting the sequence of a marR transcriptional regulator gene. Interestingly, the second type of insertion was identified in isolates obtained repeatedly from the same herd after successive brucellosis outbreaks, an observation that proves the stability and virulence of the new genotype under natural conditions. Sequence analyses revealed that the new copies probably resulted from the transposition of a single IS711 copy common to all Brucella species sequenced so far. Conclusions Our results show that the replicative transposition of IS711 can occur under field conditions. Therefore, it represents an active mechanism for the emergence of genetic diversity in B. abortus thus contributing to intra-species genetic polymorphism.

  18. Identification of new IS711 insertion sites in Brucella abortus field isolates.

    Science.gov (United States)

    Mancilla, Marcos; Ulloa, Marcos; López-Goñi, Ignacio; Moriyón, Ignacio; María Zárraga, Ana

    2011-08-03

    Brucellosis is a zoonosis caused by Brucella spp., a group of highly homogeneous bacteria. The insertion sequence IS711 is characteristic of these bacteria, and occurs in variable numbers and positions, but always constant within a given species. This species-associated polymorphism is used in molecular typing and identification. Field isolates of B. abortus, the most common species infecting cattle, typically carry seven IS711 copies (one truncated). Thus far, IS711 transposition has only been shown in vitro and only for B. ovis and B. pinnipedialis, two species carrying a high number of IS711 copies, but never in other Brucella species, neither in vitro nor in field strains. We found several B. abortus strains isolated from milk and aborted fetuses that carried additional IS711 copies in two hitherto undescribed insertion sites: one in an intergenic region near to the 3' end of a putative lactate permease gene and the other interrupting the sequence of a marR transcriptional regulator gene. Interestingly, the second type of insertion was identified in isolates obtained repeatedly from the same herd after successive brucellosis outbreaks, an observation that proves the stability and virulence of the new genotype under natural conditions. Sequence analyses revealed that the new copies probably resulted from the transposition of a single IS711 copy common to all Brucella species sequenced so far. Our results show that the replicative transposition of IS711 can occur under field conditions. Therefore, it represents an active mechanism for the emergence of genetic diversity in B. abortus thus contributing to intra-species genetic polymorphism.

  19. Molecular Characterization of Vibrio cholerae Isolated From Clinical Samples in Kurdistan Province, Iran.

    Science.gov (United States)

    Ramazanzadeh, Rashid; Rouhi, Samaneh; Shakib, Pegah; Shahbazi, Babak; Bidarpour, Farzam; Karimi, Mohammad

    2015-05-01

    Vibrio cholerae causes diarrhoeal disease that afflicts thousands of people annually. V. cholerae is classified on the basis of somatic antigens into serovars or serogroups and there are at least 200 known serogroup. Two serogroups, O1 and O139 have been associated with epidemic diseases. Virulence genes of these bacteria are OmpW, ctxA and tcpA. Due to the importance of V. cholerae infection and developing molecular diagnostics of this organism in medical and microbiology sciences, this study aimed to describe molecular characterization of V. cholerae isolated from clinical samples using a molecular method. In this study, 48 samples were provided during summer 2013 (late August and early September) by reference laboratory. Samples were assessed using biochemical tests initially. The primer of OmpW, ctxA and tcpA genes was used in Polymerase Chain Reaction (PCR) protocols. Enterobacterial Repetitive Intergenic Consensus (ERIC)-PCR and Repetitive Extragenic Palindromic (REP)-PCR methods were used to subtype V. cholerae. In this study, from a total of 48 clinical stool samples 39 (81.2 %) were positive for V. cholerae in biochemical tests and bacteria culture tests. The PCR results showed that of 39 positive isolates 35 (89.7%), 34 (87.1%) and 37 (94.8%) were positive for ctxA, tcpA and OmpW gene, respectively. Also, in the REP-PCR method with ERIC primer strains were divided into 10 groups. In the REP-PCR method with REP primer, strains were divided into 13 groups. Polymerase chain reaction has specificity and accuracy for identification of the organism and is able to differentiate biotypes. Enterobacterial repetitive intergenic consensus sequence is one of the informative and discriminative methods for the analysis of V. cholerae diversity. The REP-PCR is a less informative and discriminative method compared to other methods for the analysis of V. cholerae diversity.

  20. Isolation and Molecular Screening of Glucansucrase Gene Harboring-Lactic Acid Bacteria

    Directory of Open Access Journals (Sweden)

    Ajitya Kurnia Hermawati

    2010-04-01

    Full Text Available Exopolysaccharides (EPS have been possessed to be used in pharmaceutical, cosmetic and food industries. Lactic acid bacteria (LAB produce a wide variety of exopolysaccharides and have been well reported carrying sucrase genes glucansucrase/ glucosyltransferase (gtf and fructansucrase/fructosyltransferases (ftf, enzymes that are able to produce EPS. In this study, the isolation and screening of EPS producing-LAB (EPS-LAB were carried out on modified de Mann-Rogosa-Sharpe (MRS agar medium supplemented with 10% of sucrose on LAB isolated from various unique sugar containing-foods and -beverages originated from local sources. Besides obtaining EPS-LAB, this study aimed to screen for gtf gene as well as to molecular identify strains by using PCR technique. Degenerate primer pairs DegFor and DegRev which targeted the conserved region of gtf genes catalytic domain were used, whereas LABfw and LABrv were used to molecular identify strains using 16S rRNA gene. An approximately 660 base pairs (bp amplicons which targeted gtf gene were obtained from 13 out of 16 srains chosen. Moreover, from PCR of 16S rRNA gene identification on gtf positive strains result, all strains were molecular identified as LAB after DNA sequencing analysis of 700 bp amplicons by using blastn. A rare EPS-producing LAB were obtain from both foods and beverages i.e. Weissella. Results revealed that strains obtained in this study are potential sources for exploring novel sucrase gene/s and obtain unique EPS polymer product/s.

  1. Molecular epidemiology of Mycobacterium abscessus complex isolates in Ireland.

    Science.gov (United States)

    O'Driscoll, C; Konjek, J; Heym, B; Fitzgibbon, M M; Plant, B J; Ní Chróinín, M; Mullane, D; Lynch-Healy, M; Corcoran, G D; Schaffer, K; Rogers, T R; Prentice, M B

    2016-03-01

    The Mycobacterium abscessus complex are the rapidly growing mycobacteria (RGM) most commonly causing lung disease, especially in cystic fibrosis (CF) patients. Ireland has the world's highest CF incidence. The molecular epidemiology of M. abscessus complex in Ireland is unreported. We performed rpoB gene sequencing and multi-locus sequence typing (MLST) on M. abscessus complex strains isolated from thirty-six patients in 2006-2012 (eighteen known CF patients). Twenty-eight strains (78%) were M. abscessus subsp. abscessus, eight M. abscessus subsp. massiliense, none were M. abscessus subsp. bolletii. Sequence type 1 (ST1) and ST26 (M. abscessus subsp. abscessus) were commonest. Seven M. abscessus subsp. abscessus STs (25%) were novel (two with novel alleles). Seven M. abscessus subsp. massiliense STs were previously reported (88%), including two ST23, the globally successful clone. In 2012, of 552 CF patients screened, eleven were infected with M. abscessus complex strains (2%). The most prevalent M. abscessus subsp. abscessus and M. abscessus subsp. massiliense strains in Ireland belong to widely-distributed STs, but there is evidence of high M. abscessus subsp. abscessus diversity. Copyright © 2015 European Cystic Fibrosis Society. Published by Elsevier B.V. All rights reserved.

  2. Molecular identification and osmotolerant profile of wine yeasts that ferment a high sugar grape must.

    Science.gov (United States)

    Tofalo, Rosanna; Chaves-López, Clemencia; Di Fabio, Federico; Schirone, Maria; Felis, Giovanna E; Torriani, Sandra; Paparella, Antonello; Suzzi, Giovanna

    2009-04-15

    The objective of this study was to examine the Saccharomyces and non-Saccharomyces yeast populations involved in a spontaneous fermentation of a traditional high sugar must (Vino cotto) produced in central Italy. Molecular identification of a total of 78 isolates was achieved by a combination of PCR-RFLP of the 5.8S ITS rRNA region and sequencing of the D1/D2 domain of the 26S rRNA gene. In addition, the isolates were differentiated by RAPD-PCR. Only a restricted number of osmotolerant yeast species, i.e. Candida apicola, Candida zemplinina and Zygosaccharomyces bailii, were found throughout all the fermentation process, while Saccharomyces cerevisiae prevailed after 15 days of fermentation. A physiological characterization of isolates was performed in relation to the resistance to osmotic stress and ethanol concentration. The osmotolerant features of C. apicola, C. zemplinina and Z. bailii were confirmed, while S. cerevisiae strains showed three patterns of growth in response to different glucose concentrations (2%, 20%, 40% and 60% w/v). The ability of some C. apicola and C. zemplinina strains to grow at 14% v/v ethanol is noteworthy. The finding that some yeast biotypes with higher multiple stress tolerance can persist in the entire winemaking process suggests possible future candidates as starter for Vino cotto production.

  3. Molecular Identification of Malassezia Species in Patients with Malassezia folliculitis in Sfax, Tunisia.

    Science.gov (United States)

    Cheikhrouhou, F; Guidara, R; Masmoudi, A; Trabelsi, H; Neji, S; Sellami, H; Makni, F; Ayadi, A

    2017-06-01

    Malassezia folliculitis is caused by the invasion of hair follicles by large numbers of Malassezia cells. Several Malassezia researches still use cultures, morphology and biochemical techniques. The aim of this study was to identify Malassezia species isolated from patients diagnosed with folliculitis, at the Parasitology and Mycology Laboratory of Sfax University Hospital, and to explore the genetic diversity of Malassezia by using PCR-RFLP and PCR-sequencing targeting the rDNA region of the Malassezia genome. Specimens were taken from 27 patients with Malassezia folliculitis. For the molecular identification, PCR amplification of the 26S rDNAD1/D2 region was carried out using the Malup and Maldown primers and three restriction enzymes (BanI, MspI and HeaII) for RFLP analysis. The nucleotide sequences of each isolate were compared to those in the NCBI GenBank by using BLASTIN algorithm. Three species of Malassezia yeasts were identified among the 31 Malassezia strains isolated: M. globosa (83.9%), M. sympodialis (12. 9%) and M. furfur (3.2%). The sequence analysis of M. globosa showed six genotypes. There is a high genotypic variability of M. globosa colonizing patients with folliculitis.

  4. Isolation and identification of bacteria causing arthritis in chickens

    Directory of Open Access Journals (Sweden)

    B. Y. Rasheed

    2011-01-01

    Full Text Available Sixty chickens 30-55 days old with arthritis symptoms, were collected from different broiler chickens farms, all samples were examined clinically, post mortem and bacterial isolation were done. The results revealed isolation of 26 (50.98% of Staphylococcus aureus, which were found highly sensitive to amoxycillin. The experimental infection of 10 chickens was carried out on 35 days old by intravenous inoculated with 107 cfu/ml of isolated Staphylococcus aureus. Arthritis occurred in 8 (80% chickens. Clinical signs and post mortem findings confined to depression, swollen joints, inability to stand.

  5. Molecular identification and characterization of Fusarium spp. associated with sorghum seeds.

    Science.gov (United States)

    Divakara, Shetty Thimmappa; Santosh, Parthasarathy; Aiyaz, Mohammed; Ramana, Mudili Venkata; Hariprasad, Puttaswamy; Nayaka, Siddaih Chandra; Niranjana, Siddapura Ramachandrappa

    2014-04-01

    Fusarium spp. are not only pathogenic to plants but are also known as toxin producers that negatively affect animal and human health. The identification of Fusarium spp. remains one of the most critical issues in fungal taxonomy. In this study, different strains of Fusarium spp. were isolated from sorghum seed samples and identified at the molecular level by tef-1α gene amplification. A multiplex polymerase chain reaction (mPCR) assay was developed to differentiate toxigenic and non-toxigenic Fusarium spp. by designing a primer for the Fum21 gene along with the Fum1 and Fum8 genes. A competitive direct enzyme-linked immunosorbent assay (CD-ELISA) was employed to assess the fumonisin-producing ability of Fusarium spp. Phylogenetic analyses were performed using partial sequences of tef-1α and inter-simple sequence repeat (ISSR) markers of different Fusarium spp. All 27 isolates of Fusarium spp. were positive for the tef-1α gene and revealed the presence of F. verticillioides, F. thapsina and F. cf. incarnatum-equiseti complex. The standardized mPCR assay distinguished toxigenic and non-toxigenic F. verticillioides. Further, mPCR fumonisin-positive F. verticillioides isolates were also positive by CD-ELISA. The tef-1α gene sequence was found to be useful in revealing intraspecific polymorphism to some extent. ISSR markers revealed a high level of polymorphism among different isolates of Fusarium spp., and the dendrogram of ISSR analyses grouped the 27 isolates into two major clusters. The present method provided rapid and reliable detection of fumonisin-producing Fusarium spp. The mPCR assay could be an alternative strategy to current conventional mycotoxin analytical techniques and a reliable tool for high-throughput monitoring of major mycotoxin-producing fungi during the processing steps of food and feed commodities. © 2013 Society of Chemical Industry.

  6. Isolation and identification of antimicrobial-producing lactic acid ...

    African Journals Online (AJOL)

    , Proteus species, Staphylococcus aureus, Salmonella species, Pseudomonas flourescence, P. aeruginosa, Serratia species and Pediococcus acidilactici. Of the 42 antimicrobial producing isolates characterized, 16, 12, 6 and 8 were identified ...

  7. Isolation, Characterization and Identification of Microalgae from the Red Sea

    KAUST Repository

    Luque Alanís, Patricio

    2013-05-01

    Eukaryotic microalgae from the Red Sea were isolated, characterized and identified with the purpose of building a culture collection that will serve future research activities in the area of industrial microbiology. Seven sampling locations were surveyed using an in-house designed isolation protocol. Microalgae enrichment was carried out in vitro using the streak plate method and fluorescence activated cell sorting approaches. Colonial and cellular microscopy, growth media preference assays, as well as temperature, pH and salinity tolerance tests were carried out to describe the isolates. DNA extraction, PCR amplification, template sequencing and in silico analyses were carried out to identify the isolates and arrange them in a proper phylogenetic description. In total, 129 isolates were obtained. From these, only 39 were selected for characterization given their increased ability of accumulating large amounts of biomass in solid and liquid media in relatively short periods of time. All of these have a green color, are unicellular, non-motile, photosynthetic organisms and have a cell size ranging from 5 to 8 µm. More than half of them showed growth preference in Walne media, followed by F/2, MN and BG-11 SW. Maximum temperature tolerance of all organisms was around 38 ºC, while optimum growth was observed close to 25 ºC. pH preference was diverse and three groups were identified: acidic (6), intermediate (8 - 9) and alkaline (> 10) growing isolates. Salinity tests showed an overall growth preference at 25 PSU, approximately 10 units lower than that found at the sampling stations. Most isolates showed diminished growth at high salinity and high pH, except for OS3S1b which grew well in both cases, and could be an interesting strain to study further. Twenty four isolates were related to Ulvophyceae sp. MBIC10591 by BLAST approaches with a maximum identity of 96 - 97%. A maximum likelihood phylogenetic tree was created for these isolates, relative to the BLAST hits

  8. Isolation and identification of an ester from a crude oil

    Science.gov (United States)

    Phillips, H.F.; Breger, I.A.

    1958-01-01

    A dioctylphthalate has been isolated from a crude oil by means of adsorption column chromatography. The ester was identified by means of elemental analysis, refractive index, and its infra-red absorption spectrum. Saponification of the isolate and examination of the resultant alcohol by means of infrared absorption spectra led to the conclusion that the ester is a branched chain dioctylphthalate. This is the first reported occurrence of an ester in crude petroleum. ?? 1958.

  9. Isolation and identification of indigenous lactic acid bacteria from North Sumatra river buffalo milk

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    Heni Rizqiati

    2015-06-01

    Full Text Available Buffalo milk is a source of various lactic acid bacteria (LAB which is potential as culture starter as well as the probiotic. This study was conducted to isolate and identify LAB from indigenous North Sumatra river buffalo milk. Lactic acid bacteria was isolated and grown in medium De Man Rogosa Sharpe Agar (MRSA. The isolation was conducted to obtain pure isolate. The identification of LAB was studied in terms of morphology, physiology, biochemistry and survival on low pH. Morphology tests were conducted by Gram staining and cell forming; physiology tests were conducted for growing viability at pH 4.5 and temperature at 45oC; whereas biochemistry tests were conducted for CO2, dextran and NH3 productions. Determination of LAB species was conducted using Analytical Profile Index (API test CHL 50. Results of identification showed that 41 isolates were identified as LAB with Gram-positive, catalase-negative, rod and round shaped characteristics. Resistance test done to low pH (pH 2 for the lactic acid bacteria showed decrease of bacteria viability up to1.24±0.68 log cfu/ml. The resistant isolates at low pH were L12, L16, L17, L19, L20, M10, P8, S3, S19 and S20. Identification with API test CHL 50 for 10 isolates showed that four isolates were identified as Lactobacillus plantarum, L. brevis, L. pentosus and Lactococuslactis.

  10. Identification of metal-tolerant organisms isolated from the ...

    African Journals Online (AJOL)

    2011-01-28

    Jan 28, 2011 ... industrial effluents, agricultural runoff, leaking sewers, on-site sanitation at ... in the river water. Ten litres of river water was collected from. Site C (Fig. 1) in a 10 ℓ plastic container and transported at 4°C. Metal concentrations in river water ...... identification and in situ detection of individual microbial cells.

  11. Morphological Identification of Rumen Microbial Isolate and Rumen ...

    African Journals Online (AJOL)

    This study investigated the rumen fermentation parameters and rumen microbial identification of West African dwarf (WAD) sheep supplemented with forage – based multinutrient blocks (MNB). Twenty-five male WAD sheep, 9 – 12 months of age were used. The animals were randomly assigned to five formulated ...

