scholarly journals Investigation of virulence factors of Enterococcus faecalis strains isolated in secondary/ persistent infections

2014 ◽  
Vol 17 (1) ◽  
pp. 32
Author(s):  
Ana Claudia C. Xavier ◽  
Frederico Canato Martinho ◽  
Izabel C.G. Camões ◽  
Lilian F. Freitas
Keyword(s):  
Enterococcus Faecalis ◽  
Virulence Factor ◽  
Intergenic Spacer ◽  
Virulence Determinants ◽  
Test Species ◽  
Intergenic Spacer Region ◽  
Culture Techniques ◽  
Endodontic Infection ◽  
Species Specific ◽  
Virulence Factor Genes

<p><strong>Objective</strong>: More virulent strains may result from  the acquisition of genes by genetic exchange,  pathogenicity islands in several species encoding  toxins, adhesion factors and other factors associated  with virulence. The aim of this study was to investigate  the prevalence of E. faecalis strains in secondary  endodontic/ persistent using endodontic infection by  culture and PCR technqiues; and to investigate for  the presence of virulence factor genes of gelatinase  (gelE), cytolysin activator (Cyla), surface adhesin  of Enterococcus (ESP) and collagen adhesin of  Enterococcus (ACE). <strong>Material and methods</strong>: Microbial samples were obtained from 12 teeth with  secondary/ persistent endodontic infection showing  apical periodontitis. Culture techniques were used  including serial dilution, plating, incubation, and  biochemical identification. For PCR detection, samples  were analyzed using a species-specific primer of the  16S rDNA and the downstream intergenic spacer  region. <strong>Results</strong>: Culture and PCR detected the test  species in 3/12 (25%) and 5/12 (41.6%) of teeth,  respectively. A total of 38 Enterococcus faecalis strains  were isolated and submitted to the virulence factor  genes analysis. PCR products consistent with genes  encoding surface adhesion (ESP), gelatinase (gelE)  and collagen binding antigen (ACE) were found in  26/38 (68%), 31/38 (81%) and 38/38 (100%) of  the isolates. The Cytolysin activator (Cyla) gene was  not recovered from E. faecalis isolates. <strong>Conclusions</strong>:  In conclusion, the present study revealed by culture  and molecular methods revealed a high prevalence  of E. faecalis in teeth with secondary/ persistent  endodontic infection. Moreover, of a clinical  relevance, we found different E. faecalis strains  carrying different virulence determinants.</p><p><strong>KEYWORDS</strong> Bacteria; E. faecalis; Root canal, virulence.    </p>

scholarly journals Evaluation of Nonpathogenic Fusarium oxysporum and Pseudomonas fluorescens for Panama Disease Control

Plant Disease ◽  
2011 ◽  
Vol 95 (8) ◽  
pp. 951-959 ◽  
Author(s):  
A. Belgrove ◽  
C. Steinberg ◽  
A. Viljoen
Keyword(s):  
Fusarium Oxysporum ◽  
Pseudomonas Fluorescens ◽  
Fusarium Wilt ◽  
Intergenic Spacer ◽  
Polymorphism Analysis ◽  
Panama Disease ◽  
Intergenic Spacer Region ◽  
Soil Microbial ◽  
Fusarium Wilt Of Banana ◽  
Species Specific

Nonpathogenic Fusarium oxysporum endophytes from healthy banana roots were evaluated for their ability to reduce Fusarium wilt of banana (Panama disease). Isolates were identified morphologically and by using species-specific primers. Pathogenicity was confirmed by inoculating banana plantlets in the greenhouse. Nonpathogenic F. oxysporum isolates were grouped into 14 haplotype groups by polymerase chain reaction restriction fragment length polymorphism analysis of the intergenic spacer region, and representative isolates evaluated for biocontrol of F. oxysporum f. sp. cubense. In the greenhouse, 10 nonpathogenic F. oxysporum isolates were able to significantly reduce Fusarium wilt of banana. The isolate that protected banana plantlets best in the greenhouse, a nonpathogenic F. oxysporum from the root rhizosphere, and Pseudomonas fluorescens WCS 417 were then field tested. When the putative biological control organisms were tested in the field, neither the nonpathogenic F. oxysporum, P. fluorescens, nor combinations thereof reduced Fusarium wilt development significantly. A number of factors could contribute to the lack of field protection, including soil microbial and chemical composition and reduced survival of biocontrol organisms in banana roots. A lack of knowledge regarding the etiology of Fusarium wilt of ‘Cavendish’ banana in the subtropics and the effect of F. oxysporum f. sp. cubense race and banana cultivar in protection of banana by biocontrol organisms should be further investigated.

