scholarly journals Conventional and molecular typing of Salmonella typhi strains from Brazil

2002 ◽  
Vol 44 (6) ◽  
pp. 315-319 ◽  
Author(s):  
Bianca R. QUINTAES ◽  
Nilma C. LEAL ◽  
Eliane M. F. REIS ◽  
Érica L. FONSECA ◽  
Ernesto HOFER
Keyword(s):  
Molecular Typing ◽  
Random Amplified Polymorphic Dna ◽  
Salmonella Typhi ◽  
Discriminatory Power ◽  
Pcr Analysis ◽  
Biochemical Tests ◽  
Promising Alternative ◽  
Phage Types ◽  
Rapd Pcr ◽  
Typing Methods

Phenotypic and genotypic characteristics of Salmonella Typhi were studied in 30 strains, isolated in different years, from some areas in Brazil. Conventional typing methods were performed by biochemical tests, Vi phage-typing scheme, and antimicrobial susceptibility test. Molecular typing methods were performed by analysis of plasmid DNA and by random amplified polymorphic DNA (RAPD-PCR). For the latter, an optimization step was performed to ensure the reproducibility of the process in genetic characterization of S. Typhi. The predominance of 76.7% of biotype I (xylose +, arabinose -) was noticed in all studied areas. Three phage types were recognized, with prominence for the phage types A (73.3%) and I+IV (23.3%). All the strains were susceptible to the drugs used. However, 36.7% of the strains contained plasmids, with predominance of the 105 Kb plasmid. RAPD was capable of grouping the strains in 8 genotypic patterns using primer 784, in 6, using primer 787 and in 7, using primer 797. Conventional phenotypic typing methods, as well as the DNA plasmid analysis, presented nonsignificant discriminatory power; however, RAPD-PCR analysis showed discriminatory power, reproducibility, easy interpretation and performance, being considered as a promising alternative typing method for S. Typhi.

scholarly journals Conventional and Molecular Typing of Salmonella enterica serotype Typhi Locally Isolated In Baghdad

2012 ◽  
Vol 9 (4) ◽  
pp. 632-639
Author(s):  
Baghdad Science Journal
Keyword(s):  
Molecular Typing ◽  
Salmonella Enterica ◽  
Random Amplified Polymorphic Dna ◽  
Discriminatory Power ◽  
Universal Primers ◽  
Pcr Analysis ◽  
Biochemical Tests ◽  
Promising Alternative ◽  
Rapd Pcr ◽  
Typing Methods

Phenotypic And genotypic characteristics of Salmonella enterica serotype Typhi have been determined for 29 isolates, from Baghdad in 2007. Conventional typing methods were performed by biochemical tests, and antimicrobial susceptibility test. Molecular typing performed by analysis plasmid DNA beside using the Random Amplified Polymorphic DNA (RAPD-PCR). For the latter, two universal primers that have selected for the high discriminatory power were used for RAPD analysis. All isolates were belong one biotype according to the differention by their ability to decarboxylat lysine, 29(100%) were lysine (+). All the isolates were susceptible to the Antibiotics used. However, all the strains free of plasmids. RAPD was capable of grouping the strains in 6 genotypic patterns using primer 784, in 4 genotypic patterns using primer 787. Conventional phenotypic typing methods, as well as the DNA plasmid analysis, presented non significant discriminatory power; however, RAPD-PCR analysis showed discriminatory power, reproducibility, easy interpretation and can be considered as a promising alternative typing method for S. Typhi.

scholarly journals Characterization of verocytotoxin-producingEscherichia coliO157 isolates from patients with haemolytic uraemic syndrome in Western Europe

1995 ◽  
Vol 115 (1) ◽  
pp. 1-3 ◽  
Author(s):  
A. E. Heuvelink ◽  
N. C. A. J. van de Kar ◽  
J. F. G. M. Meis ◽  
L. A. H. Monnens ◽  
W. J. G. Melchers
Keyword(s):  
Haemolytic Uraemic Syndrome ◽  
Western Europe ◽  
Random Amplified Polymorphic Dna ◽  
E Coli ◽  
Pcr Fingerprinting ◽  
Phage Types ◽  
Rapd Pcr ◽  
Typing Methods ◽  
Dna Pcr