  12. Method for isolation and molecular characterization of extracellular microvesicles released from brain endothelial cells

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    Haqqani Arsalan S

    2013-01-01

    Full Text Available Abstract Background In addition to possessing intracellular vesicles, eukaryotic cells also produce extracellular microvesicles, ranging from 50 to 1000 nm in diameter that are released or shed into the microenvironment under physiological and pathological conditions. These membranous extracellular organelles include both exosomes (originating from internal vesicles of endosomes and ectosomes (originating from direct budding/shedding of plasma membranes. Extracellular microvesicles contain cell-specific collections of proteins, glycoproteins, lipids, nucleic acids and other molecules. These vesicles play important roles in intercellular communication by acting as carrier for essential cell-specific information to target cells. Endothelial cells in the brain form the blood–brain barrier, a specialized interface between the blood and the brain that tightly controls traffic of nutrients and macromolecules between two compartments and interacts closely with other cells forming the neurovascular unit. Therefore, brain endothelial cell extracellular microvesicles could potentially play important roles in ‘externalizing’ brain-specific biomarkers into the blood stream during pathological conditions, in transcytosis of blood-borne molecules into the brain, and in cell-cell communication within the neurovascular unit. Methods To study cell-specific molecular make-up and functions of brain endothelial cell exosomes, methods for isolation of extracellular microvesicles using mass spectrometry-compatible protocols and the characterization of their signature profiles using mass spectrometry -based proteomics were developed. Results A total of 1179 proteins were identified in the isolated extracellular microvesicles from brain endothelial cells. The microvesicles were validated by identification of almost 60 known markers, including Alix, TSG101 and the tetraspanin proteins CD81 and CD9. The surface proteins on isolated microvesicles could potentially

  13. Molecular characterization of pneumococcal isolates from pets and laboratory animals.

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    Mark van der Linden

    Full Text Available BACKGROUND: Between 1986 and 2008 Streptococcus pneumoniae was isolated from 41 pets/zoo animals (guinea pigs (n = 17, cats (n = 12, horses (n = 4, dogs (n = 3, dolphins (n = 2, rat (n = 2, gorilla (n = 1 treated in medical veterinary laboratories and zoos, and 44 laboratory animals (mastomys (multimammate mice; n = 32, mice (n = 6, rats (n = 4, guinea pigs (n = 2 during routine health monitoring in an animal facility. S. pneumoniae was isolated from nose, lung and respiratory tract, eye, ear and other sites. METHODOLOGY/PRINCIPAL FINDINGS: Carriage of the same isolate of S. pneumoniae over a period of up to 22 weeks was shown for four mastomys. Forty-one animals showed disease symptoms. Pneumococcal isolates were characterized by optochin sensitivity, bile solubility, DNA hybridization, pneumolysin PCR, serotyping and multilocus sequence typing. Eighteen of the 32 mastomys isolates (56% were optochin resistant, all other isolates were optochin susceptible. All mastomys isolates were serotype 14, all guinea pig isolates serotype 19F, all horse isolates serotype 3. Rats had serotypes 14 or 19A, mice 33A or 33F. Dolphins had serotype 23F, the gorilla serotype 14. Cats and dogs had many different serotypes. Four isolates were resistant to macrolides, three isolates also to clindamycin and tetracycline. Mastomys isolates were sequence type (ST 15 (serotype 14, an ST/serotype combination commonly found in human isolates. Cats, dogs, pet rats, gorilla and dolphins showed various human ST/serotype combinations. Lab rats and lab mice showed single locus variants (SLV of human STs, in human ST/serotype combinations. All guinea pig isolates showed the same completely new combination of known alleles. The horse isolates showed an unknown allele combination and three new alleles. CONCLUSIONS/SIGNIFICANCE: The isolates found in mastomys, mice, rats, cats, dogs, gorilla and dolphins are most likely identical to human pneumococcal isolates. Isolates from

  14. IDENTIFICATION OF THE ISOLATED COMPOUNDS FROM Zingiber amaricans BL. RHIZOME

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    Sugeng Riyanto

    2010-06-01

    Full Text Available Five extracts were obtained from extraction of rhizomes of Zingiber amaricans. Hexane, dichloromethane and methanol extracts were obtained by maceration, while dichloromethane and acetone extracts the resulted of soxhlet extraction. By column chromatography technique 2,6,9-humulantrien-9-one (zerumbone was isolated as the major constituent of the hexane, dichloromethane and methanol extracts. The minor constituents were phytosterol mixtures isolated from hexane and dichloromethane extracts. The mixtures consisted cholesterol, campesterol, stigmasterol and b-sitosterol. The structure elucidations of zerumbone was confirmed by spectroscopic method, whereas the phytosterol mixtures was identified by gas chromatography-mass.   Keywords: zerumbone, phytosterol, zingiber amaricans, spectroscopy

  15. Use of PCR-RFLP and PCR-HWP1 for identification of Candida species isolated from cystic fibrosis patients

    Directory of Open Access Journals (Sweden)

    Peyman Solimani

    2014-08-01

    Full Text Available Background: Due to the predisposing conditions in patients with cystic fibrosis (CF caused by defective mucociliary clearance facilitating colonization and invasion with Candida species has dramatically increased. Traditional methods for identifying problems are imminent and time-consuming. Therefore, molecular techniques utilizing amplification of target DNA provide quick and precise methods for the diagnosis and identification of Candida species. Therefore, the aim of the current study was to identify the most medically common isolated Candida species from the air way of CF patients by PCR-RFLP and amplification of HWP1 gene. Materials and Methods: A total of 42 CF patients presenting symptoms who referred to pediatric respiratory diseases research center were screened for the presence of Candida spp. The isolates initially were phenotypically identified and confirmed by molecular approaches based on restriction fragment length polymorphism ( PCR -RFLP for the discrimination of C. albicans of non-albicans and the amplification of HWP1 gene for the discrimination of C. albicans from C. dubliniensis and C. africana was conducted. Results: The results show that C. albicans was the most frequently isolated species (83.8% followed by non-albicans included C. parapsilosis (7.1%, C. glabrata (3.2%, and C. tropicalis (3.2%. The restriction patterns of each Candida species were perfectly specific. Since MspI could not discriminate between the three morphological related species, C. albicans, C. dubliniensis and C. africana, we used PCR amplification of HWP1 gene, which (7.1% species from C. albicans identified as C. dubliniensis, however C. africana strains were not found. Conclusion: The present study found that C. albicans as predominant species wereisolated from the CF patients. It could be concluded that molecular diagnostic methods are reliable and would be useful for the identification of medically important Candida species in clinical samples

  16. Surveillance and Molecular Identification of Acanthamoeba and Naegleria Species in Two Swimming Pools in Alexandria University, Egypt

    Science.gov (United States)

    AL-HERRAWY, Ahmad Z.; KHALIL, Mahmoud I.; EL-SHERIF, Soheir S.; OMAR, Fatima A. E.; LOTFY, Wael M.

    2017-01-01

    Background: Swimming in contaminated water was reported to be associated with Acanthamoeba and N. fowleri human infections. The present study was carried out with the aim of isolation and identification of the different species of Acanthamoeba and Naegleria from two swimming pools in Alexandria University. Methods: Samples were collected from the swimming pools of Alexandria University Stadium and Faculty of Agriculture-Alexandria University during the period from May 2012 to April 2013. Results: Free-living amoebae were prevalent in the collected samples. Molecular characterization confirmed the identity of ten Acanthamoeba isolates and seven Naegleria isolates. Acanthamoeba T3, T4, T5, T11 and T15 genotypes were identified. Acanthamoeba T4 was the most prevalent genotype. Conclusion: The relatively high prevalence of Acanthamoeba, especially genotype T4, indicates the presence of a health hazard to swimmers particularly those wearing contact lenses. Naegleria fowleri was not found during the present study. PMID:28761479

  17. Surveillance and Molecular Identification of Acanthamoeba and Naegleria Species in Two Swimming Pools in Alexandria University, Egypt

    Directory of Open Access Journals (Sweden)

    Ahmad Z. AL-HERRAWY

    2017-06-01

    Full Text Available Background: Swimming in contaminated water was reported to be associated with Acanthamoeba and N. fowleri human infections. The present study was carried out with the aim of isolation and identification of the different species of Acanthamoeba and Naegleria from two swimming pools in Alexandria University. Methods: Samples were collected from the swimming pools of Alexandria University Stadium and Faculty of Agriculture-Alexandria University during the period from May 2012 to April 2013.Results: Free-living amoebae were prevalent in the collected samples. Molecular characterization confirmed the identity of ten Acanthamoeba isolates and seven Naegleria isolates. Acanthamoeba T3, T4, T5, T11 and T15 genotypes were identified. Acanthamoeba T4 was the most prevalent genotype.Conclusion: The relatively high prevalence of Acanthamoeba, especially genotype T4, indicates the presence of a health hazard to swimmers particularly those wearing contact lenses. Naegleria fowleri was not found during the present study. 

  18. Isolation and Identification of Fungi Associated with the Spoilage of ...

    African Journals Online (AJOL)

    This study was carried out in Sokoto Metropolis to isolate and identify fungi associated with the deterioration of sweet orange fruits. A total of one hundred samples of fresh sweet Oranges (Citrus sinensis L) were used. First, a total of seventy samples were obtained from the three selected marketing centres in Sokoto ...

  19. Isolation, Identification and Determination of Six Nucleosides and ...

    African Journals Online (AJOL)

    Methods: The compounds in bamboo shoots were isolated and identified by ultraviolet (UV) spectroscopy, mass spectrometry (MS), and nuclear magnetic resonance (NMR). Quantitative analysis was performed by reversed-phase high performance liquid chromatography (RP-HPLC) using a C18 column and a mixture ...

  20. Isolation and identification of two marine-derived Streptomyces from ...

    African Journals Online (AJOL)

    Administrator

    2011-09-26

    Sep 26, 2011 ... mycelia were determined with the ISCC-NBS centroid color charts. (US National Bureau of Standard 1976). The studies on physiological characteristics of the two marine actinomycete isolates were carried out following the methods recommended by the International Streptomyces Projects (ISP), Shirling et ...

  1. Isolation and Identification of Adenovirus Recovered from the Stool ...

    African Journals Online (AJOL)

    In order to establish the role of adenovirus in gastroenteritis in Nigerian children, stool samples were collected from 138 young children with gastroenteritis and 29 other age-matched controls. The samples were inoculated into 6 different tissue culture cell lines and isolates with characteristic CPE were subjected to CFT ...

  2. Isolation and identification of bioactive compounds from kernel seed ...

    African Journals Online (AJOL)

    The ethanol extract and ethyl acetate fraction of Mangifera indica kernel seed cake inhibited the growth of Staphylococcus aureus and Pseudomonas aeruginosa. The bioactive compounds were isolated and identified by NMR, UV and mass spectrometry as methyl gallate, gallic acid and penta-O-galloylglucose. The

  3. Identification of six potato virus Y isolates from Saudi Arabia

    African Journals Online (AJOL)

    Sherif

    2012-05-22

    May 22, 2012 ... sequences were aligned together, narrowed to six (one PVY-N and five PVY-O isolates) and then aligned with all published ... The phylogenetic analysis of the cp gene nucleotide sequence revealed a cluster of PVY-saudi-N .... clustered sequences combined with the PYV coat protein, multiple alignments ...

  4. Isolation, purification and identification of bacteria from the shoes ...

    African Journals Online (AJOL)

    13 strains of bacteria were isolated from 12 shoes that were worn by children aged 6 to 12 for more than half a year. Through morphological observation, physiological and biochemical measurements, as well as 16SrRNA sequence analysis, the bacteria were identified as follows: Bacillus licheniformis, Bacillus subtilis (5 ...

  5. Identification of potential local isolated for biosurfactant production

    Science.gov (United States)

    Shafiei, Zahra; Yusoff, Wan Mohtar Wan; Hamid, Aidil Abdul; Moazami, Nasrin; Hamzah, Ainon; Fooladi, Taybeh

    2013-11-01

    Biosurfactant are amphiphilic molecule that have received increasing attention in recent years because of their role in the growth of microorganisms on water-insoluble hydrophobic materials such as hydrocarbons as well as their commercial potential in the cosmetics, food, oil recovery and agricultural industries. In this study a potential biosurfactant producing strain was isolated from several soil samples of Terengganu oil refinery, Malaysia and selected during preliminary screening using hemolytic activity, oil spreading and drop collapsed technique. Isolates with at least more than one positive response to these three methods were subjected to complementary screening by measuring surface tension reduction as well as emulsification capacity. The biosurfactant produced by isolated 5M was able to reduced surface tension of culture medium from 60 mN/m to30mN/m. The biochemical and morphological characterization, 16SrRNA gene sequencing showed that the isolated 5M belongs to bacillus groups. The maximum production of biosurfactant by Bacillus 5M was observed after 48 h of incubation.

  6. Isolation and identification of the microorganisms most prevalent in ...

    African Journals Online (AJOL)

    Infections of the external eye account for a significant percentage of ocular inflammations, some of which lead to visual losses as result of corneal involvement. This study purely isolated and identified the microorganisms most prevalent in external eye infections in Owerri urban (as seen Mercy Eye clinic). With the aid of ...

  7. Isolation and identification of microsatellite repeat motifs from the ...

    African Journals Online (AJOL)

    Jane

    2011-08-10

    Aug 10, 2011 ... isolated using streptavidin-biotin enrichment method. In total, 378 microsatellites were identified and ... researchers realized the benefits of microsatellites. Numerous reports detail the processes of ..... microsatellites for striped bass from repeat-enriched libraries. Conserv. Genet., 7: 971-982. Rivera MA ...

  8. Isolation and Identification of a Brevibacterium linens strain ...

    African Journals Online (AJOL)

    nitrophenol (PNP). Subsequent subcultures in agar, nutrient agar plates and agar slants by streaking led to isolation of pure colonies. The pure culture could degrade up to 300 mg/L PNP in presence of yeast extract. It was Gram positive rods, ...

  9. Isolation and identification of fungal species from dried date palm ...

    African Journals Online (AJOL)

    A total of 360 dried date palm (Phoenix dactylifera) fruits were collected from hawkers, shops and market places within Maiduguri metropolis for the detection of the presence of fungal species. Investigation was based on cultural, microscopically and biochemical tests. Of the 327 (90.83%) fungal isolates recovered on ...

  10. Isolation, identification and antimicrobial suscep- tibility profiles of ...

    African Journals Online (AJOL)

    The samples were examined for the presence of Salmonella following standard techniques and procedures outlined by the International Organization for Standardization. Kibry-Bauer disk diffusion test was used for the antimicrobial susceptibility testing. Salmonella was isolated from 28/266 (10.5%) of the total samples. Out.

  11. Molecular identification and histopathological study of natural Streptococcus agalactiae infection in hybrid tilapia (Oreochromis niloticus

    Directory of Open Access Journals (Sweden)

    Laith Abdul Razzak

    2017-01-01

    Full Text Available Aim: The main objective of this study was to emphasize on histopathological examinations and molecular identification of Streptococcus agalactiae isolated from natural infections in hybrid tilapia (Oreochromis niloticus in Temerloh Pahang, Malaysia, as well as to determine the susceptibility of the pathogen strains to various currently available antimicrobial agents. Materials and Methods: The diseased fishes were observed for variable clinical signs including fin hemorrhages, alterations in behavior associated with erratic swimming, exophthalmia, and mortality. Tissue samples from the eyes, brain, kidney, liver, and spleen were taken for bacterial isolation. Identification of S. agalactiae was screened by biochemical methods and confirmed by VITEK 2 and 16S rRNA gene sequencing. The antibiogram profiling of the isolate was tested against 18 standard antibiotics included nitrofurantoin, flumequine, florfenicol, amoxylin, doxycycline, oleandomycin, tetracycline, ampicillin, lincomycin, colistin sulfate, oxolinic acid, novobiocin, spiramycin, erythromycin, fosfomycin, neomycin, gentamycin, and polymyxin B. The histopathological analysis of eyes, brain, liver, kidney, and spleen was observed for abnormalities related to S. agalactiae infection. Results: The suspected colonies of S. agalactiae identified by biochemical methods was observed as Gram-positive chained cocci, β-hemolytic, and non-motile. The isolate was confirmed as S. agalactiae by VITEK 2 (99% similarity, reconfirmed by 16S rRNA gene sequencing (99% similarity and deposited in GenBank with accession no. KT869025. The isolate was observed to be resistance to neomycin and gentamicin. The most consistent gross findings were marked hemorrhages, erosions of caudal fin, and exophthalmos. Microscopic examination confirmed the presence of marked congestion and infiltration of inflammatory cell in the eye, brain, kidney, liver, and spleen. Eye samples showed damage of the lens capsule

  12. [Isolation and identification of proteins with anti-tumor and fibrinolysogen kinase activities from Eisenia foetida].

    Science.gov (United States)

    Zhao, Rui; Ji, Jian-Guo; Tong, Yuan-Peng; Chen, Qian; Pu, Hai; Ru, Bing-Gen

    2002-09-01

    Proteins from Eisenia foetida possess many biological activities. A group of proteins precipitated by ethanol were isolated and purified by Sephadex G-75 and HiPrep 16/60 DEAE columns, then identified by one- or two- dimensional electrophoresis and mass spectrometry. 2D gel experiments displayed that the pI of proteins from Eisenia foetida were mainly from 3.0 to 4.0. Anti-tumor and kinase activities were determined by in vitro experiments. The enthanol fraction D2(8) showed both of the activities. These ethanol-precipitated proteins were identified further by native polyacrylamide electrophoresis, the protein spots were cut off from gels and digested by trypsin, the peptide mass fingerprints (PMFs) were determined by mass spectrometry. PMF, molecular weight, amino acid composition and N-terminus of 6 proteins were characterized, and band 9 was identified as D2(8). The results suggested that there exist proteins in Eisenia foetida possessed both anti-tumor and fibrinolysogen kinase activities. These methods can be used for identification of the natural bioactive proteins.

  13. COMPARATIVE ANALYSIS OF METHODS FOR IDENTIFICATION OF NONTUBERCULOUS MYCOBACTERIA ISOLATED FROM CLINICAL MATERIAL

    OpenAIRE

    A. V. Lyamin; D. D. Ismatullin; A. V. Zhestkov; A. M. Kovalyov; L. A. Baryshnikova; S. S. Nenjajkin

    2017-01-01

    Recently there has been a significant increase in the incidence of mycobacteriosis, which is due to an increase in the proportion of immunosuppressed patients, the presence of these various comorbid conditions, as well as the improvement of diagnostic methods. Selecting the most accurate method of identification is extremely important in determining treatment strategy of patients. The aim of the study was to conduct a comparative analysis of modern methods of identification NTMB isolated from...

  14. Isolation and identification of differentially expressed genes between ...

    African Journals Online (AJOL)

    Plants have evolved sophisticated molecular defense mechanisms in order to survive disease conditions. So far, a number of pathogen resistance (R) genes have been reported in plants. These R genes are thought to be involved in activating the signals that lead to disease resistance. The structural specificity of R genes ...

  15. Species distribution of clinical Acinetobacter isolates revealed by different identification techniques.

    Directory of Open Access Journals (Sweden)

    Jianfeng Wang

    Full Text Available A total of 2582 non-duplicate clinical Acinetobacter spp. isolates were collected to evaluate the performance of four identification methods because it is important to identify Acinetobacter spp. accurately and survey the species distribution to determine the appropriate antimicrobial treatment. Phenotyping (VITEK 2 and VITEK MS and genotyping (16S rRNA and rpoB gene sequencing methods were applied for species identification, and antimicrobial susceptibility test of imipenem and meropenem was performed with a disk diffusion assay. Generally, the phenotypic identification results were quite different from the genotyping results, and their discrimination ability was unsatisfactory, whereas 16S rRNA and rpoB gene sequencing showed consistent typing results, with different resolution. Additionally, A. pittii, A. calcoaceticus and A. nosocomialis, which were phylogenetically close to A. baumannii, accounted for 85.5% of the non-A. baumannii isolates. One group, which could not be clustered with any reference strains, consisted of 11 isolates and constituted a novel Acinetobacter species that was entitled genomic species 33YU. None of the non-A. baumannii isolates harbored a blaOXA-51-like gene, and this gene was disrupted by ISAba19 in only one isolate; it continues to be appropriate as a genetic marker for A. baumannii identification. The resistance rate of non-A. baumannii isolates to imipenem and/or meropenem was only 2.6%, which was significantly lower than that of A. baumannii. Overall, rpoB gene sequencing was the most accurate identification method for Acinetobacter species. Except for A. baumannii, the most frequently isolated species from the nosocomial setting were A. pittii, A. calcoaceticus and A. nosocomialis.

