Conventional and molecular typing of Salmonella typhi strains from Brazil

Abstract

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.

Optimization of randomly amplified polymorphic DNA-polymerase chain reaction for molecular typing of Salmonella enterica serovar Typhi

“…Thus, the use of random amplification of polymorphic DNA fingerprinting technique (RAPD), a modification of the polymerase chain The number and the intensity of fragments produced varied in function of the thermal cycling profile, the MgCl 2 , DNA template, primer and Taq DNA polymerase enzyme concentrations. Maintaining the DNA and primer 784 concentrations at 20ng/µl and 20pM/µl, respectively, and varying the MgCl 2 concentration, we observed that the best fingerprinting pattern was obtained using 2.0mM MgCl 2 at reaction (PCR) in which arbitrary oligonucleotides primers are used to promote DNA synthesis at low stringency conditions in order to determine genomic diversity, may be considered a promising alternative typing method capable of discriminating between S. enterica serovar Typhi strains of the same phage type 14 . Besides differentiating isolates of serovar Typhi and other Salmonella isolates, RAPD-PCR proved capable of discriminating between S. enterica serovar Typhi strains 16 .…”

“…As described previously 14 . It has been demonstrated that RAPD-PCR reaction has the potential to provide a discriminatory, reproducible and easy to interpret method to type S. enterica serovar Typhi strains.…”

Molecular typing of Salmonella from Sergipe, Northeastern Brazil, showing the epidemiological relationship between poultry and human infection

“…PCR-based fingerprinting using RAPD has been used widely for genome identification, and the RAPD technique using up to six primers has been reported to be a good tool for molecular typing, permitting the execution of public health measures necessary to control infection (Chansiripornchai et al, 2000;De Cesare et al, 2001;Khoodoo et al, 2002;Quintaes et al, 2002). During this study, we found that the best amplification conditions for RAPD-PCR were an annealing temperature of 35ºC with 3 µL primer.…”

“…It does not require any previous knowledge of the DNA sequence of the target organisms, and results in simpler, low-cost, and rapid detection of the genetic variations present in isolated samples (Colauto et al, 2002;Ruiz et al, 2003;Santos and Bäumler, 2004). RAPD has been used as a tool to trace Salmonella epidemiologically, to distinguish Salmonella strains from different geographical origins, and for complementing serotyping and phage typing methods (Soto et al, 1999;De Cesare et al, 2001;Quintaes et al, 2002;Lim et al, 2005;Smith et al, 2011;Rezk et al, 2012).…”

Variabilidade genética de amostras de Salmonella Typhi isoladas de surto e de casos esporádicos ocorridos em Belém, Brasil

“…Uma nítida e peculiar distribuição sazonal é verificada no estado do Pará, já que mais de 70% dos casos da doença são registrados durante o período de intenso calor, entre os meses de julho e dezembro (13,14) .…”