SummaryFifty verocytotoxin (VT)-producingEscherichia coli(VTEC) strains of serogroup O157 were characterized by phage typing, polymerase chain reaction (PCR) for VT genes and theE. coliattaching and effacing (eae) gene, and random amplified polymorphic DNA–PCR (RAPD–PCR) fingerprinting. The collection represented isolates obtained from patients with diarrhoea-associated haemolytic-uraemic syndrome (D+ HUS) and their family contacts, isolated in the Netherlands, Belgium and Germany between 1989 and 1993. Based on isolates from separate families (n= 27) seven different phage types were identified, types 2 (44%) and 4 (33%) were predominant. Eighty-five percent of the strains contained only VT2 gene sequences and 15% both VT1 and VT2. All strains of the dominant phage types 2 and 4 carried the VT2 gene. Strains that belonged to the minor phage types 8, 14, 32 carried both VT1 and VT2 genes, with the exception of two isolates identified as phage types 49 and 54 which contained only VT2 genes. All O157 VTEC strains possessed the chromosomally-locatedeaegene, which indicates its usefulness as virulence marker. RAPD–PCR fingerprinting identified four distinct banding patterns, with one profile found among 79% of the strains. Based on the combined results of all typing methods used in this study, the collection of 50 O157 VTEC strains could be divided into nine distinct groups. Strains isolated from different persons within one family could not be distinguished by any of these methods. The data suggest that O157 VTEC strains are members of one clone that has become widely distributed.

scholarly journals Genetic Diversity of Pierce's Disease Strains and Other Pathotypes of Xylella fastidiosa

2001 ◽  
Vol 67 (2) ◽  
pp. 895-903 ◽  
Author(s):  
Mavis Hendson ◽  
Alexander H. Purcell ◽  
Deqiao Chen ◽  
Chris Smart ◽  
Magalie Guilhabert ◽  
...  
Keyword(s):  
Gel Electrophoresis ◽  
Xylella Fastidiosa ◽  
Random Amplified Polymorphic Dna ◽  
Consensus Sequence ◽  
23S Rrna ◽  
Pierce's Disease ◽  
Pcr Analysis ◽  
Leaf Scorch ◽  
Pierce’S Disease ◽  
Rapd Pcr

ABSTRACT Strains of Xylella fastidiosa isolated from grape, almond, maple, and oleander were characterized by enterobacterial repetitive intergenic consensus sequence-, repetitive extragenic palindromic element (REP)-, and random amplified polymorphic DNA (RAPD)-PCR; contour-clamped homogeneous electric field (CHEF) gel electrophoresis; plasmid content; and sequencing of the 16S-23S rRNA spacer region. Combining methods gave greater resolution of strain groupings than any single method. Strains isolated from grape with Pierce's disease (PD) from California, Florida, and Georgia showed greater than previously reported genetic variability, including plasmid contents, but formed a cluster based on analysis of RAPD-PCR products,NotI and SpeI genomic DNA fingerprints, and 16S-23S rRNA spacer region sequence. Two groupings of almond leaf scorch (ALS) strains were distinguished by RAPD-PCR and CHEF gel electrophoresis, but some ALS isolates were clustered within the PD group. RAPD-PCR, CHEF gel electrophoresis, and 16S-23S rRNA sequence analysis produced the same groupings of strains, with RAPD-PCR resolving the greatest genetic differences. Oleander strains, phony peach disease (PP), and oak leaf scorch (OLS) strains were distinct from other strains. DNA profiles constructed by REP-PCR analysis were the same or very similar among all grape strains and most almond strains but different among some almond strains and all other strains tested. Eight of 12 ALS strains and 4 of 14 PD strains of X. fastidiosa isolated in California contained plasmids. All oleander strains carried the same-sized plasmid; all OLS strains carried the same-sized plasmid. A plum leaf scald strain contained three plasmids, two of which were the same sizes as those found in PP strains. These findings support a division of X. fastidiosaat the subspecies or pathovar level.