  16. Species Distribution of Clinical Acinetobacter Isolates Revealed by Different Identification Techniques

    Science.gov (United States)

    Feng, Ye; Fu, Ying; Jiang, Yan; Wang, Haiping; Yu, Yunsong

    2014-01-01

    A total of 2582 non-duplicate clinical Acinetobacter spp. isolates were collected to evaluate the performance of four identification methods because it is important to identify Acinetobacter spp. accurately and survey the species distribution to determine the appropriate antimicrobial treatment. Phenotyping (VITEK 2 and VITEK MS) and genotyping (16S rRNA and rpoB gene sequencing) methods were applied for species identification, and antimicrobial susceptibility test of imipenem and meropenem was performed with a disk diffusion assay. Generally, the phenotypic identification results were quite different from the genotyping results, and their discrimination ability was unsatisfactory, whereas 16S rRNA and rpoB gene sequencing showed consistent typing results, with different resolution. Additionally, A. pittii, A. calcoaceticus and A. nosocomialis, which were phylogenetically close to A. baumannii, accounted for 85.5% of the non-A. baumannii isolates. One group, which could not be clustered with any reference strains, consisted of 11 isolates and constituted a novel Acinetobacter species that was entitled genomic species 33YU. None of the non-A. baumannii isolates harbored a bla OXA-51-like gene, and this gene was disrupted by ISAba19 in only one isolate; it continues to be appropriate as a genetic marker for A. baumannii identification. The resistance rate of non-A. baumannii isolates to imipenem and/or meropenem was only 2.6%, which was significantly lower than that of A. baumannii. Overall, rpoB gene sequencing was the most accurate identification method for Acinetobacter species. Except for A. baumannii, the most frequently isolated species from the nosocomial setting were A. pittii, A. calcoaceticus and A. nosocomialis. PMID:25120020

  17. MALDI-TOF mass spectrometry for rapid identification of clinical fungal isolates based on ribosomal protein biomarkers.

    Science.gov (United States)

    Panda, Ashutosh; Ghosh, Anup K; Mirdha, Bijay R; Xess, Immaculata; Paul, Saikat; Samantaray, Jyotish C; Srinivasan, Alagiri; Khalil, Shehla; Rastogi, Neha; Dabas, Yubhisha

    2015-02-01

    This study aimed to evaluate the identification of clinical fungal isolates (yeast and molds) by protein profiling using Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF/MS). A total of 125 clinical fungal culture isolates (yeast and filamentous fungi) were collected. The test set included 88 yeast isolates (Candida albicans, Candida glabrata, Candida guilliermondii, Candida kefyr, Candida krusei, Candida parapsilosis, Candida rugosa, Candida tropicalis and Cryptococcus neoformans) and 37 isolates of molds (Alternaria spp., Aspergillus flavus, Aspergillus fumigatus, Aspergillus niger, Cunninghamella spp., Histoplasma capsulatum, Microsporum gypseum, Microsporum nanum, Rhizomucor spp. and Trichophyton spp.). The correlation between MALDI TOF MS and conventional identification for all these 125 fungal isolates included in the study was 87.2% at the species level and 90.4% at the genus level. MALDI TOF MS results revealed that the correlation in yeast (n=88) identification was 100% both at the genus and species levels whereas, the correlation in mold (n=37) identification was more heterogeneous i.e. 10.81% isolates had correct identification up to the genus level, 56.7% isolates had correct identification both at the genus and species levels, whereas 32.42% isolates were deemed Not Reliable Identification (NRI). But, with the modification in sample preparation protocol for molds, there was a significant improvement in identification. 86.4% isolates had correct identification till the genus and species levels whereas, only 2.7% isolates had Not Reliable Identification. In conclusion, this study demonstrates that MALDI-TOF MS could be a possible alternative to conventional techniques both for the identification and differentiation of clinical fungal isolates. However, the main limitation of this technique is that MS identification could be more precise only if the reference spectrum of the fungal species is available in the

  18. Molecular identification of Lactobacillus spp. associated with puba, a Brazilian fermented cassava food

    Science.gov (United States)

    Crispim, S.M.; Nascimento, A.M.A.; Costa, P.S.; Moreira, J.L.S.; Nunes, A.C.; Nicoli, J.R.; Lima, F.L.; Mota, V.T.; Nardi, R.M.D.

    2013-01-01

    Puba or carimã is a Brazilian staple food obtained by spontaneous submerged fermentation of cassava roots. A total of 116 lactobacilli and three cocci isolates from 20 commercial puba samples were recovered on de Man, Rogosa and Sharpe agar (MRS); they were characterized for their antagonistic activity against foodborne pathogens and identified taxonomically by classical and molecular methods. In all samples, lactic acid bacteria were recovered as the dominant microbiota (7.86 ± 0.41 log10 CFU/g). 16S–23S rRNA ARDRA pattern assigned 116 isolates to the Lactobacillus genus, represented by the species Lactobacillus fermentum (59 isolates), Lactobacillus delbrueckii (18 isolates), Lactobacillus casei (9 isolates), Lactobacillus reuteri (6 isolates), Lactobacillus brevis (3 isolates), Lactobacillus gasseri (2 isolates), Lactobacillus nagelii (1 isolate), and Lactobacillus plantarum group (18 isolates). recA gene-multiplex PCR analysis revealed that L. plantarum group isolates belonged to Lactobacillus plantarum (15 isolates) and Lactobacillus paraplantarum (3 isolates). Genomic diversity was investigated by molecular typing with rep (repetitive sequence)-based PCR using the primer ERIC2 (enterobacterial repetitive intergenic consensus). The Lactobacillus isolates exhibited genetic heterogeneity and species-specific fingerprint patterns. All the isolates showed antagonistic activity against the foodborne pathogenic bacteria tested. This antibacterial effect was attributed to acid production, except in the cases of three isolates that apparently produced bacteriocin-like inhibitory substances. This study provides the first insight into the genetic diversity of Lactobacillus spp. of puba. PMID:24159278

  19. Molecular identification of Lactobacillus spp. associated with puba, a Brazilian fermented cassava food

    Directory of Open Access Journals (Sweden)

    S.M. Crispim

    2013-01-01

    Full Text Available Puba or carimã is a Brazilian staple food obtained by spontaneous submerged fermentation of cassava roots. A total of 116 lactobacilli and three cocci isolates from 20 commercial puba samples were recovered on de Man, Rogosa and Sharpe agar (MRS; they were characterized for their antagonistic activity against foodborne pathogens and identified taxonomically by classical and molecular methods. In all samples, lactic acid bacteria were recovered as the dominant microbiota (7.86 ± 0.41 log10 CFU/g. 16S-23S rRNA ARDRA pattern assigned 116 isolates to the Lactobacillus genus, represented by the species Lactobacillus fermentum (59 isolates, Lactobacillus delbrueckii (18 isolates, Lactobacillus casei (9 isolates, Lactobacillus reuteri (6 isolates, Lactobacillus brevis (3 isolates, Lactobacillus gasseri (2 isolates, Lactobacillus nagelii (1 isolate, and Lactobacillus plantarum group (18 isolates. recA gene-multiplex PCR analysis revealed that L. plantarum group isolates belonged to Lactobacillus plantarum (15 isolates and Lactobacillus paraplantarum (3 isolates. Genomic diversity was investigated by molecular typing with rep (repetitive sequence-based PCR using the primer ERIC2 (enterobacterial repetitive intergenic consensus. The Lactobacillus isolates exhibited genetic heterogeneity and species-specific fingerprint patterns. All the isolates showed antagonistic activity against the foodborne pathogenic bacteria tested. This antibacterial effect was attributed to acid production, except in the cases of three isolates that apparently produced bacteriocin-like inhibitory substances. This study provides the first insight into the genetic diversity of Lactobacillus spp. of puba.

  20. Proteomic identification of gender molecular markers in Bothrops jararaca venom.

    Science.gov (United States)

    Zelanis, André; Menezes, Milene C; Kitano, Eduardo S; Liberato, Tarcísio; Tashima, Alexandre K; Pinto, Antonio F M; Sherman, Nicholas E; Ho, Paulo L; Fox, Jay W; Serrano, Solange M T

    2016-04-29

    Variation in the snake venom proteome is a well-documented phenomenon; however, sex-based variation in the venom proteome/peptidome is poorly understood. Bothrops jararaca shows significant sexual size dimorphism and here we report a comparative proteomic/peptidomic analysis of venoms from male and female specimens and correlate it with the evaluation of important venom features. We demonstrate that adult male and female venoms have distinct profiles of proteolytic activity upon fibrinogen and gelatin. These differences were clearly reflected in their different profiles of SDS-PAGE, two-dimensional electrophoresis and glycosylated proteins. Identification of differential protein bands and spots between male or female venoms revealed gender-specific molecular markers. However, the proteome comparison by in-solution trypsin digestion and label-free quantification analysis showed that the overall profiles of male and female venoms are similar at the polypeptide chain level but show striking variation regarding their attached carbohydrate moieties. The analysis of the peptidomes of male and female venoms revealed different contents of peptides, while the bradykinin potentiating peptides (BPPs) showed rather similar profiles. Furthermore we confirmed the ubiquitous presence of four BPPs that lack the C-terminal Q-I-P-P sequence only in the female venom as gender molecular markers. As a result of these studies we demonstrate that the sexual size dimorphism is associated with differences in the venom proteome/peptidome in B. jararaca species. Moreover, gender-based variations contributed by different glycosylation levels in toxins impact venom complexity. Bothrops jararaca is primarily a nocturnal and generalist snake species, however, it exhibits a notable ontogenetic shift in diet and in venom proteome upon neonate to adult transition. As is common in the Bothrops genus, B. jararaca shows significant sexual dimorphism in snout-vent length and weight, with females being

  1. Isolation and Identification of Burkholderia glumae from Symptomless Rice Seeds

    Directory of Open Access Journals (Sweden)

    Bo Zhu

    2008-06-01

    Full Text Available A survey on isolation and detection of the casual organism of bacterial grain rot of rice was conducted during 1997–2006. In 2006, six pathogenic bacterial strains were isolated from two symptomless seed samples of rice (Oryza sativa L. originally produced in Hainan Province and then planted in Zhejiang Province, China. They were identified as Burkholderia glumae which is the causal organism of bacterial grain rot of rice by physiological characteristics, colony morphology, pathogenicity test, Biolog, fatty acid methyl ester (FAME analysis and RAPD-PCR compared with the four standard reference strains. It is confirmed that there is the infection of B. glumae in so-called ‘health looking seeds’.

  2. Isolation, identification and antimicrobial activity of propolis-associated fungi.

    Science.gov (United States)

    de Souza, Giovanni Gontijo; Pfenning, Ludwig Heinrich; de Moura, Fabiana; Salgado, Mírian; Takahashi, Jacqueline Aparecida

    2013-01-01

    Propolis is a natural product widely known for its medicinal properties. In this work, fungi present on propolis samples were isolated, identified and tested for the production of antimicrobial metabolites. Twenty-two fungal isolates were obtained, some of which were identified as Alternaria alternata, Aspergillus flavus, Bipolaris hawaiiensis, Fusarium merismoides, Lasiodiplodia theobromae, Penicillium citrinum, Penicillium crustosum, Penicillium janthinellum, Penicillium purpurogenum, Pestalotiopsis palustris, Tetracoccosporium paxianum and Trichoderma koningii. These fungi were grown in liquid media to obtain crude extracts that were evaluated for their antibiotic activity against pathogenic bacteria, yeast and Cladosporium cladosporioides and A. flavus. The most active extract was obtained from L. theobromae (minimum inhibitory concentration = 64 μg/mL against Listeria monocitogenes). Some extracts showed to be more active than the positive control in the inhibition of Staphylococcus aureus and L. monocitogenes. Therefore, propolis is a promising source of fungi, which produces active agents against relevant food poisoning bacteria and crop-associated fungi.

  3. Comparison of a commercial biochemical kit and an oligonucleotide probe for identification of environmental isolates of Vibrio vulnificus

    DEFF Research Database (Denmark)

    Dalsgaard, A.; Dalsgaard, Inger; Høi, L.

    1996-01-01

    Methods for the identification and isolation of environmental isolates of Vibrio vulnificus were evaluated. Alkaline peptone water supplemented with polymyxin B and colistin-polymyxin B- cellobiose agar were employed for the isolation of suspected V. vulnificus from water, sediment and shellfish ...... hybridizing with the probe. The results indicated that, compared with colony hybridization, the API 20E assay was not adequate for the identification of environmental isolates of V. vulnificus....

  4. Identification of leptospiral isolates by bacterial restriction endonuclease analysis (Brenda

    Directory of Open Access Journals (Sweden)

    Venkatesha M

    2001-01-01

    Full Text Available DNA samples from 19 reference serovars belonging to 19 different serogroups of Leptospira interrogans and two serovars belonging to Leptospira biflexa were examined by bacterial restriction endonuclease analysis using EcoR I and Hae III enzymes. All the serovars gave unique restriction patterns that differed from each other. DNA from 10 local isolates digested with these enzymes produced patterns which on comparison with the standard patterns produced by reference strains could be identified to serovar level.

  5. Selection, isolation, and identification of fungi for bioherbicide production

    OpenAIRE

    Souza, Ang?lica Rossana Castro de; Baldoni, Daiana Bortoluzzi; Lima, Jessica; Porto, Vit?ria; Marcuz, Camila; Machado, Carolina; Ferraz, Rafael Camargo; Kuhn, Raquel C.; Jacques, Rodrigo J.S.; Guedes, Jerson V.C.; Mazutti, Marcio A.

    2016-01-01

    Abstract Production of a bioherbicide for biological control of weeds requires a series of steps, from selection of a suitable microbial strain to final formulation. Thus, this study aimed to select fungi for production of secondary metabolites with herbicidal activity using biological resources of the Brazilian Pampa biome. Phytopathogenic fungi were isolated from infected tissues of weeds in the Pampa biome. A liquid synthetic culture medium was used for production of metabolites. The phyto...

  6. [Isolation and identification of bacteria with ferro-oxidase activity].

    Science.gov (United States)

    Zheng, Hong; Zhang, Wensen; Zhang, Xiaorong; Wu, Xiaomei; Zhan, Xingdai; Deng, Jiacong

    2014-12-04

    We screened and isolated Ferro-oxidase producing bacteria, for adsorbing iron and manganese. The strains producing Ferro-oxidase were isolated from three samples of water. Ferro-oxidase producing strains were screened in shake flask culture, and identified according to morphological features, physiological and biochemical analysis as well as 16S rRNA gene sequence analysis. We isolated a bacterium S9. The strain was identified as Sphaerotilus natans. This strain had strongest adsorption on iron and manganese among the strains we identified, with 29.02 mg/g iron adsorption amount in water, and 66.77% adsorption rate for 4 hours' adsorption. When the adsorption time is 6 h, the adsorption amount of manganese was 34.49 mg/g, and the adsorption rate was 70.68%. The optimum temperature and pH value of Ferro-oxidase were 30 degrees C and 7.5, respectively. Mg2+, Na+, K+ could activate Ferro-oxidase, whereas Cu2+ had little impact. While Mn2+, Zn2+ could strongly inhibit Ferro-Oxidase, Pb2+, Ag+ had only modest inhibitory effect. Strain S9 had a high Ferro-oxidase activity, and has application potential in sewage treatment.

  7. Molecular characterization of Histoplasma capsulatum isolated from an outbreak in treasure hunters Histoplasma capsulatum in treasure hunters

    Directory of Open Access Journals (Sweden)

    Muñoz Bertha

    2010-09-01

    Full Text Available Abstract Background In Mexico, primary pulmonary histoplasmosis is the most relevant clinical form of the disease. The geographical distribution of specific strains of Histoplasma capsulatum circulating in Mexico has not been fully established. Outbreaks must be reported in order to have current, updated information on this disease, identifying new endemic areas, manner of exposure to the fungi, and molecular characterization of the causative agents. We report a recent outbreak of histoplasmosis in treasure hunters and the molecular characterization of two isolates obtained from these patients. Methods Six patients admitted to the National Institute of Respiratory Diseases (INER in Mexico City presented severe respiratory symptoms suggestive of histoplasmosis. They acquired the infection in the Veracruz (VZ endemic zone. Diagnosis was made by X-ray and Computed tomography (CT, liver function, immunological techniques, and culture. Identification of H. capsulatum isolates was confirmed by using Polymerase chain reaction (PCR was conducted with a probe from the M antigen, and the isolates were characterized by means of Random amplification of polymorphic DNA (RAPD-PCR employed the 1253 oligonucleotide and a mixture of oligonucleotides 1281 and 1283. These were compared to eight reference strain isolates from neighboring areas. Results X-ray and CT revealed disseminated micronodular images throughout lung parenchyma, as well as bilateral retrocaval, prevascular, subcarinal, and hilar adenopathies, hepatosplenomegaly, and altered liver function tests. Five of the six patients developed disseminated histoplasmosis. Two H. capsulatum strains were isolated. The same band profile was detected in both strains, indicating that both isolates corresponded to the sole H. capsulatum strain. Molecular characterization of the isolates was similar in 100% with the EH-53 Hidalgo human (HG strain (reference strain integrated into the LAm A clade described for

  8. Identification and diversity of Fusarium species isolated from tomato fruits

    Directory of Open Access Journals (Sweden)

    Murad Nur Baiti Abd

    2016-07-01

    Full Text Available Fruit rot of tomato is a serious disease caused by Fusarium species. Sampling was conducted throughout Selangor, Malaysia and fungal species identification was conducted based on morphological and gene encoding translation elongation factor 1-α (tef1-α sequence analysis. Five species of Fusarium were discovered namely F. oxysporum (including F. oxysporum f. sp. lycopersici, F. solani, F. equiseti, F. proliferatum and F. verticillioides. Our results provide additional information regarding the diversity of Fusarium species associated with fruit rot disease of tomato.

  9. Isolation, identification and subtyping of Campylobacter: where to from here?

    Science.gov (United States)

    On, Stephen L W

    2013-10-01

    Campylobacter species are widely regarded as the most frequent bacterial cause of gastroenteritis in humans worldwide. Their main transmission routes are via contaminated food and water. For interventions to be effective, methods for the detection, identification and epidemiological subtyping must be sensitive, accurate and rapid. As yet, methods are not perfect, although several significant advances have been made in these areas in recent years. This paper provides a brief review and commentary on the current state of the art in the hope that it will help provide context for others in selecting, improving or developing these vital tools for research and diagnoses. Copyright © 2013 Elsevier B.V. All rights reserved.

