Evaluation of a Multilocus Variable-Number Tandem-Repeat Analysis Scheme for Typing Human Brucella Isolates in a Region of Brucellosis Endemicity▿ †

Brucellosis remains an important anthropozoonosis worldwide. Brucella species are genetically homogeneous, and thus, the typing of Brucella species for epidemiological purposes by conventional molecular typing methods has remained elusive. Although many methods could segregate isolates into the phylogenetically recognized taxa, limited within-species genetic diversity has been identified. Recently, multilocus variable-number tandem-repeat analysis (MLVA) was found to have a high degree of resolution when it was applied to collections of Brucella isolates from geographically widespread locations, and an assay comprising 16 such loci (MLVA-16) was proposed. This scheme includes eight minisatellite loci (panel 1) and eight microsatellites (panel 2, which is subdivided into panels 2A and 2B). The utility of MLVA-16 for the subtyping of human Brucella isolates from geographically restricted regions needs to be further evaluated, and genotyping databases with worldwide coverage must be progressively established. In the present study, MLVA-16 was applied to the typing of 42 human Brucella isolates obtained from 41 patients recovered from 2002 to 2006 at a tertiary-care center in Lebanon. All isolates were identified as Brucella melitensis by MLVA-16 and were found to be closely related to B. melitensis isolates from neighboring countries in the Middle East when their genotypes were queried against those in the web-based Brucella2007 MLVA database (http://mlva.u-psud.fr/). Panel 2B, which comprised the most variable loci, displayed a very high discriminatory power, while panels 1 and 2A showed limited diversity. The most frequent genotype comprised seven isolates obtained over 7 weeks in 2002, demonstrating an outbreak from a common source. Two isolates obtained from one patient 5 months apart comprised another genotype, indicating relapsing disease. These findings confirm that MLVA-16 has a good discriminatory power for species determination, typing of B. melitensis isolates, and inferring their geographical origin. Abbreviated panel 2B could be used as a short-term epidemiological tool in a small region of endemicity

Prevalence and clinical relevance of Helicobacter pylori cagA and vacA genes in Lebanese patients with gastritis and peptic ulcer disease

Background: The prevalence and clinical relevance of H. pylori cagA and vacA virulence genes in the pathogenesis of disease phenotype was assessed by a novel approach for this organism consisting of determination and comparisons of H. pylori gene transcription levels directly in gastric biopsies according to disease phenotype.Methodology: Gastric mucosal biopsies were collected from patients with peptic ulcer disease (PUD), gastritis, and normal mucosa in an academic medical center in Lebanon. H. pylori was detected in these biopsies by rapid urease (CLO™) test and PCR amplification of the ureA gene. H. pylori virulence genes, their transcription and transcription levels were determined respectively by PCR, RT-PCR and real time RT-PCR. Results: Forty-five percent of patients were H. pylori positive by PCR of the ureA gene, 37.5% of whom had cagA and 59.4% vacA.Conclusions: The cagA and vacA genes were detected more frequently in PUD patients with significantly higher transcription levels than in gastritis and normal mucosa