Characterization and epidemiological relationships of SpanishBrachyspira hyodysenteriaefield isolates

2009 ◽  
Vol 138 (1) ◽  
pp. 76-85 ◽  
Author(s):  
Á. HIDALGO ◽  
A. CARVAJAL ◽  
M. PRINGLE ◽  
P. RUBIO ◽  
C. FELLSTRÖM
Keyword(s):  
Phenotypic Diversity ◽  
Random Amplified Polymorphic Dna ◽  
Pulsed Field Gel Electrophoresis ◽  
Pcr Analysis ◽  
Biochemical Tests ◽  
Diagnostic Pcr ◽  
Brachyspira Hyodysenteriae ◽  
Antimicrobial Susceptibility Tests ◽  
Susceptibility Tests ◽  
First Time

SUMMARYThis research aimed to describe the genetic and phenotypic diversity of 74 SpanishBrachyspira hyodysenteriaefield isolates, to establish epidemiological relationships between the isolates and to confirm the presence of tiamulin-resistant isolates in Spain. For these purposes, we performed biochemical tests in combination with diagnostic PCR analysis for the identification ofBrachyspiraspp. and for detection of thesmpA/smpBgene. We also used antimicrobial susceptibility tests, random amplified polymorphic DNA (RAPD) and a new pulsed-field gel electrophoresis (PFGE) protocol. The combination of RAPD and PFGE allowed the study of epidemiological relationships. Both indole-negative and tiamulin-resistant isolates ofB. hyodysenteriaeare reported in Spain for the first time. The genetic analyses indicated a relationship between these Spanish isolates and indole-negative isolates previously obtained from Germany and Belgium.

Characterization of two non-native invasive bark beetles, Scolytus schevyrewi and Scolytus multistriatus (Coleoptera: Curculionidae: Scolytinae)

2008 ◽  
Vol 140 (5) ◽  
pp. 527-538 ◽  
Author(s):  
Patricia L. Johnson ◽  
Jane L. Hayes ◽  
John Rinehart ◽  
Walter S. Sheppard ◽  
Steven E. Smith
Keyword(s):  
Bark Beetle ◽  
Bark Beetles ◽  
Developmental Stages ◽  
Random Amplified Polymorphic Dna ◽  
Morphological Characters ◽  
Pcr Analysis ◽  
Scolytus Multistriatus ◽  
Blind Test ◽  
Rapd Pcr ◽  
Elm Bark Beetle

AbstractScolytus schevyrewi Semenov, the banded elm bark beetle, and S. multistriatus Marsham, the smaller European elm bark beetle, are morphologically similar. Reliance on adult external morphological characters for identification can be problematic because of wide within-species variability and the need for good-quality specimens. The inability to identify developmental stages can also hamper early-detection programs. Using two character identification systems, genitalic (aedeagus) morphology, and DNA markers (random amplified polymorphic DNA polymerase chain reaction (RAPD-PCR)) to distinguish S. schevyrewi from S. multistriatus, we examined specimens from geographically distinct populations of both species collected from infested host trees or semiochemical-baited funnel traps. We found that aedeagus morphology can be used to identify the two species. The use of two oligonucleotide primers in the RAPD-PCR analysis yielded distinct DNA banding patterns for the two species. Species identification using RAPD-PCR analysis was validated by a blind test and used to make species identifications of larval specimens. These tools improve the ability to differentiate between S. schevyrewi and S. multistriatus at immature and adult stages, and could be developed and used for other scolytines as well.

Development of an Efficient Fungal DNA Extraction Method To Be Used in Random Amplified Polymorphic DNA–PCR Analysis To Differentiate Cyclopiazonic Acid Mold Producers

2008 ◽  
Vol 71 (12) ◽  
pp. 2497-2503 ◽  
Author(s):  
BEATRIZ SÁNCHEZ ◽  
MAR RODRÍGUEZ ◽  
EVA M. CASADO ◽  
ALBERTO MARTÍN ◽  
JUAN J. CÓRDOBA
Keyword(s):  
Dna Extraction ◽  
Extraction Method ◽  
Rapd Analysis ◽  
Random Amplified Polymorphic Dna ◽  
Cyclopiazonic Acid ◽  
Proteinase K ◽  
Vacuum System ◽  
Pcr Analysis ◽  
Dna Extraction Method ◽  
Rapd Pcr