  10. Isolation and identification of lactic acid bacteria from koumiss in Eastern Inner Mongolia of China

    Science.gov (United States)

    Bai, Lijuan; Ji, Shujuan

    2017-01-01

    Koumiss is a traditional fermented dairy product and known as its unique physiological actions. Isolation and identification of LAB in it will yield valuable knowledge. In total, 55 LAB strains were isolated and identified of 12 koumiss samples collected in limited regions of Eastern Inner Mongolia. 16S rRNA sequence analysis results showed that were Lactobacillus helveticus, Lactobacillus casei, Lactobacillus plantarum, Lactobacillus delbrueckii, Enterococcus. durans, Leuconostoc lactis and Leuconostoc mesenteroides. It is benefit to further research on koumiss.

  11. Characterization and identification of newly isolated Acinetobacter baumannii strain serdang 1 for phenol removal

    Science.gov (United States)

    Yadzir, Z. H. M.; Shukor, M. Y.; Nazir, M. S.; Abdullah, M. A.

    2012-09-01

    A new indigenous bacterial strain from Malaysian soil contaminated with petroleum waste had been successfully isolated, characterized and identified for phenol removal. The gram negative bacteria showed 98% identity with Acinetobacter baumannii based on Biolog{trade mark, serif} Identification System and the determination of a partial 16S ribosomal RNA (rRNA) sequence. The isolate clustered with species belonging to Acinetobacter clade in a 16S rDNA-based neighbour-joining phylogenetic tree.

  12. Molecular identification of Anopheles (Cellia) sundaicus from the Andaman and Nicobar islands of India.

    Science.gov (United States)

    Alam, Mohammad Tauqeer; Das, Manoj K; Ansari, Musharraf A; Sharma, Yagya D

    2006-01-01

    Anopheles (Cellia) sundaicus (Rodenwaldt) is an important malaria vector in the Andaman and Nicobar islands of India where it breeds in freshwater as well as in brackish water. To establish the molecular identity of An. sundaicus on these islands we analyzed samples from four geographically isolated areas-Teressa, Nancowry, Car Nicobar and Katchal islands. PCR-amplification and nucleotide sequence analysis were performed for internal transcribed spacer 2 (ITS2) and domain-3 (D3) of 28S rRNA. The ITS2 region of An. sundaicus from all four islands was identical but different from An. sundaicus A of Vietnam and An. sundaicus s.s of Malaysia. Furthermore, freshwater and brackish water forms of An. sundaicus did not reveal any sequence variation. Similarly, the D3 sequences were identical among all An. sundaicus samples from the four islands. D3 sequences for a species of the Sundaicus Complex are reported here for the first time and thus could not be compared with other regional isolates of this species. In conclusion, probably only one member of the Sundaicus Complex exists on the Andaman and Nicobar islands, which breeds in freshwater as well as in brackish water and is different from the An. sundaicus A and Malaysian An. sundaicus s.s. The identification of a new sibling species of the Sundaicus Complex in these islands is significant from the viewpoint of vector control strategies.

  13. Morphological-anatomical characterization and molecular identification of Tomentella stuposa ectomycorrhizae and related anatomotypes.

    Science.gov (United States)

    Jakucs, Erzsébet; Kovács, G M; Agerer, R; Romsics, C; Eros-Honti, Z

    2005-06-01

    Species in the genus Tomentella (Thelephoraceae) belong to the most frequent and widespread ectomycorrhizal (EM) fungi found in temperate and boreal forests. Although several unidentified tomentelloid morphotypes have been presented as common members of EM communities in coniferous and broad-leaved forests, few tomentelloid EM have been identified and described in detail. In this study, ten tomentelloid EM isolates collected from Populus alba, Quercus cerris and Picea abies stands in Hungary and Germany are characterized and documented by morphological-anatomical methods using light microscopy. The investigated ectomycorrhizae belong to the same brown-black tomentelloid morphotype but form two different anatomotype groups (At I and At II). Molecular taxonomical identification was accomplished using phylogenetic analysis (neighbor joining method) of 49 Tomentella nrDNA-ITS nucleotide sequences including the 10 new and 39 GenBank sequences. The EM isolates clustered into two adjoining clades identical with the two anatomotypes. At II clustered with Tomentella stuposa while At I could not be identified to species. Based on the morphological similarity and the low genetic difference it must be a closely related taxon. A comparison of the recently known tomentelloid EM to T. stuposa is presented. Ecological questions involving abundance and host relationships are discussed.

  14. Isolation and identification of bacterial pathogen from mastitis milk in Central Java Indonesia

    Science.gov (United States)

    Harjanti, D. W.; Ciptaningtyas, R.; Wahyono, F.; Setiatin, ET

    2018-01-01

    Mastitis is a multi-etiologic disease of the mammary gland characterized mainly by reduction in milk production and milk quality due to intramammary infection by pathogenic bacteria. Nearly 83% of lactating dairy cows in Indonesia are infected with mastitis in various inflammation degrees. This study was conducted to isolate and identify the pathogen in milk collected from mastitis-infected dairy cows. The study was carried out in ten smallholder dairy farms in Central Java Indonesia based on animal examination, California mastitis test, isolation bacterial pathogens, Gram staining, Catalase and Coagulase test, and identification of bacteria species using Vitek. Bacteriological examination of milk samples revealed 15 isolates where Streptococcus was predominant species (73.3%) and the coagulase negative Staphylococcus species was identified at the least bacteria (26.7%). The Streptococcus bacteria found were Streptococcus uberis (2 isolates), Streptococcus sanguinis(6 isolates), Streptococcus dysgalactiaessp dysgalactiae(1 isolate) , Streptococcus mitis (1 isolate) and Streptococcus agalactiae (1 isolate). The Staphylococcus isolates comprising of Staphylococcus simulans (1 isolate) and Staphylococcus chromogens (3 isolates). Contamination of raw milkwith pathogenic bacteria can cause outbreaks of human disease (milk borne disease). Thus, proper milk processing method that couldinhibit the growth or kill these pathogenic bacteria is important to ensure the safety of milk and milk products.

  15. [Isolation and identification methods of enterobacteria group and its technological advancement].

    Science.gov (United States)

    Furuta, Itaru

    2007-08-01

    In the last half-century, isolation and identification methods of enterobacteria groups have markedly improved by technological advancement. Clinical microbiology tests have changed overtime from tube methods to commercial identification kits and automated identification. Tube methods are the original method for the identification of enterobacteria groups, that is, a basically essential method to recognize bacterial fermentation and biochemical principles. In this paper, traditional tube tests are discussed, such as the utilization of carbohydrates, indole, methyl red, and citrate and urease tests. Commercial identification kits and automated instruments by computer based analysis as current methods are also discussed, and those methods provide rapidity and accuracy. Nonculture techniques of nucleic acid typing methods using PCR analysis, and immunochemical methods using monoclonal antibodies can be further developed.

  16. Molecular characterization of two isolates of sweet potato leaf curl ...

    African Journals Online (AJOL)

    Comparison analysis showed that DNA-A sequence of JS1 isolate was closely related to that of sweet potato leaf curl virus (SPLCV) from United States with nucleotide sequence identity of 97.0% and DNA-A of Y338 showed highest sequence identity at 97.8% with an isolate of SPLCV from China. Phylogenetic analysis ...

  17. Molecular characterization of bacteria isolated from the Kingdom of ...

    African Journals Online (AJOL)

    ola

    2012-11-06

    Nov 6, 2012 ... Esherichia coli strains isolated from different habitats in relation to their antimicrobial activity against pathogenic fungi causing dermatological diseases. MATERIALS AND METHODS. Microbial strains, media and culture conditions. Bacterial isolates from soil samples were collected from different localities ...

  18. Molecular detection of Edwardsiella tarda with gyr B gene isolated ...

    African Journals Online (AJOL)

    The 16S rRNA gene was identical and exhibited 99% sequence similarity with the other known isolates of E. tarda available in the GenBank. This paper reports the isolation and detection of E. tarda with the gyrB gene in pirarucu, A. gigas, which was exhibited in an indoor private commercial aquarium in Seoul, South Korea ...

  19. Molecular characterization of Citrus tristeza virus isolates from ...

    African Journals Online (AJOL)

    The CTV CP gene of all isolates was amplified by reverse transcriptase polymerase chain reaction (RT–PCR) using CP gene-specific primers yielding 672 bp. The maximum disease incidence was found in sweet orange followed by mandarin and grapefruit. These isolates were then subjected to CPG/Hinf I restriction ...

  20. Molecular fingerprinting of Fusarium oxysporum f. sp. passiflorae isolates using AFLP markers

    Directory of Open Access Journals (Sweden)

    Aline dos Santos Silva

    2013-04-01

    Full Text Available Fusarium oxysporum f. sp. passiflorae W.L. Gordon (FOP is one of the most important fungal pathogens of passion fruits. Understanding molecular variation of isolates from different areas is of utmost importance. Molecular fingerprinting on 14 isolates of FOP were conducted using AFLP molecular markers (Amplified Fragment Length Polymorphism, and their genetic variability were estimated. Twenty-five AFLP primer combinations were selected for amplification of FOP isolates and one for Fusarium oxysporum f. sp. cubense W.C. Snyder & H.N. Hansen (FOC, resulting in 99% polymorphic fragments, with an average of 40 fragments per primer combination. Specific fingerprints could be generated for most of the isolates evaluated; we observed a high power of discrimination of the AFLP primer combinations, with the presence/absence of up to 26 specific fragments per isolate. Thus, specific fingerprinting was obtained for 10 of the 15 isolates analyzed. The values of the polymorphic information content, the index and the resolving power of the markers showed wide variation and reflected the high informative contents of the primers used in the characterization of the FOP isolates. The FOP isolates were divided into four groups, irrespective of their geographic origins, with the allocation of 5, 7, 1 and 1 FOP isolates into Groups II, III, IV and V, respectively. A wide genetic diversity was observed in FOP isolates, which should be taken into consideration when implementing strategies for the improvement of passion fruit in the search for cultivars with multiple resistance to different isolates.

  1. Molecular Identification of Bacteria Involved in Degradation of Crude ...

    African Journals Online (AJOL)

    Abstract. In this present study, bacteria were isolated from soil obtained from oil contaminated mechanic site in Enugu. Out of the 206 forming colonies, 16 colonies were selected based on observable morphological differences and screened for crude oil degraders. Out of the. 16 isolates, 11 isolates were assumed to be ...

  2. Identification of clinically relevant Corynebacterium strains by Api Coryne, MALDI-TOF-mass spectrometry and molecular approaches.

    Science.gov (United States)

    Alibi, S; Ferjani, A; Gaillot, O; Marzouk, M; Courcol, R; Boukadida, J

    2015-09-01

    We evaluated the Bruker Biotyper matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry (MS) for the identification of 97 Corynebacterium clinical in comparison to identification strains by Api Coryne and MALDI-TOF-MS using 16S rRNA gene and hypervariable region of rpoB genes sequencing as a reference method. C. striatum was the predominant species isolated followed by C. amycolatum. There was an agreement between Api Coryne strips and MALDI-TOF-MS identification in 88.65% of cases. MALDI-TOF-MS was unable to differentiate C. aurimucosum from C. minutissimum and C. minutissimum from C. singulare but reliably identify 92 of 97 (94.84%) strains. Two strains remained incompletely identified to the species level by MALDI-TOF-MS and molecular approaches. They belonged to Cellulomonas and Pseudoclavibacter genus. In conclusion, MALDI-TOF-MS is a rapid and reliable method for the identification of Corynebacterium species. However, some limits have been noted and have to be resolved by the application of molecular methods. Copyright © 2015. Published by Elsevier SAS.

  3. Identification of carcinogenic tannin isolated from Bracken fern (Pteridium aquilinum).

    Science.gov (United States)

    Wang, C Y; Chiu, C W; Pamukcu, A M; Bryan, G T

    1976-01-01

    We attempted to isolate a carcinogenic substance from bracken fern (Pteridium aquilinum), a naturally occurring toxicant responsible for the production of chronic enzootic hematuria and urinary bladder cancer of cattle and carcinogenic for various target organs of several species. Hot methanol extracts of bracken fern were solubilized in water and extracted with chloroform followed by a mixture of n-butanol-butanone (1:1). That fraction was dried and triturated with ether-methanol (4:1), n-butanol, and finally absolute ethanol. The insoluble residue was dissolved in 10% aqueous methanol and passed through Dowex 1 OH-, Dowex 50 H+, or Dowex 1 OH- and then Dowex 50 H+ ion exchange resins. A condensed tannin, isolated from one ot the fractions, was identical to that isolated from bracken fern by the caffeine procedure used for the separation of tannins from other plant constituents. Three systems were used for bioassay; induction of bladder carcinoma by implantation of cholesterol pellets containing bracken fern fractions into the bladder lumens of mice; acute toxicity by ip injection of brachen fern fraction into mice; and growth inhibition of Escherichia coli. The following fractions induced significantly greater incidences of bladder carcinoma than did cholesterol pellets only: tannin, Dowex 50 H+, residue, n-butanol, and methanol. Tiliroside, a component of bracken fern fractions into the bladder lumens of mice; acute genic acid, and quercetin were not carcinogenic. Tannin was the most toxic (mean lethal dose: 0.16 mg/g) and carcinogenic. None of the carcinogenic fractions inhibited growth of E. coli.

  4. Isolation and identification of an allelopathic substance from Hibiscus sabdariffa.

    Science.gov (United States)

    Suwitchayanon, Prapaipit; Pukclai, Piyatida; Ohno, Osamu; Suenaga, Kiyotake; Kato-Noguchi, Hisashi

    2015-05-01

    In this study, an allelopathic substance was isolated from an aqueous methanol extract of Hibiscus sabdariffa L. by column chromatography and reverse phase HPLC. The chemical structure of the substance was determined by 1H NMR spectroscopy and mass spectrometry as trimethyl allo-hydroxycitrate. Trimethyl allo-hydroxycitrate inhibited the growth of cress hypocotyls and roots at concentrations greater than 10 mM. The concentrations required for 50% growth inhibition of the hypocotyls and roots of cress were 20.3 and 14.4 mM, respectively. The inhibitory activity of trimethyl allo-hydroxycitrate suggests that the substance may act as an allelopathic substance of H. sabdariffa.

  5. Environmental issue identification for the Basalt Waste Isolation Project

    International Nuclear Information System (INIS)

    Carrell, D.J.; Jones, K.A.

    1980-04-01

    A preliminary evaluation of environmental issues is provided in this report. It contains summary of the thought process that was used during the area characterization studies for a geological repository for high-level radioactive wastes. Environmental issues are discussed separately for construction, operation, and long term isolation aspects of a repository in basalt. During construction the primary environmental concerns are public perception and water resources; intermediate concerns are air quality, ecosystems, physical resources, and cultural and social resources. During operation, the primary environmental issues concern the transport of radioactive materials and physical resources. Long term environmental issues envolve water resources and borehole plugging

  6. Pityriasis versicolor: isolation and identification of the main species

    OpenAIRE

    FRAMIL, Valéria Maria de Souza; MELHEM, Márcia S. C; SZESZS, Maria Walderez; CORNETA, Elaine Cristina; ZAITZ, Clarisse

    2010-01-01

    As espécies do gênero Malassezia isoladas foram: Malassezia sympodialis (16,66%), Malassezia furfur (12,50%), Malassezia globosa (11,45%) e Malassezia slooffiae (2,10%). A Malassezia sympodialis foi a espécie que predominou em nosso estudo. As espécies de Malassezia identificadas não mostraram correlação com as variantes clínicas e com a distribuição das lesões de pitiríase versicolor quanto às regiões do corpo.Species of the genus Malassezia isolated were: Malassezia sympodialis (16.66%), Ma...

  7. Isolation and identification of a novel radio-resistant strain

    International Nuclear Information System (INIS)

    Zhang Zhidong; Mao Jun; Wang Wei; Tang Qiyong; Shi Yuhu

    2008-01-01

    A novel radio-resistant strain named RL2 was studied polyphasically, which was isolated from the soils in the Gurban-Tunggut Desert, Xinjiang. The strain is Gam-positive, sphere-shaped and pink pigmented; The DNA (G+C) contents of RL2 is 71.62mo1%; The 16S rDNA genes of RL2 and D. radiodurans type strain DSM20539 shows a high level of similarity (97.2%). According to phenotypic characteristics and phylogenetic analysis, it can be suggested that the strain RL2 has been identified as Deinococcus. sp and it may be a novel species. (authors)

  8. Microbiological and molecular characterization of Staphylococcus hominis isolates from blood.

    Directory of Open Access Journals (Sweden)

    Soraya Mendoza-Olazarán

    Full Text Available BACKGROUND: Among Coagulase-Negative Staphylococci (CoNS, Staphylococcus hominis represents the third most common organism recoverable from the blood of immunocompromised patients. The aim of this study was to characterize biofilm formation, antibiotic resistance, define the SCCmec (Staphylococcal Chromosomal Cassette mec type, and genetic relatedness of clinical S. hominis isolates. METHODOLOGY: S. hominis blood isolates (n = 21 were screened for biofilm formation using crystal violet staining. Methicillin resistance was evaluated using the cefoxitin disk test and the mecA gene was detected by PCR. Antibiotic resistance was determined by the broth microdilution method. Genetic relatedness was determined by pulsed-field gel electrophoresis (PFGE and SCCmec typed by multiplex PCR using two different methodologies described for Staphylococcus aureus. RESULTS: Of the S. hominis isolates screened, 47.6% (10/21 were categorized as strong biofilm producers and 23.8% (5/21 as weak producers. Furthermore, 81% (17/21 of the isolates were methicillin resistant and mecA gene carriers. Resistance to ampicillin, erythromycin, and trimethoprim was observed in >70% of isolates screened. Each isolate showed a different PFGE macrorestriction pattern with similarity ranging between 0-95%. Among mecA-positive isolates, 14 (82% harbored a non-typeable SCCmec type: eight isolates were not positive for any ccr complex; four contained the mec complex A ccrAB1 and ccrC, one isolate contained mec complex A, ccrAB4 and ccrC, and one isolate contained the mec complex A, ccrAB1, ccrAB4, and ccrC. Two isolates harbored the association: mec complex A and ccrAB1. Only one strain was typeable as SCCmec III. CONCLUSIONS: The S. hominis isolates analyzed were variable biofilm producers had a high prevalence of methicillin resistance and resistance to other antibiotics, and high genetic diversity. The results of this study strongly suggested that S. hominis isolates harbor

  9. Molecular identification of Nocardia species using the sodA gene

    Directory of Open Access Journals (Sweden)

    K. Sánchez-Herrera

    2017-09-01

    Full Text Available Currently for bacterial identification and classification the rrs gene encoding 16S rRNA is used as a reference method for the analysis of strains of the genus Nocardia. However, it does not have enough polymorphism to differentiate them at the species level. This fact makes it necessary to search for molecular targets that can provide better identification. The sodA gene (encoding the enzyme superoxide dismutase has had good results in identifying species of other Actinomycetes. In this study the sodA gene is proposed for the identification and differentiation at the species level of the genus Nocardia. We used 41 type species of various collections; a 386 bp fragment of the sodA gene was amplified and sequenced, and a phylogenetic analysis was performed comparing the genes rrs (1171 bp, hsp65 (401 bp, secA1 (494 bp, gyrB (1195 bp and rpoB (401 bp. The sequences were aligned using the Clustal X program. Evolutionary trees according to the neighbour-joining method were created with the programs Phylo_win and MEGA 6. The specific variability of the sodA genus of the genus Nocardia was analysed. A high phylogenetic resolution, significant genetic variability, and specificity and reliability were observed for the differentiation of the isolates at the species level. The polymorphism observed in the sodA gene sequence contains variable regions that allow the discrimination of closely related Nocardia species. The clear specificity, despite its small size, proves to be of great advantage for use in taxonomic studies and clinical diagnosis of the genus Nocardia.