scholarly journals Identification of Porphyromonas gingivalis Strains by Heteroduplex Analysis and Detection of Multiple Strains

1999 ◽  
Vol 37 (12) ◽  
pp. 3906-3911 ◽  
Author(s):  
Eugene J. Leys ◽  
James H. Smith ◽  
Sharon R. Lyons ◽  
Ann L. Griffen
Keyword(s):  
Porphyromonas Gingivalis ◽  
Dna Sequences ◽  
Allelic Variation ◽  
Intergenic Spacer ◽  
Heteroduplex Analysis ◽  
Clinical Samples ◽  
Specific Primers ◽  
Intergenic Spacer Region ◽  
Species Specific ◽  
Multiple Strains

Heteroduplex analysis has been used extensively to identify allelic variation among mammalian genes. It provides a rapid and reliable method for determining and cataloging minor differences between two closely related DNA sequences. We have adapted this technique to distinguish among strains or clonal types of Porphyromonas gingivalis. The ribosomal intergenic spacer region (ISR) was amplified directly from a subgingival plaque sample by PCR with species-specific primers, avoiding the need for culturing the bacteria. The PCR products were then directly compared by heteroduplex analysis with known strains of P. gingivalis for identification. We identified 22 distinct but closely related heteroduplex types ofP. gingivalis in 1,183 clinical samples. Multiple strains were found in 34% of the samples in which P. gingivaliswas detected. Heteroduplex types were identified from these multistrain samples without separating them by culturing or molecular cloning. PCR with species-specific primers and heteroduplex analysis makes it possible to reliably and sensitively detect and identify strains ofP. gingivalis in large numbers of samples.

PCR Detection Assay of Fumonisin-Producing Fusarium verticillioides Strains

2004 ◽  
Vol 67 (6) ◽  
pp. 1278-1283 ◽  
Author(s):  
BELÉN PATIÑO ◽  
SALVADOR MIRETE ◽  
M. TERESA GONZÁLEZ-JAÉN ◽  
GIUSEPPINA MULÉ ◽  
M. TERESA RODRÍGUEZ ◽  
...  
Keyword(s):  
Fusarium Species ◽  
Fusarium Verticillioides ◽  
Animal Health ◽  
Intergenic Spacer ◽  
Fungal Species ◽  
Intergenic Spacer Region ◽  
Detection And Identification ◽  
Geographical Regions ◽  
Species Specific ◽  
A Minor

Fusarium verticillioides is considered to be the main source of fumonisins, a group of toxins that contaminate commodities and result in chronic and acute diseases affecting humans and animals. The detection and control of this species is crucial to prevent fumonisins from entering the food chain. The objective of the present research was to develop a specific, sensitive, and robust PCR assay to detect F. verticillioides strains using two pairs of specific primers for F. verticillioides, which have been designed on the basis of the intergenic spacer region of the rDNA units. The first pair of primers was F. verticillioides species specific, whereas the second pair of primers detected fumonisin-producing F. verticillioides strains. This second pair of primers allowed for the discrimination between the major group of F. verticillioides strains, fumonisin-producing strains that are mainly associated with crops, and a minor group of strains, non–fumonisin-producing strains that are associated with bananas. Fifty-four strains of F. verticillioides from different geographical regions and hosts were tested using both sets of primers. Sixteen additional Fusarium species were examined. The specificity of the primer sequences provides the basis for a simple, rapid, accurate, and sensitive detection and identification method of this fungal species that represents a risk for human and animal health.

scholarly journals Simultaneous Detection and Identification of Common Cell Culture Contaminant and Pathogenic Mollicutes Strains by Reverse Line Blot Hybridization

2004 ◽  
Vol 70 (3) ◽  
pp. 1483-1486 ◽  
Author(s):  
Hui Wang ◽  
Fanrong Kong ◽  
Peter Jelfs ◽  
Gregory James ◽  
Gwendolyn L. Gilbert
Keyword(s):  
Cell Culture ◽  
16S Rrna ◽  
Cell Line ◽  
Intergenic Spacer ◽  
23S Rrna ◽  
Pcr Assay ◽  
Intergenic Spacer Region ◽  
Specific Pcr ◽  
Specific Pcr Assay ◽  
Species Specific