A variety of previously established mechanical and chemical treatments to achieve fungal cell lysis combined with a semiautomatic system operated by a vacuum pump were tested to obtain DNA extract to be directly used in randomly amplified polymorphic DNA (RAPD)–PCR to differentiate cyclopiazonic acid–producing and –nonproducing mold strains. A DNA extraction method that includes digestion with proteinase K and lyticase prior to using a mortar and pestle grinding and a semiautomatic vacuum system yielded DNA of high quality in all the fungal strains and species tested, at concentrations ranging from 17 to 89 ng/μl in 150 μl of the final DNA extract. Two microliters of DNA extracted with this method was directly used for RAPD-PCR using primer (GACA)4. Reproducible RAPD fingerprints showing high differences between producer and nonproducer strains were observed. These differences in the RAPD patterns did not differentiate all the strains tested in clusters by cyclopiazonic acid production but may be very useful to distinguish cyclopiazonic acid producer strains from nonproducer strains by a simple RAPD analysis. Thus, the DNA extracts obtained could be used directly without previous purification and quantification for RAPD analysis to differentiate cyclopiazonic acid producer from nonproducer mold strains. This combined analysis could be adaptable to other toxigenic fungal species to enable differentiation of toxigenic and nontoxigenic molds, a procedure of great interest in food safety.

scholarly journals Molecular Typing of Hospital-Acquired Staphylococcus aureus Isolated from Isfahan, Iran

2014 ◽  
Vol 2014 ◽  
pp. 1-6 ◽  
Author(s):  
Seyed Asghar Havaei ◽  
Fahimeh Ghanbari ◽  
Ali Asghar Rastegari ◽  
Amir Azimian ◽  
Farzad Khademi ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Molecular Typing ◽  
Diffusion Method ◽  
Biochemical Tests ◽  
Spa Typing ◽  
Disk Diffusion Method ◽  
Type Iv ◽  
Typing Methods ◽  
Hospital Acquired ◽  
Spa Type

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.

scholarly journals VARIASI GENETIK Lactobacillus fermentum Beijerink ASAL SAYUR ASIN BERDASARKAN ANALISIS RFLP 16S-23S rDNA ISR, RAPD-PCR DAN ERIC-PCR

2017 ◽  
Vol 16 (2) ◽  
Author(s):  
Sulistiani Sulistiani ◽  
Wibowo Mangunwardoyo ◽  
Abinawanto Abinawanto ◽  
Endang Sukara ◽  
Achmad Dinoto ◽  
...  
Keyword(s):  
Genetic Variation ◽  
Random Amplified Polymorphic Dna ◽  
Intergenic Spacer ◽  
Molecular Techniques ◽  
Lactobacillus Fermentum ◽  
Pcr Analysis ◽  
Intergenic Spacer Region ◽  
Rapd Pcr ◽  
Upgma Cluster Analysis ◽  
Process Combination

Molecular analysis of Lactobacillus fermentum isolates is essential to understand their genetic variation in relations to their roles in sayur asin fermentation process. Combination of three molecular techniques which is restriction fragment length polymorphism (RFLP) of 16S23S rDNA intergenic spacer region (ISR), random amplified polymorphic DNA (RAPD-PCR) and an enterobacterial repetitive intergenic consensus (ERIC-PCR) analysis were performed to discriminate 19 representative isolates of L. fermentum isolated from sayur asin. The result showed that L. fermentum strain D11 is distantly related to other isolates based on RFLP using HhaI restriction enzyme and RAPDPCR analyses. In addition, both of RAPD-PCR and ERIC-PCR successfully determined the genetic variation among L. fermentum strains by exhibiting distinct 4-8 bands (800-2080 bp) and 4-10 bands (280-3050 bp), respectively. A dendogram generated from UPGMA cluster analysis of both RAPD-PCR and ERIC-PCR data showed two distinct genotypic groups exist among L. fermentum isolated from sayur asin in Indonesia.

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