  10. Chemicals from Agave sisalana Biomass: Isolation and Identification

    Science.gov (United States)

    Santos, Jener David Gonçalves; Vieira, Ivo Jose Curcino; Braz-Filho, Raimundo; Branco, Alexsandro

    2015-01-01

    Agave sisalana (sisal) is known worldwide as a source of hard fibers, and Brazil is the largest producer of sisal. Nonetheless, the process of removing the fibers of the sisal leaf generates 95% waste. In this study, we applied chemical sequential steps (hydrothermal extraction, precipitation, liquid-liquid extraction, crystallization, SiO2 and Sephadex LH 20 column chromatography) to obtain pectin, mannitol, succinic acid, kaempferol and a mixture of saponins as raw chemicals from sisal biomass. The structural identification of these compounds was performed though spectrometric methods, such as Infrared (IR), Ultraviolet (UV), Mass spectrometry (MS) and Nuclear magnetic resonance (NMR). All the sisal chemicals found in this work are used by both the chemical and pharmaceutical industries as excipients or active principles in products. PMID:25903149

  11. Chemicals from Agave sisalana Biomass: Isolation and Identification

    Directory of Open Access Journals (Sweden)

    Jener David Gonçalves Santos

    2015-04-01

    Full Text Available Agave sisalana (sisal is known worldwide as a source of hard fibers, and Brazil is the largest producer of sisal. Nonetheless, the process of removing the fibers of the sisal leaf generates 95% waste. In this study, we applied chemical sequential steps (hydrothermal extraction, precipitation, liquid-liquid extraction, crystallization, SiO2 and Sephadex LH 20 column chromatography to obtain pectin, mannitol, succinic acid, kaempferol and a mixture of saponins as raw chemicals from sisal biomass. The structural identification of these compounds was performed though spectrometric methods, such as Infrared (IR, Ultraviolet (UV, Mass spectrometry (MS and Nuclear magnetic resonance (NMR. All the sisal chemicals found in this work are used by both the chemical and pharmaceutical industries as excipients or active principles in products.

  12. Evaluation of MALDI-TOF-MS for the Identification of Yeast Isolates Causing Bloodstream Infection.

    Science.gov (United States)

    Turhan, Ozge; Ozhak-Baysan, Betil; Zaragoza, Oscar; Er, Halil; Sarıtas, Zubeyde Eres; Ongut, Gozde; Ogunc, Dilara; Colak, Dilek; Cuenca-Estrella, Manuel

    2017-04-01

    Infections due to Candida species are major causes of morbidity and mortality in humans, causing a diverse spectrum of clinical disease ranging from superficial and mucosal infections to invasive disease. Several authors have demonstrated that mortality is closely linked to both timing of therapy and/or source control. The rapid identification of pathogenic species is helpful to start timely and effective antifungal therapy. The aim of this study was to assess the performance of the matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) system for the correct and rapid identification of yeast isolates causing bloodstream infection. Between January 2014 and January 2015, a total of 117 yeast like organisms isolated from blood culture samples of 117 episodes from 102 patients who had blood stream infections were included in the study. The isolates were identified by MALDI-TOF MS. The results were compared with those obtained by the standard mycological methods and/or sequence analysis. One hundred and seventeen yeast isolates including 115 Candida spp and two non-Candida yeasts were analysed. The Biotyper correctly identified 115 (98.3%) isolates to the genus level and 102 (87.2%) isolates to the species level using the manufacturer's recommended cutoff scores. The Bruker Biotyper is a rapid, easy, inexpensive, and highly reliable system for the identification of yeast isolates. Early identification with MALDI-TOF MS would save time for determination of antifungal susceptibility and proper treatment strategy. The expansion of the database of the library by addition of less common species will improve the performance of the system.

  13. Fusarium proliferatum isolated from garlic in Spain: identification, toxigenic potential and pathogenicity on related Allium species

    Directory of Open Access Journals (Sweden)

    Daniel PALMERO

    2012-05-01

    Full Text Available Fusarium proliferatum has been reported on garlic in the Northwest USA, Spain and Serbia, causing water-soaked tan-colored lesions on cloves. In this work, Fusarium proliferatum was isolated from 300 symptomatic garlic bulbs. Morphological identification of Fusarium was confirmed using species-specific PCR assays and EF-1α sequencing. Confirmation of pathogenicity was conducted with eighteen isolates. Six randomly selected F. proliferatum isolates from garlic were tested for specific pathogenicity and screened for fusaric acid production. Additionally, pathogenicity of each F. proliferatum isolate was tested on healthy seedlings of onion (Allium cepa, leek (A. porrum, scallions (A. fistulosum, chives (A. schoenoprasum and garlic (A. sativum. A disease severity index (DSI was calculated as the mean severity on three plants of each species with four test replicates. Symptoms on onion and garlic plants were observed three weeks after inoculation. All isolates tested produced symptoms on all varieties inoculated. Inoculation of F. proliferatum isolates from diseased garlic onto other Allium species provided new information on host range and pathogenicity. The results demonstrated differences in susceptibility with respect to host species and cultivar. The F. proliferatum isolates tested all produced fusaric acid (FA; correlations between FA production and isolate pathogenicity are discussed. Additionally, all isolates showed the presence of the FUM1 gene suggesting the ability of Spanish isolates to produce fumonisins.

  14. Isolation and identification of the human homolog of a new p53-binding protein, Mdmx

    NARCIS (Netherlands)

    Shvarts, A.; Bazuine, M.; Dekker, P.; Ramos, Y. F.; Steegenga, W. T.; Merckx, G.; van Ham, R. C.; van der Houven van Oordt, W.; van der Eb, A. J.; Jochemsen, A. G.

    1997-01-01

    We recently reported the identification of a mouse cDNA encoding a new p53-associating protein that we called Mdmx because of its structural similarity to Mdm2, a well-known p53-binding protein. Here we report the isolation of a cDNA encoding the human homolog of Mdmx. The ORF of the cDNA encodes a

  15. Identification and typing of the yeast strains isolated from bili bili, a ...

    African Journals Online (AJOL)

    Seventy six yeast strains isolated form bili bili and others sample were identified and typed in purpose of selecting appropriate starter culture. Identification techniques included conventional phenetic method, PCR/RFLP of NTS2 rDNA region, partial sequencing of the D1/D2 region of 26S rDNA and karyotyping using ...

  16. Isolation and identification methods of Rothia species in oral cavities.

    Science.gov (United States)

    Tsuzukibashi, Osamu; Uchibori, Satoshi; Kobayashi, Taira; Umezawa, Koji; Mashimo, Chiho; Nambu, Takayuki; Saito, Masanori; Hashizume-Takizawa, Tomomi; Ochiai, Tomoko

    2017-03-01

    Rothia dentocariosa and Rothia mucilaginosa which are Gram-positive bacteria are part of the normal flora in the human oral cavity and pharynx. Furthermore, Rothia aeria, which was first isolated from air samples in the Russian space station Mir, is predicted to be an oral inhabitant. Immunocompromised patients are often infected by these organisms, leading to various systemic diseases. The involvement of these organisms in oral infections has attracted little attention, and their distribution in the oral cavity has not been fully clarified because of difficulties in accurately identifying these organisms. A suitable selective medium for oral Rothia species, including R. aeria, is necessary to assess the veritable prevalence of these organisms in the oral cavity. To examine the bacterial population in the oral cavity, a novel selective medium (ORSM) was developed for isolating oral Rothia species in this study. ORSM consists of tryptone, sodium gluconate, Lab-Lemco powder, sodium fluoride, neutral acriflavin, lincomycin, colistin, and agar. The average growth recovery of oral Rothia species on ORSM was 96.7% compared with that on BHI-Y agar. Growth of other representative oral bacteria, i.e. genera Streptococcus, Actinomyces, Neisseria, and Corynebacterium, was remarkably inhibited on the selective medium. PCR primers were designed based on partial sequences of the 16S rDNA genes of oral Rothia species. These primers reacted to each organism and did not react to other non-oral Rothia species or representative oral bacteria. These results indicated that these primers are useful for identifying oral Rothia species. A simple multiplex PCR procedure using these primers was a reliable method of identifying oral Rothia species. The proportion of oral Rothia species in saliva samples collected from 20 subjects was examined by culture method using ORSM. Rothia dentocariosa, Rothia mucilaginosa, and R. aeria accounted for 1.3%, 5.9%, and 0.8% of the total cultivable

  17. Molecular characterization of Acinetobacter baumannii isolated from Iraqi hospital environment

    Directory of Open Access Journals (Sweden)

    I.M.S. AL-Kadmy

    2018-01-01

    Full Text Available Healthcare-associated items are a common source of acquired infections, and hospital-acquired infections cause significant mortality and morbidity worldwide. Acinetobacter baumannii is the most prevalent infection-causing organism in the hospital environment. Hospital articles and objects are the main sources of infection with the ability to transmit some of the pathogenic microorganisms such as A. baumannii, which is considered a serious problem in therapeutic treatments. In the current study, we isolated A. baumannii from hospital sources and evaluated its antibiotic resistance, virulence factors and resistance gene determinants. The isolates were identified phenotypically as well as genotypically using PCR. In addition, their capability for biofilm formation and ten other virulence factors were measured. Of 112 samples, 21 showed growth of the target organism. Apart from A. baumannii, isolates of Candida albicans, Staphylococcus sp., Pseudomonas aeruginosa, Escherichia coli and Klebsiella pneumoniae were also grown. Antibiotic susceptibility test results considered all the A. baumannii to be multidrug-resistant isolates with the highest resistance being 100% to gentamycin, ciprofloxacin; the most effective antibiotics with 100% susceptibility was colistin and tigecycline. All A. baumannii isolates had MIC for ceftriaxone >32 mg/L. All A. baumannii isolates from the hospital environment showed multidrug resistance and had many virulence factors. They have long-term resistance to dry conditions and cause a serious public health issue.

  18. Isolation and identification of acetogenic bacteria obtained from deer rumen and their potential for methanogenesis inhibitor

    Directory of Open Access Journals (Sweden)

    Amlius Thalib

    2008-10-01

    Full Text Available Methanogenesis can be inhibited by various chemicals through different mechanism reaktion. The use of acetogenic bacteria as H2 sink is assumed to be a promising approach. Isolation and identification of acetogenic bacteria obtained from deer rumen had been conducted. Two types of media used for isolation were hydrogen-carbondioxide utilizing acetogens and carbonmonoxide utilizing acetogens. Identification of species of acetogens isolates was based on descriptions of morphology, Gram type, motility, bioreaction results, and oksygen requirement. The compositions of methane and volatile fatty acids (VFA were determined on minimal media or added with sheep rumen liquid innoculated with pure isolates. The identification results showed that the isolate cultured on media of hydrogen-carbondioxide utilizing acetogens was Acetoanaerobium noterae and the ones cultured on media of carbonmonoxide utilizing acetogens was Acetobacterium woodii. Inoculumn of A. noterae and A. woodii could decreased the composition of methane resulted from substrate fermented by fresh rumen liquid of sheep (CRDF, that is culture of A. noterae added FPM and defaunator decreased methane production by 28.8% (P CH3COOH + 2H2O by which reduction of CO2 with H2 producing CH4 can be inhibited or decreased. Their function as methanogenesis inhibitor would be more significant when they are combined with microbial growth factors and defaunator.

  19. Dental morphology and variation in theropod dinosaurs: implications for the taxonomic identification of isolated teeth.

    Science.gov (United States)

    Smith, Joshua B; Vann, David R; Dodson, Peter

    2005-08-01

    Isolated theropod teeth are common Mesozoic fossils and would be an important data source for paleoecology biogeography if they could be reliably identified as having come from particular taxa. However, obtaining identifications is confounded by a paucity of easily identifiable characters. Here we discuss a quantitative methodology designed to provide defensible identifications of isolated teeth using Tyrannosaurus as a comparison taxon. We created a standard data set based as much as possible on teeth of known taxonomic affinity against which to compare isolated crowns. Tooth morphology was described using measured variables describing crown length, base length and width, and derived variables related to basal shape, squatness, mesial curve shape, apex location with respect to base, and denticle size. Crown curves were described by fitting the power function Y = a + bX(0.5) to coordinate data collected from lateral-view images of mesial curve profiles. The b value from these analyses provides a measure of curvature. Discriminant analyses compared isolated teeth of various taxonomic affinities against the standard. The analyses classified known Tyrannosaurus teeth with Tyrannosaurus and separated most teeth known not to be Tyrannosaurus from Tyrannosaurus. They had trouble correctly classifying teeth that were very similar to Tyrannosaurus and for which there were few data in the standard. However, the results indicate that expanding the standard should facilitate the identification of numerous types of isolated theropod teeth. (c) 2005 Wiley-Liss, Inc.

  20. Molecular epidemiology and antifungal susceptibility of Saprochaete capitata (Blastoschizomyces capitatus) isolates causing nosocomial infection in Kayseri/Turkey.

    Science.gov (United States)

    Koç, A Nedret; Atalay, M Altay; Timur, Demet; Demir, Gonca; Kaynar, Leylagül

    2016-08-01

    Saprochaete capitata isolates have emerged as important nosocomial pathogens, among immunosuppressed or neutropenic patients, and a rare cause of nosocomial infection in the hematology-bone marrow unit (HBMU) and the intensive care unit (ICU). The purpose of this study was to molecular epidemiology and antifungal susceptibility of S. capitata (Blastoschizomyces capitatus) isolates causing nosocomial infection at Kayseri in Turkey. During a period from 2012 to 2015, a total of 20 S. capitata strains were obtained from patients hospitalized at Erciyes University Hospital. The identification of S. capitata was performed by phenotypic and biochemical methods; this was confirmed by molecular methods by DNA sequencing analysis. Genotyping of S.capitata isolates from different patients was determined to by the repetitive sequence PCR (repPCR) using the DiversiLab System (BioMerieux). More than half of the patients with S. capitata infections were hospitalized in the hematology-oncology unit (60%). The patients mainly included those using intravascular devices (90%), and receiving parenteral antibiotics (85%); the mortality rate was 55%. The microbiological investigation failed to identify S. capitata in the hospital environment. All isolates were resistant to caspofungin (>32). However, the MIC90 values for voriconazole, amphotericin B, and fluconazole against all of the isolates were 0.125, 0.25, and 1μg/ml, respectively. The S. capitata strains belonged to five clones (A-E) which were determined by the use of rep-PCR and Clone C was found to be predominant. S. capitata isolates are an important cause of nosocomial infection in the HBMU and ICUs.

  1. Identification and antimicrobial susceptibility patterns of Staphylococcus spp. isolated from canine chronic otitis externa

    Directory of Open Access Journals (Sweden)

    Silva N.

    2001-01-01

    Full Text Available Swab samples obtained from 96 dogs with chronic otitis externa were cultured for the isolation of Staphylococcus species. Of 57 staphylococcal strains, 41 (72% were coagulase-negative (CNS. The identification of staphylococci strains was made by standard procedures for the routine identification of staphylococci in clinical practice. S. sciuri was the most frequent species isolated (22.8% from chronic otitis externa in dogs followed by S. intermedius (12.3%, S. auricularis (10.5% and S. aureus (8.8%. Three (5.2% CNS strains could not be identified. Bacterial isolates were susceptible to enrofloxacin, gentamicin, cephalothin, chloramphenicol and neomycin. Resistance was most common to penicillin G, oxacillin and ampicillin.

  2. Identification of Malassezia species isolated from patients with extensive forms of pityriasis versicolor in Siena, Italy.

    Science.gov (United States)

    Romano, Clara; Mancianti, Francesca; Nardoni, Simona; Ariti, Gaetano; Caposciutti, Paola; Fimiani, Michele

    2013-01-01

    Pityriasis versicolor (PV) is an infection caused by various species of Malassezia yeast. There is no agreement in the literature concerning the species of Malassezia and the demographic, clinical, and mycological data. To prospectively identify Malassezia species isolated from lesions of patients with extensive, long standing and recurrent forms of PV and to estimate the relationship between Malassezia species and the demographic and clinical data of the patients. All patients with PV were enrolled over a four-year period. Malassezia species were isolated in cultures and identified by morphological features and physiological tests. In the last 2 years a PCR-based technique was used to confirm the species' identification. A total of 74 patients (43 males and 31 females, mean age 39.5 years) were enrolled. Only one species was isolated in 45 patients, and more than one species were identified in the remaining 28 patients (38%). M. globosa was the most frequently isolated (60.3%) species. There was a significant association between the isolation of 2 or more species and the presence of at least one predisposing factor. In the last 29 cases, which were subjected to PCR, there were no differences in the identification of isolated species as compared to traditional methods. The isolation of more than one species in a single lesion is not infrequent in PV and is related to the presence of one predisposing factor. The isolated species isolated were not influenced by demographic and clinical features. The traditional and more recent (PCR) procedures gave the same results in the isolated species. Copyright © 2012 Revista Iberoamericana de Micología. Published by Elsevier Espana. All rights reserved.

  3. Molecular characterization of two isolates of sweet potato leaf curl ...

    African Journals Online (AJOL)

    Jane

    2011-08-17

    Aug 17, 2011 ... Xie Y, Zhou XP, Li ZH, Zhang ZK, Li GX (2002). Identification of a novel. DNA molecule associated with Tobacco leaf curl virus. Chin. Sci. Bull. 47: 1273-1276. Zhou XP, Xie Y, Peng Y, Zhang ZK (2003). Malvastrum yellow vein virus, a new begomovirus species associated with satellite DNA molecule. Chin.

  4. Molecular and serological characterization of Leptospira interrogans serovar Canicola isolated from dogs, swine, and bovine in Brazil

    NARCIS (Netherlands)

    Miraglia, Fabiana; de Morais, Zenaide M.; Dellagostin, Odir A.; Seixas, Fabiana K.; Freitas, Julio C.; Zacarias, Francielle G. S.; Delbem, Adina C.; Ferreira, Thaís S. P.; Souza, Gisele O.; Hartskeerl, Rudy A.; Vasconcellos, Silvio A.; Moreno, Andrea M.