ABSTRACT We have developed a reverse line blot (RLB) hybridization assay to detect and identify the commonest mollicutes causing cell line contamination (Mycoplasma arginini, Mycoplasma fermentans, Mycoplasma hyorhinis, Mycoplasma orale, and Acholeplasma laidlawii) and human infection (Mycoplasma pneumoniae, Mycoplasma hominis, Mycoplasma genitalium, Ureaplasma parvum, and Ureaplasma urealyticum). We developed a nested PCR assay with “universal” primers targeting the mollicute 16S-23S rRNA intergenic spacer region. Amplified biotin-labeled PCR products were hybridized to membrane-bound species-specific oligonucleotide probes. The assay correctly identified reference strains of 10 mollicute species. Cell cultures submitted for detection of mollicute contamination, clinical specimens, and clinical isolates were initially tested by PCR assay targeting a presumed mollicute-specific sequence of the 16S rRNA gene. Any that were positive were assessed by the RLB assay, with species-specific PCR assay as the reference method. Initially, 100 clinical and 88 of 92 cell culture specimens gave concordant results, including 18 in which two or more mollicute species were detected by both methods. PCR and sequencing of the 16S-23S rRNA intergenic spacer region and subsequent retesting by species-specific PCR assay of the four cell culture specimens for which results were initially discrepant confirmed the original RLB results. Sequencing of amplicons from 12 cell culture specimens that were positive in the 16S rRNA PCR assay but negative by both the RLB and species-specific PCR assays failed to identify any mollicute species. The RLB hybridization assay is sensitive and specific and able to rapidly detect and identify mollicute species from clinical and cell line specimens.

scholarly journals Prevalence and Characteristics of Streptococcus canis Strains Isolated from Dogs and Cats

2007 ◽  
Vol 76 (4) ◽  
pp. 619-625 ◽  
Author(s):  
P. Lysková ◽  
M. Vydržalová ◽  
D. Královcová ◽  
J. Mazurová
Keyword(s):  
Antimicrobial Agent ◽  
Intergenic Spacer ◽  
Penicillin G ◽  
23S Rrna ◽  
Phenotypic Properties ◽  
Intergenic Spacer Region ◽  
Arginine Dihydrolase ◽  
Specific Amplification ◽  
Streptococcus Canis ◽  
Species Specific

To determine the prevalence of Streptococcus canis in dogs and cats, a total of 926 swabs were examined bacteriologically in the period from 2003 to 2005. Eighty-six isolates obtained from various anatomical locations were further characterized for their phenotypic properties. The most frequently isolated biotype produced phosphatase, leucine amidopeptidase, arginine dihydrolase, alpha-D- and beta-D-galactosidase and fermented lactose and ribose. Additional identification by species-specific amplification of the 16S-23S rRNA intergenic spacer region was consistent with S. canis. All isolates were susceptible to penicillin G and ampicillin. The least effective antimicrobial agent was found to be tetracycline (only 33.8% of susceptible strains).

scholarly journals Identification of Nile Perch (Lates niloticus), Grouper (Epinephelus guaza), and Wreck Fish (Polyprion americanus) Fillets by PCR Amplification of the 5S rDNA Gene

2001 ◽  
Vol 84 (3) ◽  
pp. 777-781 ◽  
Author(s):  
Luis Asensio ◽  
Isabel González ◽  
Alicia Fernández ◽  
Ana Céspedes ◽  
Miguel A Rodríguez ◽  
...  
Keyword(s):  
Pcr Amplification ◽  
Intergenic Spacer ◽  
5S Rdna ◽  
Nile Perch ◽  
Intergenic Spacer Region ◽  
Rdna Gene ◽  
Lates Niloticus ◽  
Polymerase Chain ◽  
Species Specific ◽  
Polyprion Americanus

Abstract Nile perch (Lates niloticus), grouper (Epinephelus guaza), and wreck fish (Polyprion americanus) were differentiated by polymerase chain reaction (PCR) amplification of the nuclear 5S rDNA gene. The design of 3 species-specific primers complementary to the nontranscribed intergenic spacer region from the 5S rDNA molecule allowed amplification of clearly distinguishable gene fragments in each fish species. This approach is rapid and reliable and offers the potential to detect fraudulent or unintentional mislabeling of these species in routine seafood authentication analysis.

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