    2012-01-01

    The identification of Leptospira clinical isolates through genotyping and serotyping, besides the recognition of its reservoirs, are important tools for understanding the epidemiology of leptospirosis, and they are also keys for identifying new species and serovars. Fourteen clinical isolates from

  5. Molecular identification and characterization of Anaplasma platys and Ehrlichia canis in dogs in Mexico.

    Science.gov (United States)

    Almazán, Consuelo; González-Álvarez, Vicente H; Fernández de Mera, Isabel G; Cabezas-Cruz, Alejandro; Rodríguez-Martínez, Rafael; de la Fuente, José

    2016-03-01

    The tick-borne pathogens Ehrlichia canis and Anaplasma platys are the causative agents of canine monocytic ehrlichiosis (CME) and canine cyclic thrombocytopenia (CCT). Although molecular evidence of E. canis has been shown, phylogenetic analysis of this pathogen has not been performed and A. platys has not been identified in Mexico, where the tick vector Rhipicephalus sanguineus sensu lato (s.l.) is common. The aim of this research was to screen, identify and characterize E. canis and A. platys by PCR and phylogenetic analysis in dogs from La Comarca Lagunera, a region formed by three municipalities, Torreon, Gomez-Palacio and Lerdo, in the Northern states of Coahuila and Durango, Mexico. Blood samples and five engorged R. sanguineus s.l. ticks per animal were collected from 43 females and 57 male dogs presented to veterinary clinics or lived in the dog shelter from La Comarca Lagunera. All the sampled dogs were apparently healthy and PCR for Anaplasma 16S rRNA, Ehrlichia 16S rRNA, and E. canis trp36 were performed. PCR products were sequenced and used for phylogenetic analysis. PCR products were successfully amplified in 31% of the samples using primers for Anaplasma 16S rRNA, while 10% and 4% amplified products using primers for Ehrlichia 16S rRNA and E. canis trp36 respectively. Subsequent sequencing and phylogenetic analyses of these products showed that three samples corresponded to A. platys and four to E. canis. Based on the analysis of trp36 we confirmed that the E. canis strains isolated from Mexico belong to a conservative clade of E. canis and are closely related to strains from USA. In conclusion, this is the first molecular identification of A. platys and the first molecular characterization and phylogenetic study of both A. platys and E. canis in dogs in Mexico. Copyright © 2015 Elsevier GmbH. All rights reserved.

  6. Species identification and antifungal susceptibility pattern of Candida isolates in cases of vulvovaginal

    Directory of Open Access Journals (Sweden)

    Dalia Saad ElFeky

    2016-09-01

    Full Text Available Vulvovaginal candidiasis (VVC remains one of the most common infections of the female genital tract. Correct identification of the isolated Candida species is essential to direct the empirical antifungal therapy. Objectives: This local study was conducted to identify the spectrum of Candida species associated with VVC using different phenotypic and genotypic methods and assess their antifungal susceptibility pattern. Materials and methods: High vaginal swabs were collected from 125 patients presenting with a clinical picture suggestive of VVC. Swabs were subjected to Gram-stain and culture on Sabouraud dextrose agar. Species identification of Candida isolates was done using phenotypic methods including germ tube test, Rice Tween-80 agar, Chrom ID (CAN2 agar and API 20C AUX, while PCR-RFLP was used as the gold standard method. Antifungal susceptibility testing was done using the disk diffusion method. Results: Vaginal swab cultures yielded Candida growth in 63 cases (50.4%. Candida albicans was the predominant isolated species (60.3% while the most common non-albicans species was Candida glabrata (12.7%. Forty-five (71.4% and fifty-five (87.3% Candida isolates were correctly speciated by Rice Tween-80 Agar and API 20C AUX, respectively, while fifty-seven isolates (90.5% were correctly assigned into the 3 groups of yeasts identified by CAN2 agar. Amphotericin B was more effective than azoles against vaginal Candida isolates. Conclusion: C. albicans is the most common species associated with VVC. API 20C AUX was the most accurate phenotypic method for the proper identification of most Candida species whereas PCR-RFLP could properly confirm Candida species identification genotypically.

  7. Isolation and identification of main mastitis pathogens in Mexico

    Directory of Open Access Journals (Sweden)

    H. Castañeda Vázquez

    2013-04-01

    Full Text Available The present work is a large epidemiological study aiming to detect the prevalence of subclinical mastitis and to investigate the major udder pathogens in Jalisco State, western Mexico. For this purpose, 2205 dairy cows, representing 33 Mexican dairy herds, were involved. Of 2205 cows, 752 mastitic animals were diagnosed and only 2,979 milk samples could be obtained for further investigation. All 2979 milk samples were subjected to California Mastitis Test (CMT to differentiate clinical cases from subclinical ones where 1996 samples (67 % reacted positively. Of these, 1087 samples (54.5% came from cows suffering from clinical cases of mastitis. Bacteriological identification of the causative agents revealed the presence of a major group of pathogens including the Coagulase negative staphylococci (CNS, S.aureus, S.agalactiae, Corynebacterium spp. and Coliform bacteria which were detected in 464 (15.6%, 175 (5.9%, 200 (6.8%, 417 (14% and 123 (4.1% of the 2927 investigated quarters, 295 (15.4%, 118 (15.7%, 111 (14.8%, 227 (30.2% and 109 (14.5% of the 752 examined cows and in 33 (100%, 22 (66.7%, 19 (57.6%, 30 (90.1% and 27 (81.8% of the 33 herds involved, respectively. Other pathogens could be detected in the investigated milk samples such as S. dysgalactiae (0.4%, S.uberis (0.37%, Bacillus spp. (1%, Nocardia spp. (0.6% und Candida spp. (0.1%. Meanwhile, others were present in a negligible ratio; including the Aerococcus viridans, and Enterococcus spp., Lactococcus lactis, S. bovis.

  8. Isolation, identification and characterization of an electrogenic microalgae strain.

    Directory of Open Access Journals (Sweden)

    Yicheng Wu

    Full Text Available Extracellular electron transfer involving microbes is important as it closely reflects the ability of cells to communicate with the environment. However, there are few reports on electron transfer mechanisms of pure microalgae and a lack of any model alga to study the transfer processes. In the present study, nine green microalgae species were isolated from wastewater and characterized in terms of their ability to transfer electrons between cells and an electrode. One species showed direct electron transfer via membrane-associated proteins and indirect electron transfer via secreted oxygen. The microalga was identified as Desmodesmus sp. based on phylogenetic analysis and electron microscopy. Electrochemical tests demonstrated that Desmodesmus sp. was able to act as a cathodic microorganism. Stable current densities of -0.24, 35.54 and 170 mA m(-2 were achieved at potentials of +0.2, -0.2 and -0.4 V, respectively, under illumination. Dissolved oxygen concentration measurement showed gradients within the microalgae biofilm: 18.3 mg L(-1 in light decreasing to 4.29 mg L(-1 in the dark. This study diversified the exoelectrogen library and provided a potential model microalga to explore the associated mechanism of extracellular electron transfer.

  9. Isolation, identification and characterization of novel Bacillus subtilis.

    Science.gov (United States)

    Lu, Zhenxiang; Guo, Weina; Liu, Chang

    2018-03-24

    In this study, we have identified a bacterium that can inhibit the growth of Staphylococcus aureus, and further analyzed its antibacterial activity and other biological characteristics and laid the foundation for its future application. Through isolation and culture of the unknown bacteria, the culture characteristics, morphology observation, biochemical test, preliminary antibacterial test, 16S rRNA PCR amplification, sequence analysis, and homology analysis were performed. It was found that the bacteria are Gram positive spore chain Bacillus. The bacteria could only ferment glucose for acid production, but could not utilize lactose and maltose. The VP test for this bacteria was positive, while indole and methyl red tests were negative. Further analysis showed that these bacteria shared a homology up to 99.4% with Bacillus subtilis DQ198162.1. Thus, this newly identified bacterium was classified as Bacillus subtilis. Importantly, the crude bacteriocin of this Bacillus subtilis could inhibit the growth of Staphylococcus aureus, Escherichia coli, Enterococcus and Salmonella, which implies its potential usage in the future.

  10. Identification of Clinical Isolates of Candida using Duplex PCR

    Directory of Open Access Journals (Sweden)

    Homeyra Babaei

    2016-11-01

    Full Text Available Abstract Background: Candida albicans is still the main etiologic agent of candidiasis. However, infections of non-albicans Candida species are increasing. Candida dubliniensis is similar to C. albicans phenotypically and must be identified due to the better management of infection. The aim of the present study is to defferentiate and identify Candida species by Duplex PCR for get-ting an epidemiological data of Candida species among clinical specimens. Materials and Methods: DNA was extracted using phenol-chloroform method from fresh colonies. Internal Transcribed Spacer region was amplified by polymerase chain reaction using specific primers. Based on differences of bands sizes on agarose gel electrophoresis, species were identified. Results: Ninety four out of 100 patients (49 males and 51 females had predisposing fac-tors in the present study. Diabetes (73.4%, use of antibiotic (6.3%, vitamin deficiency (4.3% were the main predisposing factors. The most specimens belonged to mouth (75%, vagina (5%, and blood (4%. All isolates were identified as C. albicans. Conclusion: Duplex PCR is a rapid and precise method for the detection and differentia-tion of Candida species carefully, and in this method, phenotypic tests like germ-tube and chla-mydoconidia production, as well as biochemical tests are not required for clinical laboratories that have limited resources and time for response to the patients, and it can replace with the traditional methods.

  11. Caracterização morfofisiológica e identificação molecular de isolados de Guignardia citricarpa, agente patogênico da mancha preta dos citros = Morphophysiological characterization and molecular identification of isolates of Guignardia citricarpa, a pathogenic agent of the citrus black spot

    Directory of Open Access Journals (Sweden)

    Marilda Pereira Caixeta

    2008-12-01

    Full Text Available O presente trabalho teve como objetivo identificar 11 isolados de Guignardia citricarpa, agente causal da mancha preta dos citros (MPC, obtidos de frutas cítricas sintomáticas de diferentes regiões geográficas, por meio da PCR e caracterização morfofisiológica das estruturas propagativas, esporulação e crescimento micelial emdiferentes meios de cultura, temperaturas e regimes de luz, nas condições de laboratório. Pelo teste de PCR, todos os isolados foram identificados como o patógeno G. citricarpa. Os isolados caracterizados foram submetidos às temperaturas de 20, 25 e 30ºC, em regime de luz contínua, escuro contínuo e fotoperíodos de 12 horas, durante 24 dias. Utilizaram-se os meios de cultura aveia-ágar (AA, batata-dextrose-ágar (BDA e cenoura-dextrose-ágar (CDA. Os resultados mostraram que ocorreu interação entre os diferentes meios de cultura, temperaturas e fotoperíodos. O meio de cultura que melhor estimulou o crescimento micelial foi o CDA a 25ºC sob o fotoperíodo de 12h. A maior produção de esporos (conídios foi verificada no meio BDA a 20ºC, no fotoperíodo de 12 horas. No meio CDA, não ocorreu esporulação de nenhum isolado. Sob a temperatura de 30ºC, foiverificada apenas a produção de hifas e picnídios para a maioria dos isolados, em todos os meios de cultura e fotoperíodo testados.Thepresent work aims to identify 11 isolates of Guignardia citricarpa, the causal agent of the citrus black spot (CBS, obtained from affected fruit in different geographical regions, through PCR and morphophysiological characterization of propagative structures, sporing and mycelial growth in different means of culture, temperatures and photoperiods, under laboratory conditions. Through the PCR test, all isolates were identified as being the G.citricarpa pathogen. The characterized isolates were subjected to evaluations at temperatures of 20, 25 and 30ºC, in continuous light, continuous darkness, and alternating 12

  12. Molecular identification in metabolomics using infrared ion spectroscopy

    NARCIS (Netherlands)

    Martens, J.; Berden, G.; van Outersterp, R.E.; Kluijtmans, L.A.J.; Engelke, U.F.; van Karnebeek, C.D.M.; Wevers, R.A.; Oomens, J.

    2017-01-01

    Small molecule identification is a continually expanding field of research and represents the core challenge in various areas of (bio) analytical science, including metabolomics. Here, we unequivocally differentiate enantiomeric N-acetylhexosamines in body fluids using infrared ion spectroscopy,

  13. Molecular identification of Paragonimus species by DNA pyrosequencing technology.

    Science.gov (United States)

    Tantrawatpan, Chairat; Intapan, Pewpan M; Janwan, Penchom; Sanpool, Oranuch; Lulitanond, Viraphong; Srichantaratsamee, Chutatip; Anamnart, Witthaya; Maleewong, Wanchai

    2013-06-01

    DNA pyrosequencing for PCR amplicons is an attractive strategy for the identification of microorganisms because of its short time performance for large number of samples. In this study, the primers targeting the fragment of ITS2 region of nuclear ribosomal RNA gene were newly developed for pyrosequencing-based identification of 6 Paragonimus species, Paragonimus bangkokensis, Paragonimus harinasutai, Paragonimus heterotremus, Paragonimus macrorchis, Paragonimus siamensis and Paragonimus westermani. Pyrosequencing determination of 39 nucleotides of partial ITS2 region could discriminate 6 Paragonimus species, and could also detect intra-species genetic variation of P. macrorchis. This DNA pyrosequencing-based identification can be a valuable tool to improve species-level identification of Paragonimus in the endemic areas. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

  14. Molecular identification of Anopheles gambiae sensu stricto Giles ...

    African Journals Online (AJOL)

    User

    2016-09-28

    baited bed net traps for subsequent identification of the .... climatic and ecological zone (NEMA, 2007) and were surrounded by a variety of ... mosquitoes that entered a trap during any hour were those actively seeking hosts, and, in ...

  15. Molecular Identification of Bacteria Involved in Degradation of Crude ...

    African Journals Online (AJOL)

    Lactobacillus spp. G22 respectively. However, isolate 1, 8, 9, 10, 11, 12, 13, 14, 15 and 16 did not produce any result. This study shows that microorganisms isolated from a non native crude oil contaminated site can be utilized for bioremediation. Key words: Bioremediation, 16s rRNA gene, DNA sequencing, Biotechnology ...

  16. Molecular identification of uncommon clinical yeast species in Iran

    Directory of Open Access Journals (Sweden)

    Ladan Karimi

    2015-03-01

    Conclusion: We identified several rare clinical isolates selected from a big collection at the species level by ITS-sequencing. As the list of yeast species as opportunistic human fungal infections is increasing dramatically, and many isolates remain unidentified using conventional methods, more sensitive and specific advanced approaches help us to clarify the aspects of microbial epidemiology of the yeast infections.

  17. Molecular and pathological identification of feline coronavirus type I

    African Journals Online (AJOL)

    DR. NJ TONUKARI

    2012-06-05

    Jun 5, 2012 ... characterization of naturally occurring feline coronavirus from domestic cat in Malaysia. Additionally, the resultant ... to virus isolation in Felis catus whole fetus cell cultures (Fcwf-4). The result of virus ... coronavirus in Malaysian cats and importantly the isolated virus was confirmed to be type I using S-.

  18. Molecular typing of Mycobacterium tuberculosis isolates circulating in Jiangsu province, China.

    Science.gov (United States)

    Liu, Qiao; Yang, Dandan; Xu, Weiguo; Wang, Jianming; LV, Bing; Shao, Yan; Song, Honghuan; Li, Guoli; Dong, Haiyan; Wan, Kanglin; Wang, Hua

    2011-10-26

    Globally, China is the second place with high burden of tuberculosis (TB). To explore the characteristics of the pathogens of Mycobacterium tuberculosis (MTB) circulating in this area is helpful for understanding and controlling the spread of the strains. Recent developments in molecular biology have allowed prompt identification and tracking specific strains of MTB spreading through the population. Spacer-oligonucleotide typing (spoligotyping) and mycobacterial interspersed repetitive units variable number tandem repeat (MIRU-VNTR) were performed in combination to yield specific genetic profiles of 260 MTB strains isolated from 30 counties of Jiangsu province in China between June and July 2010. The spoligotyping results were in comparison to the world Spoligotyping Database of Institute Pasteur de Guadeloupe (SpolDB4). Drug susceptibility test (DST) was performed on all strains by proportion method on Lowenstein-Jensen (LJ) culture media. Based on the spoligotyping method, 246 strains displayed known patterns and 14 were absent in the database. Predominant spoligotypes belonged to the Beijing family (80.4%). By using the 24-loci VNTR typing scheme, 224 different patterns were identified, including 20 clusters and 204 unique patterns. The largest clade comprised 195 strains belonging to the Beijing family. The combination of spoligotyping and 24-loci MIRU-VNTR demonstrated maximal discriminatory power. Furthermore, we observed a significant association between Beijing family strains and drug-resistant phenotypes. The Beijing family strains presented increased risks for developing multi-drug resistant TB, with the OR (95% CI) of 11.07(1.45-84.50). The present study demonstrated that Beijing family isolates were the most prevalent strains circulating in Jiangsu province of China. The utility of spoligotyping in combination with 24-loci MIRU-VNTR might be a useful tool for epidemiological analysis of MTB transmission.

  19. Molecular typing of mycobacterium tuberculosis isolates circulating in Jiangsu Province, China

    Directory of Open Access Journals (Sweden)

    Dong Haiyan

    2011-10-01

    Full Text Available Abstract Background Globally, China is the second place with high burden of tuberculosis (TB. To explore the characteristics of the pathogens of Mycobacterium tuberculosis (MTB circulating in this area is helpful for understanding and controlling the spread of the strains. Recent developments in molecular biology have allowed prompt identification and tracking specific strains of MTB spreading through the population. Methods Spacer-oligonucleotide typing (spoligotyping and mycobacterial interspersed repetitive units variable number tandem repeat (MIRU-VNTR were performed in combination to yield specific genetic profiles of 260 MTB strains isolated from 30 counties of Jiangsu province in China between June and July 2010. The spoligotyping results were in comparison to the world Spoligotyping Database of Institute Pasteur de Guadeloupe (SpolDB4. Drug susceptibility test (DST was performed on all strains by proportion method on Lowenstein-Jensen (LJ culture media. Results Based on the spoligotyping method, 246 strains displayed known patterns and 14 were absent in the database. Predominant spoligotypes belonged to the Beijing family (80.4%. By using the 24-loci VNTR typing scheme, 224 different patterns were identified, including 20 clusters and 204 unique patterns. The largest clade comprised 195 strains belonging to the Beijing family. The combination of spoligotyping and 24-loci MIRU-VNTR demonstrated maximal discriminatory power. Furthermore, we observed a significant association between Beijing family strains and drug-resistant phenotypes. The Beijing family strains presented increased risks for developing multi-drug resistant TB, with the OR (95% CI of 11.07(1.45-84.50. Conclusions The present study demonstrated that Beijing family isolates were the most prevalent strains circulating in Jiangsu province of China. The utility of spoligotyping in combination with 24-loci MIRU-VNTR might be a useful tool for epidemiological analysis of MTB

  20. Molecular analysis of clinical isolates previously diagnosed as Mycobacterium intracellulare reveals incidental findings of "Mycobacterium indicus pranii" genotypes in human lung infection.

    Science.gov (United States)

    Kim, Su-Young; Park, Hye Yun; Jeong, Byeong-Ho; Jeon, Kyeongman; Huh, Hee Jae; Ki, Chang-Seok; Lee, Nam Yong; Han, Seung-Jung; Shin, Sung Jae; Koh, Won-Jung

    2015-09-30

    Mycobacterium intracellulare is a major cause of Mycobacterium avium complex lung disease in many countries. Molecular studies have revealed several new Mycobacteria species that are closely related to M. intracellulare. The aim of this study was to re-identify and characterize clinical isolates from patients previously diagnosed with M. intracellulare lung disease at the molecular level. Mycobacterial isolates from 77 patients, initially diagnosed with M. intracellulare lung disease were re-analyzed by multi-locus sequencing and pattern of insertion sequences. Among the 77 isolates, 74 (96 %) isolates were designated as M. intracellulare based on multigene sequence-based analysis. Interestingly, the three remaining strains (4 %) were re-identified as "Mycobacterium indicus pranii" according to distinct molecular phylogenetic positions in rpoB and hsp65 sequence-based typing. In hsp65 sequevar analysis, code 13 was found in the majority of cases and three unreported codes were identified. In 16S-23S rRNA internal transcribed spacer (ITS) sequevar analysis, all isolates of both species were classified within the Min-A ITS sequevar. Interestingly, four of the M. intracellulare isolates harbored IS1311, a M. avium-specific element. Two of three patients infected with "M. indicus pranii" had persistent positive sputum cultures after antibiotic therapy, indicating the clinical relevance of this study. This analysis highlights the importance of precise identification of clinical isolates genetically close to Mycobacterium species, and suggests that greater attention should be paid to nontuberculous mycobacteria lung disease caused by "M. indicus pranii".

  1. [Isolation identification and characterization of halotolerant petroleum-degrading bacteria].

    Science.gov (United States)

    Wu, Tao; Xie, Wen-Jun; Yi, Yan-Li; Li, Xiao-Bin; Wang, Jun; Hu, Xiang-Ming

    2012-11-01

    To obtain efficient halotolerant petroleum-degrading bacteria, 39 bacteria strains were isolated from 30 petroleum contaminated saline soil samples in Yellow River Delta, an important base of petroleum production in China. One bacterium (strain BM38) was found to efficiently degrade crude oil in highly saline environments based on a series of liquid and soil incubation experiments. According to its morphology, physiochemical characteristics and 16S rDNA sequence analysis, this strain was identified as Pseudomonas putida. Moreover, a series of liquid incubation experiments were conducted to investigate its characteristics such as halotolerance, biosurfactants production and degrading efficiency for various hydrocarbons. The salt resistance test demonstrated that strain BM38 grew well at NaCl concentrations ranging from 0.5% to 6.0%. Petroleum degradation experiments showed that strain BM38 could degrade 73.5% crude oil after 7 days in a liquid culture medium containing 1.0% NaCl and remove more than 40% of total petroleum hydrocarbons after 40 days in the soil with 0.22% and 0.61% of salinity, these results proved that the strain was effective in removing petroleum hydrocarbons. Strain BM38 could produce a bioemulsifier in a liquid culture medium. The NaCl concentration had the significant effect on the EI24 of fermentation broth, which decreased sharply if the NaCl concentration was greater than 1.0%. However, the EI24 of BM38 was still quite high in the presence of 2.0% of NaCl, and the value was 61.0%. Furthermore, this strain was also able to grow in mineral liquid media amended with hexadecane, toluene, phenanthrene, isooctane and cyclohexane as the sole carbon sources. Among these hydracarbons, strain BM38 showed relatively high ability in degrading n-alkanes and aromatic hydracarbons. The results indicated that strain BM38 had potential for application in bioremediation of petroleum-contaminated saline soil.

  2. Isolation and molecular characterization of Acanthamoeba genotypes isolated from soil sources of public and recreational areas in Iran.

    Science.gov (United States)

    Karamati, Seyed Ahmad; Niyyati, Maryam; Lorenzo-Morales, Jacob; Lasjerdi, Zohreh

    2016-12-01

    Pathogenic strains of Acanthamoeba are causative agents of a sight threating infection of the cornea known as Acanthamoeba keratitis. AK cases have been reported in Iran recently due to inappropriate usage of contact lens maintenance and most patients report a contact with contaminated sources such as dust, water or soil. Sixty soil samples were collected from public and recreational areas in the province of East Azerbaijan, Iran and checked for the presence of Acanthamoeba spp. Samples were cultured on non-nutrient agar plates seeded with heat killed Escherichia coli. PCR and sequencing of the DF3 region were carried out in order to genotype the isolated strains of Acanthamoeba. Thermotolerance and osmotolerance assays were performed in order to investigate the pathogenic potential of isolated Acanthamoeba strains. Acanthamoeba spp. was isolated from 41.6% of soil samples and genotyping of the strains resulted in the identification of genotypes T3, T4, T5 and T11. Most of the isolates belonging to genotypes T3 and T4 showed high pathogenic potential, indicating that they might present a potential health hazard for humans and other animals in this region. To the best of our knowledge, this is the first report on the identification of genotypes T3 and T11 from soil sources in the country.

  3. IDENTIFICATION OF SOIL FUNGI ISOLATED FROM ALFALFA (Medicago sativa L) TO FIND SPECIFIC FUNGI WHICH IMPROVED THE GROWTH OF ALFALFA

    OpenAIRE

    T. Yudiarti; S. Sumarsono; D.W. Widjayanto

    2014-01-01

    Objective of the study was to identify all kinds of fungi which can life in the alfalfa plantation inBaturaden Purwokerto-Central Java. Fungi used in this study was 38 isolates. All fungi have been takenfrom the isolation of soil and root of diseased plant. Macroscopic and microscopic methods were usedfor identification. Potato Dextrose Agar (PDA) medium was used to grow the fungi. All fungi wereidentified using book identification of fungi. The results showed that from 38 isolates, six speci...

  4. Molecular characterization of Azotobacter spp. nifH gene Isolated ...

    African Journals Online (AJOL)

    The nifH gene sequence of the nitrogen-fixing bacterium Azotobacter spp. was determined with the use of polymerase chain reaction (PCR). The Azotobacter species was isolated from marine source in two different seasons. They were cultivated under laboratory conditions using Nitrogen free Azotobacter specific medium.

  5. Isolation and molecular characterization of an ethylene response

    Indian Academy of Sciences (India)

    Apetala2/Ethylene Response Factors (AP2/ERF) play important roles in regulating gene expression under abiotic and biotic stress in the plant kingdom. Here, we isolated a member of the AP2/ERF transcription factors, NtERF1-1, from Nicotiana tabcum cv. Xanthi NN carrying the N gene, which is resistant to Tobacco ...

  6. First isolation and molecular characterization of Suid herpesvirus ...

    African Journals Online (AJOL)

    Since Aujeszky`s disease (pseudorabies), which is caused by Suid herpesvirus type 1 (SuHV-1), was first notified in Argentina in 1978, many SuHV-1 strains have been isolated from swine. However, this disease can affect other vertebrates, such as dogs (secondary hosts), and lead to fatal neurological disease.

  7. Molecular phylogeny of Escherichia coli isolated from clinical ...

    African Journals Online (AJOL)

    A, B1, B2 and D). Strains of these groups differ in their phenotypic characteristics, including the ability to use certain sugars, antibiotic resistance profiles and growth rate-temperature relationships. A total of 45 E. coli isolates were obtained from ...

  8. Molecular phylogeny of Escherichia coli isolated from clinical ...

    African Journals Online (AJOL)

    lames

    2011-11-09

    A, B1, B2 and D). Strains of these groups differ in their phenotypic characteristics, including the ability to use certain sugars, antibiotic resistance profiles and growth rate-temperature relationships. A total of 45 E. coli isolates ...

  9. Molecular detection and isolation of avian metapneumovirus in Mexico.

    Science.gov (United States)

    Rivera-Benitez, José Francisco; Martínez-Bautista, Rebeca; Ríos-Cambre, Francisco; Ramírez-Mendoza, Humberto

    2014-01-01

    We conducted a longitudinal study to detect and isolate avian metapneumovirus (aMPV) in two highly productive poultry areas in Mexico. A total of 968 breeder hens and pullets from 2 to 73 weeks of age were analysed. Serology was performed to detect aMPV antibodies and 105 samples of tracheal tissue were collected, pooled by age, and used for attempted virus isolation and aMPV nested reverse transcriptase-polymerase chain reaction (nRT-PCR). The serological analysis indicated that 100% of the sampled chickens showed aMPV antibodies by 12 weeks of age. Five pools of pullet samples collected at 3 to 8 weeks of age were positive by nRT-PCR and the sequences obtained indicated 98 to 99% similarity with the reported sequences for aMPV subtype A. Virus isolation of nRT-PCR-positive samples was successfully attempted using chicken embryo lung and trachea mixed cultures with subsequent adaptation to Vero cells. This is the first report of detection and isolation of aMPV in Mexico.

  10. Rapid isolation of high molecular weight DNA from single dried ...

    African Journals Online (AJOL)

    ANAND

    For studying genetic diversity in populations of predatory coccinellid, Cryptolaemus montrouzieri. Mulsant (Coccinellidae: Coleoptera), our attempts to isolate high quality DNA from individual adult beetle using several previously reported protocols and even modifications were quite unsuccessful as the insect size was small ...

  11. Isolation and molecular genetic characterization of a yeast strain ...

    African Journals Online (AJOL)

    STORAGESEVER

    2009-05-18

    May 18, 2009 ... optimal in situ bioremediation strategies. In the current study, three yeast strains were isolated from oil- contaminated soil by enrichment technique in mineral basal salts (MBS) medium supplemented with phenanthrene as a sole carbon source. Out of these, strain AH70 was selected for PAHs degradation,.

  12. Molecular characterization of Pseudomonas syringae pv. tomato isolates from Tanzania

    DEFF Research Database (Denmark)

    Shenge, K.C.; Stephan, D.; Mabagala, R. B.

    2008-01-01

    Bacterial speck caused by Pseudomonas syringae pv. tomato is an emerging disease of tomato in Tanzania. Following reports of outbreaks of the disease in many locations in Tanzania, 56 isolates of P. syringae pv. tomato were collected from four tomato- producing areas and characterized using patho...

  13. Rapid isolation of high molecular weight DNA from single dry ...

    African Journals Online (AJOL)

    For studying genetic diversity in populations of predatory coccinellid, Cryptolaemus montrouzieri Mulsant (Coccinellidae: Coleoptera), our attempts to isolate high quality DNA from individual adult beetle using several previously reported protocols and even modifications were quite unsuccessful as the insect size was small ...

  14. Molecular characterization of Azotobacter spp. nifH gene Isolated ...

    African Journals Online (AJOL)

    STORAGESEVER

    2009-12-15

    Dec 15, 2009 ... The nifH gene sequence of the nitrogen-fixing bacterium Azotobacter spp. was determined with the use of polymerase chain reaction (PCR). The Azotobacter species was isolated from marine source in two different seasons. They were cultivated under laboratory conditions using Nitrogen free Azotobacter.

  15. Molecular characterization of Staphylococcus aureus isolates causing skin and soft tissue infections (SSTIs

    Directory of Open Access Journals (Sweden)

    Zhang Xue-qing

    2010-05-01

    Full Text Available Abstract Background Staphylococcus aureus, particularly methicillin-resistant S. aureus (MRSA, is an important cause of pyogenic skin and soft tissue infections (SSTIs. The aim of present study is to investigate the molecular characteristic of Staphylococcus aureus isolates isolated from the pus samples from the patients with purulent skin and soft tissue infections in Wenzhou, China. Methods Between December 2002 and June 2008, a total of 111 nonduplicate S. aureus isolates were collected from the pus samples of the patients with SSTIs in a teaching hospital in Wenzhou, China. All the tested isolates were confirmed as S. aureus using a Staph SPA agglutination kit, Gram's stain and a Vitek-60 microbiology analyzer. The homology among the tested isolates was determined by pulsed-field gel electrophoresis (PFGE. Multilocus sequence typing (MLST was used to determine the sequence types (STs of the selected isolates. The genotypes of SCCmec were determined by a multiplex PCR in the MRSA isolates. Panton-Valentine leukocidin (PVL genes and mecA were also determined by another multiplex PCR. Results Among the 111 S. aureus isolates, 48 and 63 isolates were community-acquired and hospital-acquired respectively. Sixty isolates were confirmed as MRSA harboring mecA detected by PCR. A total of 32 PFGE clonal types were obtained by PFGE, with 10 predominant patterns (types A to J. Twenty-five different STs including ST398 and three novel STs were found among 51 selected isolates. The main STs were ST239, ST1018, ST59, ST7 and ST88. Of 60 MRSA isolates, SCCmec II, III, IV and SCCmec V were found in three, 50, three and two isolates, respectively. The positive rates of PVL genes in overall isolates, HA-isolates, CA-isolates, MRSA isolates and MSSA isolates were 23.4% (26/111, 20.6% (13/63, 27.1% (13/48, 21.7% (13/60 and 25.5% (13/51, respectively. Eight (33.3%, 8/24 of 24 CA-MRSA isolates and 5 (13.9%, 5/36 of 36 HA-MRSA isolates were positive for PVL genes

  16. Molecular identification of tsetse fly (Diptera: Glossinidae) species ...

    African Journals Online (AJOL)

    Christopher

    2015-05-13

    May 13, 2015 ... species group, palpalis or morsitans demonstrated a common ancestry and closer relatedness by .... Molecular methods. DNA extraction and PCR was done at the Molecular Biology. Laboratory of the National Veterinary Research Institute (NVRI), ..... Training Manual for Tsetse fly Control Personnel Vol.

  17. Identification of Streptococcus canis Isolated from Milk of Dairy Cows with Subclinical Mastitis

    OpenAIRE

    Hassan, Abdulwahed Ahmed; Akineden, Ömer; Usleber, Ewald

    2005-01-01

    Streptococcus canis was isolated from 31 milk samples from 11 cows in a dairy herd (with 49 lactating cows) affected by subclinical mastitis in north Rhine-Westphalia, Germany. Thirty-one isolates from the infected udder quarters were further characterized for their phenotypic and molecular properties. Most isolates (83.9%) produced α-galactosidase, and all were negative for β-d-glucuronidase. Amplification of the 16S rRNA gene by the PCR method and digestion with the restriction enzymes RsaI...

  18. Isolation and molecular genetic characterization of a yeast strain ...

    African Journals Online (AJOL)

    The yeast was identified by molecular genetics technique based on sequence analysis of the variable D1/D2 domain of the large subunit (26S) ribosomal DNA. Subsequent 26S rRNA gene sequencing showed 100% base sequence homology and it was identified as Candida viswanathii. The degradation of PAHs

  19. Molecular characterization of multidrug-resistant Klebsiella pneumoniae isolates

    Directory of Open Access Journals (Sweden)

    Xiang-hua Hou

    2015-09-01

    Full Text Available Klebsiella pneumoniae is an important cause of healthcare-associated infections worldwide. Selective pressure, the extensive use of antibiotics, and the conjugational transmission of antibiotic resistance genes across bacterial species and genera facilitate the emergence of multidrug-resistant (MDR K. pneumoniae. Here, we examined the occurrence, phenotypes and genetic features of MDR K. pneumoniae isolated from patients in intensive care units (ICUs at the First Affiliated Hospital of Xiamen University in Xiamen, China, from January to December 2011. Thirty-eight MDR K. pneumoniae strains were collected. These MDR K. pneumoniae isolates possessed at least seven antibiotic resistance determinants, which contribute to the high-level resistance of these bacteria to aminoglycosides, macrolides, quinolones and β-lactams. Among these isolates, 24 strains were extended-spectrum β-lactamase (ESBL producers, 2 strains were AmpC producers, and 12 strains were both ESBL and AmpC producers. The 38 MDR isolates also contained class I (28/38 and class II integrons (10/38. All 28 class I-positive isolates contained aacC1, aacC4, orfX, orfX’ and aadA1 genes. β-lactam resistance was conferred through blaSHV (22/38, blaTEM (10/38, and blaCTX-M (7/38. The highly conserved blaKPC-2 (37/38 and blaOXA-23(1/38 alleles were responsible for carbapenem resistance, and a gyrAsite mutation (27/38 and the plasmid-mediated qnrB gene (13/38 were responsible for quinolone resistance. Repetitive-sequence-based PCR (REP-PCR fingerprinting of these MDR strains revealed the presence of five groups and sixteen patterns. The MDR strains from unrelated groups showed different drug resistance patterns; however, some homologous strains also showed different drug resistance profiles. Therefore, REP-PCR-based analyses can provide information to evaluate the epidemic status of nosocomial infection caused by MDR K. pneumoniae; however, this test lacks the power to discriminate some

  20. Identification and characterization of chitinolytic bacteria isolated from a freshwater lake.

    Science.gov (United States)

    Tran, Dinh Minh; Sugimoto, Hayuki; Nguyen, Dzung Anh; Watanabe, Takeshi; Suzuki, Kazushi

    2018-02-01

    To develop a novel type of biocontrol agent, we focus on bacteria that are characterized by both chitinase activity and biofilm development. Chitinolytic bacteria were isolated from sediments and chitin flakes immersed in the water of a sand dune lake, Sakata, in Niigata, Japan. Thirty-one isolates from more than 5100 isolated strains were examined chitinase activity and biofilm formation. Phylogenetic analysis of these isolates based on the 16S rRNA gene sequences revealed that most isolates belonged to the family Aeromonadaceae, followed by Paenibacillaceae, Enterobacteriaceae, and Neisseriaceae. The specific activity of chitinase of four selected strains was higher than that of a reference strain. The molecular size of one chitinase produced by Andreprevotia was greater than that of typical bacterial chitinases. The dialyzed culture supernatant containing chitinases of the four strains suppressed hyphal growth of Trichoderma reesei. These results indicate that these four strains are good candidates for biocontrol agents.

  1. Isolation, Identification, and Characterization of a New Highly Pathogenic Field Isolate of Mycobacterium avium spp. avium

    Directory of Open Access Journals (Sweden)

    Liangquan Zhu

    2018-01-01

    Full Text Available Avian tuberculosis is a chronic, contagious zoonotic disease affecting birds, mammals, and humans. The disease is most often caused by Mycobacterium avium spp. avium (MAA. Strain resources are important for research on avian tuberculosis and vaccine development. However, there has been little reported about the newly identified MAA strain in recent years in China. In this study, a new strain was isolated from a fowl with symptoms of avian tuberculosis by bacterial culture. The isolated strain was identified to be MAA by culture, staining, and biochemical and genetic analysis, except for different colony morphology. The isolated strain was Ziehl-Zeelsen staining positive, resistant to p-nitrobenzoic acid, and negative for niacin production, Tween-80 hydrolysis, heat stable catalase and nitrate production. The strain had the DnaJ gene, IS1245, and IS901, as well. Serum agglutination indicated that the MAA strain was of serotype 1. The MAA strain showed strong virulence via mortality in rabbits and chickens. The prepared tuberculin of the MAA strain had similar potency compared to the MAA reference strain and standard tuberculin via a tuberculin skin test. Our studies suggested that this MAA strain tends to be a novel subtype, which might enrich the strain resource of avian tuberculosis.

  2. Isolation and identification of antifungal peptides from Bacillus BH072, a novel bacterium isolated from honey.

    Science.gov (United States)

    Zhao, Xin; Zhou, Zhi-jiang; Han, Ye; Wang, Zhan-zhong; Fan, Jie; Xiao, Hua-zhi

    2013-11-07

    A bacterial strain BH072 isolated from a honey sample showed antifungal activity against mold. Based on morphological, biochemical, physiological tests, and analysis of 16S rDNA sequence, the strain was identified to be a new subspecies of Bacillus sp. It had a broad spectrum of antifungal activity against various mold, such as Aspergillus niger, Pythium, and Botrytis cinerea. Six pairs of antifungal genes primers were designed and synthesized, and ituA, hag, tasA genes were detected by PCR analysis. The remarkable antifungal activity could be associated with the co-production of these three peptides. One of them was purified by 30-40% ammonium sulfate precipitation, Sephadex G-75 gel filtration and anion exchange chromatography on D201 resin. The purified peptide was estimated to be 35.615 kDa and identified to be flagellin by micrOTOF-Q II. By using methanol extraction, another substance was isolated from fermentation liquor, and determined to be iturin with liquid chromatography-mass spectrometry (LC-MS) method. The third possible peptide encoded by tasA was not isolated in this study. The culture liquor displayed antifungal activity in a wide pH range (5.0-9.0) and at 40-100°C. The result of the present work suggested that Bacillus BH072 might be a bio-control bacterium of research value. Copyright © 2013 Elsevier GmbH. All rights reserved.

  3. Molecular Identification of Streptomyces producing antibiotics and their antimicrobial activities

    Directory of Open Access Journals (Sweden)

    Latifa A. Al_husnan

    2016-12-01

    Full Text Available Five strains of Streptomyces, namely S, N, W, E and C (designations should be mentioned in detail here isolated from the rhizosphere soil cultivated with palm Alajua (date, pressed dates, AlMedina city, Saudi Arabia, were induced to produce antibiotics. Antimicrobial activities were determined on solid medium supplemented with starch. The detection was based on the formation of transparent zones around colonies. The results indicated that isolates had antibacterial activities against Staphylococcus aureus, Bacillus cereus, B. subtilis, Pseudomonas aeruginosa and also showed antifungal activity against Candida albicans and Aspergillus niger. DNA extracted from five isolates was used as template for 16s rDNA gene amplification. The expected PCR size was 1.5 kbp;1.6 kbp; 1.25 kbp; 1.25kbp and 1.0 k bp for S, N, W, E and C isolates respectively using universal 16s rDNA gene primers using direct PCR. The isolates varied morphologically on the basis of spore color, aerial and substrate mycelium formation, and production of diffusible pigment. Isolates were tested under a microscope by using slide culture technique. The results indicate that the soil of this region is source of Streptomyces having antibacterial and antifungal activity and thus better utilization of these microorganisms as biological control agents.

  4. Isolation and Molecular Structure of Hexacyanoruthenate(III)

    DEFF Research Database (Denmark)

    Bendix, J; Steenberg, P; Søtofte, Inger

    2003-01-01

    The [Ru(CN)(6)](3-) ion is synthesized in aqueous solution and isolated as [Ph(4)AS](3)[Ru(CN)(6)].2H(2)O (1). Compound 1 crystallizes as orange needles in the monoclinic space group P2(1)/n with cell parameters a = 11.346(2) Angstrom, b = 23.107(5) Angstrom, c = 25.015(5) Angstrom, beta = 99...

  5. Molecular Typing of Turkish Apple Chlorotic Leaf Spot Virus Isolates Based on Partial Coat Protein Gene

    Directory of Open Access Journals (Sweden)

    C. Ulubas Serce

    2006-08-01

    Full Text Available Apple chlorotic leaf spot virus (ACLSV isolates from various hosts and geographic locations in Turkey were molecularly characterized by RFLP, nucleotide sequence analysis and the construction of a phylogenetic tree including ACLSV isolates from GenBank. Based on nucleotide sequence alignment and the phylogenetic tree, we proposed a classification of ACLSV isolates in which isolates were divided into three major groups. The first group contained mainly Far-Eastern isolates, the second group the Hungarian (eastern-European ACLSV isolates, and the third group, which contained isolates of variable characteristics, was again divided into two subgroups, subgroup I containing mixed European isolates, and subgroup II containing central European isolates. Three representative Turkish ACLSV isolates belonged to the third group; of these, one was from the mixed European cluster (subgroup I and two from the central European cluster (subgroup II. The nucleotide sequence divergence and geographic origin of the ACLSV isolates were correlated, which indicated the possible extraction of the Turkish isolates.

  6. Isolation, Characterization and Identification of Environmental Bacterial Isolates with Screening for Antagonism Against Three Bacterial Targets

    Science.gov (United States)

    2017-04-01

    making the broad-spectrum antimicrobial ineffective. When this occurs, the current strategy is to replace an ineffective antimicrobial agent with...Shlae. “Fix the Antibiotic Pipeline ”. Nature 472:32. (2011) Cotter, P.A., C. Hin, and R.P. Ross. “Bacteriocin Developing Innate Immunity for Food...and resistance. Clin. Microbiol. Rev. 12:147—179. (1999) Silver, S. “Bacterial silver resistance: molecular biology and uses and misuses of silver

  7. Comparative study for 147 Candida spp. identification and echinocandins susceptibility in isolates obtained from blood cultures in 15 hospitals, Medellin, Colombia.

    Science.gov (United States)

    Berrio, Indira; Maldonado, Natalia; De Bedout, Catalina; Arango, Karen; Cano, Luz Elena; Valencia, Yorlady; Jiménez-Ortigosa, Cristina; Perlin, David S; Gómez, Beatriz L; Robledo, Carlos; Robledo, Jaime

    2017-11-26

    Invasive candidiasis has high impact on morbidity and mortality in hospitalized patients. Accurate and timely methods for identification of Candida species and determination of equinocandins susceptibility become a priority for laboratories of clinical microbiology. A study was performed to compare MALDI-TOF MS identification to sequencing of D1/D2 region of rRNA gene complex 28 subunit, in 147 isolates obtained from patients with candidemia. Susceptibility testing was performed by broth microdilution method and Etest ® . DNA sequencing of FKS1 and FKS2 genes was performed. In this study, the most common species were C. albicans (40.8%), followed by C. parapsilosis (23.1%) and C. tropicalis (17.0%). Overall agreement between the results of identification by MALDI-TOF MS and molecular identification was 99.3%. Anidulafungin and caspofungin susceptibility by broth microdilution method were 98% and 88.4%, respectively. Susceptibility to anidulafungin and caspofungin by Etest was 93.9% and 98.6%. Categorical agreement between Etest and broth microdilution method was 91.8% for anidulafungin and 89.8% for caspofungin; with lower agreements in C. parapsilosis for anidulafungin (76.5%) and C. glabrata for caspofungin (40.0%). No mutations related to resistance were found in FKS genes, although 54 isolates presented synonymous polymorphisms in the hot spots sequenced. MALDI TOF MS is a good alternative for routine identification of Candida isolates. DNA sequencing of FKS genes suggests that isolates analyzed are susceptible to echinocandins; alternatively, unknown resistance mechanisms or limitations related to antifungal susceptibility tests may explain the resistance found in few isolates. Copyright © 2017. Published by Elsevier Ltd.

  8. Detection, molecular typing and phylogenetic analysis of Leishmania isolated from cases of leishmaniasis among Syrian refugees in Lebanon

    Directory of Open Access Journals (Sweden)

    Tamara Salloum

    2016-06-01

    Two molecular typing methods of 39 FFPE Leishmania isolates were used: the ITS1-PCR RFLP and the nested ITS1-5.8S rDNA gene amplification followed by sequencing and phylogenetic analysis. The efficiency of these two techniques in Leishmania identification was compared and the phylogenetic relationships among these isolates were illustrated based on the neighbor-joining (NJ method. The results were statistically correlated with the parasitic index (PI. The DNA storage in formalin-fixed paraffin embedded (FFPE tissues was assessed as well. The parasites identified were all L. tropica as determined by both techniques. ITS1-5.8S rDNA gene based typing proved to be more sensitive in the detection of parasites (positive in 69.2% of the isolates as opposed to the ITS1-PCR RFLP method that was successful in identifying L. tropica in only 43.6% of the isolates. Sequencing and phylogenetic analysis revealed high levels of heterogeneity. A statistically significant correlation was observed between PI and the results of the nested ITS1-5.8S rDNA gene PCR. Genotyping at the species level is essential for monitoring the relative frequency of CL in the Mediterranean area that is correlated to three different Leishmania species (Leishmania infantum, Leishmania major and L. tropica, each characterized by distinct epidemiological features. The obtained results highlight the need to find a universally accepted diagnostic tool for Leishmania typing.

  9. Molecular identification of Candida dubliniensis isolated from oral lesions of HIV-positive and HIV-negative patients in São Paulo, Brazil Identificação molecular de amostras de Candida dubliniensis isoladas de lesões orais de pacientes HIV positivos e negativos em São Paulo, Brasil

    Directory of Open Access Journals (Sweden)

    Jorge Kleber Chavasco

    2006-02-01

    Full Text Available Candida dubliniensis is a new, recently described species of yeast. This emerging oral pathogen shares many phenotypic and biochemical characteristics with C. albicans, making it hard to differentiate between them, although they are genotypically distinct. In this study, PCR (Polymerase Chain Reaction was used to investigate the presence of C. dubliniensis in samples in a culture collection, which had been isolated from HIV-positive and HIV-negative patients with oral erythematous candidiasis. From a total of 37 samples previously identified as C. albicans by the classical method, two samples of C. dubliniensis (5.4% were found through the use of PCR. This study underscores the presence of C. dubliniensis, whose geographical and epidemiological distribution should be more fully investigated.Candida dubliniensis é uma nova espécie recentemente descrita. Este patógeno oral emergente compartilha muitas características fenotípicas e bioquímicas com C. albicans dificultando assim a diferenciação entre elas. As mesmas, porém, mostram-se genotipicamente distintas. Este trabalho tem como objetivo identificar, pela técnica de PCR (Polymerase Chain Reaction, a possível presença de C. dubliniensis dentre amostras isoladas de candidose oral eritematosa, provenientes de pacientes HIV positivos e HIV negativos. Em um total de 37 amostras identificadas anteriormente, por método clássico, como C. albicans encontramos duas amostras de C. dubliniensis (5,4% utilizando a técnica do PCR. Esta técnica mostrou-se útil, prática e com identificação taxonômica mais acurada.

  10. Application of PCR-based restriction fragment length polymorphism for the identification of mycobacterial isolates.

    Science.gov (United States)

    Deepa, P; Therese, K L; Madhavan, H N

    2005-05-01

    Conventional identification of mycobacteria is achieved by standard biochemical tests that are time consuming, laborious and is not always conclusive. This study was thus undertaken to standardize a simple, rapid and cost-effective polymerase chain reaction based restriction fragment length polymorphism (PCR-RFLP) using primers coding for the 16S - 23S rRNA spacer region to identify the mycobacterial isolates to the species level. The PCR with primers targeting the 16S-23S rRNA spacer region was standardized using the standard mycobacterial strains and applied on 51 clinical isolates. The PCR amplified products were subjected to RFLP using the restriction enzymes, Hae III, MspI and BstXI. The results obtained were compared with those of conventional biochemical tests. PCR was sensitive to detect 2.5 pg of H37Rv DNA (370 bp for slow grower mycobacteria) and 1.5 pg of M. fortuitum DNA (450 bp for rapid grower mycobacteria). Based on the PCRRFLP products obtained the 51 mycobacterial isolates were classified into 41 slow growers and 10 rapid growers. Among the 41 slow growers, 40 were identified as M. tuberculosis, one as M. xenopi and 10 rapid growers as M. fortuitum. PCR using primers targeting the 16S-23S rRNA spacer region was a reliable tool for rapid identification of mycobacterial isolates into slow and rapid growers within 4 h of isolation and further speciation by PCR-RFLP within 6-8 h.

  11. Molecular identification and phylogenetic analysis of human Trichostrongylus species from an endemic area of Iran.

    Science.gov (United States)

    Sharifdini, Meysam; Derakhshani, Sedigheh; Alizadeh, Safar Ali; Ghanbarzadeh, Laleh; Mirjalali, Hamed; Mobedi, Iraj; Saraei, Mehrzad

    2017-12-01

    Human infections with Trichostrongylus species have been reported in most parts of Iran. The aim of this study was the identification, molecular characterization and phylogenetic analysis of human Trichostrongylus species based on ITS2 region of ribosomal DNA from Guilan Province, northern Iran. Stool samples were collected from rural inhabitants and examined by formalin-ether concentration and agar plate culture techniques. After anthelmintic treatment, male adult worms were collected from five infected cases. Genomic DNA was extracted from one male worm of each species in every treated individual and one filariform larva isolated from each case. PCR amplification of ITS2-rDNA region was performed and the products were sequenced. Among 1508 individuals, 46 (3.05%) were found infected with Trichostrongylus species using parasitological methods. Male worms of T. colubriformis, T. vitrinus and T. longispicularis were expelled from five patients after treatment. Out of 41 filariform larvae, 40 were T. colubriformis, and the other one was T. axei. Phylogenetic analysis showed that each species was placed together with reference sequences submitted to GenBank database. Intra-species similarity for all species obtained in the current study was 100%. T. colubriformis was found to be probably the most common species in this region of Iran. For the first time, the authors of the present study report the occurrence of natural human infection by T. longispicularis in the world. Therefore, the number of Trichostrongylus species infecting human in Iran now increased to ten. Copyright © 2017. Published by Elsevier B.V.

  12. Molecular identification of endophytic fungi from Aquilaria sinensis ...

    African Journals Online (AJOL)

    Molecular phylogenetic analysis demonstrated that Botryosphaeria, Colletotrichum gloeosporioides, Phomopsis and Cylindrocladium species are members of the agarwood-producing wounded tree, while Phoma, Mycosphaerella, Sagenomella, Alternaria and Ramichloridium species is able to colonize the non-resinous ...

  13. A simple and rapid molecular method for Leptospira species identification

    NARCIS (Netherlands)

    Ahmed, Ahmed; Anthony, Richard M.; Hartskeerl, Rudy A.

    2010-01-01

    Serological and DNA-based classification systems only have little correlation. Currently serological and molecular methods for characterizing Leptospira are complex and costly restricting their world-wide distribution and use. Ligation mediated amplification combined with microarray analysis

  14. Identification of a myometrial molecular profile for dystocic labor.

    LENUS (Irish Health Repository)

    Brennan, Donal J

    2011-01-01

    The most common indication for cesarean section (CS) in nulliparous women is dystocia secondary to ineffective myometrial contractility. The aim of this study was to identify a molecular profile in myometrium associated with dystocic labor.

  15. Isolation and molecular characterization of an ethylene response ...

    Indian Academy of Sciences (India)

    2014-10-20

    Oct 20, 2014 ... Xanthi NN carrying the N gene, which is resistant to Tobacco mosaic virus (TMV). NtERF1-1 encoded a putative protein of 229 amino acids with a predicted molecular mass of 24.58 kDa. Nucleotide sequence analysis showed that NtERF1-1 contained a conserved DNA-binding domain at the N-terminal.

  16. Molecular identification of original plants of Fritillariae cirrhosae ...

    African Journals Online (AJOL)

    Background: DNA barcoding is a widely used tool that enables rapid and accurate identification of species based on standardized DNA regions. Materials and Methods: In this study, potential DNA barcodes, namely three plastid regions (rbcL, trnH-psbA and matK) and one nuclear ribosomal internal transcribed spacer (ITS) ...

  17. Seasonal abundance and molecular identification of West Nile virus ...

    African Journals Online (AJOL)

    tion of the Ace2 gene by PCR confirmed morphormetric identification of the mosquitoes. Results: A total of 751 mosquitoes were ... WestNile virus, (WNV), Yellow fever virus. (YFV), Dengue virus (DNV) and Rift valley fever virus ..... West Nile virus in mosquitoes and febrile patients in a semi-arid zone in Nigeria. J Am Sci.

  18. Prevalence, incidence and molecular identification of root-knot ...

    African Journals Online (AJOL)

    Tomato is a widely grown vegetable in Pakistan. However, its production is severely constrained by root knot nematodes (RKNs). Accurate identification of RKNs is essential for an appropriate control program. The current study evaluated the prevalence, incidence and diversity of RKNs of tomato crops grown in the Khyber ...

  19. ISOLATION AND MOLECULAR CHARACTERIZATION OF EGYPTIAN TRICHODERMA AND ASSESSMENT OF THEIR ANTAGONISTIC POTENTIAL AGAINST RHIZOCTONIA SOLANI

    Directory of Open Access Journals (Sweden)

    Gamal Mohamedin Hassan

    2015-06-01

    Full Text Available Morphological and molecular characterization of antagonistic ability of Trichoderma species was studied. Soil dilution plate method was used to isolate trichoderma from rhizosphere of bean, cowpea, cucumber, wheat and faba bean plants. Based on morphological and cultural characteristics, the Trichoderma isolates were identified as T. harzianum (10 isolates, T. koningii (8 isolates, and T. viride (2 isolates. A portion of rDNA, 560-600 bp was amplified from six biocontrol isolates using ITS1 and ITS 4 primers, and was sequenced and aligned against ex-type strain sequences from TrichoBlast and established Trichoderma taxonomy. Molecular phylogenetic analysis were performed based on nucleotide sequences in order to examine these isolates among 15 accession numbers of Trichoderma spp. found in GenBank. The results indicate that the FUE3, FUE5, FUE6, FUE9 and FUE18 Trichoderma isolates are closely related to Trichoderma koningii, while FUE15 isolate is closely related to Trichoderma harzianim .This result was in accordance with the result obtained from morphological and cultural characteristics. Production of volatile inhibitors and mycoparasitism were investigated using in vitro and in vivo tests in dual culture PDA medium and infected soils. The percent inhibitory effect against growth of Rhizoctonia solani was calculated, T. koningii FUE3 showed the greatest antagonistic effect to the pathogen (57.77% in vitro experiment whereas T. koningii FUE6 and FUE18 were gave the highest reduction 96% of disease incidence caused by R. solani in greenhouse conditions.

  20. Isolation and molecular characterization of a new Neospora caninum isolate from cattle in Argentina.

    Science.gov (United States)

    Campero, L M; Venturini, M C; Moore, D P; Massola, L; Lagomarsino, H; García, B; Bacigalupe, D; Rambeaud, M; Pardini, L; Leunda, M R; Schares, G; Campero, C M

    2015-08-01

    Neospora caninum is one of the most important causes of bovine abortion, but isolation of live parasites from infected tissue is difficult. The aims of the present study were to obtain new isolates of N. caninum from congenitally infected asymptomatic newborn cattle in Argentina and to perform characterization by multilocus-microsatellite analysis. Five clinically normal born calves, with demonstrable N. caninum antibodies in precolostrum serum by indirect fluorescent antibody test, were euthanized and their brain samples were processed for histopathological, immunohistochemical, polymerase chain reaction (PCR) analysis, and for bioassay in γ-interferon knockout (GKO) mice. Although N. caninum DNA was detected in brain from all the calves by PCR, viable N. caninum was isolated in GKO mice from only one calf. Neospora caninum tachyzoites of this Argentinean isolate, designated NC-Argentina LP1, were propagated in VERO cell cultures seeded with tachyzoites from the infected GKO mice tissues. Multilocus-microsatellite typing on DNA derived from cell cultured tachyzoites revealed a unique genetic pattern, different from reported isolates. This is the first bovine isolation and genetic characterization of N. caninum in Argentina. Copyright © 2015 Elsevier Inc. All rights reserved.