Phylogenetic and recombination analysis was performed on 32 complete hepatitis E virus (HEV) genomes from infected humans and pigs. For the first time, evidence for recombination between divergent HEV strains was obtained, with at least two strains being found to have discordant phylogenetic relationships consistent with the occurrence of intragenotype recombination. This finding confirms that humans can be dually infected with divergent HEV strains and has implications for the emergence and evolution of new HEV epidemics.Hepatitis E virus (HEV) is a causative agent of enterically transmitted hepatitis disease and, apart from infections of pregnant women, is generally associated with low mortality (11,17). Outbreaks of HEV have been described to occur in many regions of the world, including Asia, Africa, and Mexico (1, 33, 53), while sporadic cases have been documented in the United States, Africa (9), and Europe (27,37,38,60). HEV is presumed to be a zoonotic disease, as surveys of both wild and domestic animals have found evidence for human-related HEVs in rats (12, 22), pigs (2,7,21,29,32,56), deer (45), and wild boar (41). Direct animal-to-human transmission of HEV has been demonstrated, for example, from deer to human as a result of eating uncooked meat (45). Interestingly, HEV characterized from swine in countries where HEV is not endemic is usually closely related to cases of sporadic human HEVs from the same country, for example, in the United States and Japan (29,30,42,57), while in India, where HEV is endemic, human and swine HEV strains are not closely related (2). This suggests that in certain regions HEV is more established in the human population and that in other regions HEV is emerging as a result of direct contact with animal reservoirs for the virus. Divergent HEVs with no human counterparts have also been found in chickens (avian HEV). Avian HEV shares only 50 to 60% nucleotide sequence identity with HEV from humans and swine (15,16).HEV is a positive single-stranded RNA virus that is approximately 7,200 nucleotides (nt) in length. HEV's genome is capped and polyadenylated and has three open reading frames (ORF) (Fig. 1). As the organization of HEV's genome resembles that of Caliciviridae, at one time it was considered a mem-FIG. 1. Genomic organization of HEV. The scale at the bottom of the figure corresponds to nucleotides (in thousands). ORF1 encodes a nonstructural polyprotein which provides guanylyl-methyltransferase and RNA-dependent RNA polymerase activities and possibly other functions (23,25). The ORF2 and ORF3 proteins are believed to be encoded by individual subgenomic RNAs generated during replication (44, 58). ORF2 encodes the viral capsid protein; ORF3, which contains one potential phosphorylation site, encodes a very small protein of about 123 amino acids (59). ORF3 proteins partition with the cytoskeleton, and it has been suggested that an interaction between ORF2 and ORF3 is associated with the assembly of virions (47,50). ORF3 is also believed to interact with proteins involv...

BackgroundStreptococcus pneumoniae infections remain a major cause of morbidity and mortality worldwide. The diversity of pneumococci was first evidenced by serotyping of their capsular polysaccharides, responsible of virulence, resolving into more than 93 serotypes. Molecular tools have been developed to track the emergence and the spread of resistant, hyper virulent or non-vaccine type clones, particularly DNA-based methods using genetic polymorphism. Pulsed-Field Gel Electrophoresis analysis (PFGE) and Multiple Loci Sequence Typing (MLST) are the most frequently used genotyping techniques for S. pneumoniae. MLST is based on sequence comparison of housekeeping genes clustering isolates within sequence types. The availability of genome sequence data from different S. pneumoniae strains facilitated the search for other class of genetic markers as polymorphic DNA sequences for a Multiple-Locus Variable-Number Tandem-Repeat Analysis (MLVA). This study aims at confirming the relevance of MLVA of S. pneumoniae, comparing MLST and MLVA performances when discriminating subgroups of strains belonging to the same Sequence Type (ST), and defining a restricted but universal set of MLVA markers that has at least the same discriminatory power as MLST for S. pneumoniae by applying marker sets used by different authors on 331 isolates selected in UK.ResultsA minimum spanning tree was built including the serotypes distribution and comparing MLVA and MLST results. 220 MLVA types were determined grouped in 10 Sequence Types (ST). MLVA differentiated ST162 in two clonal complexes. A minimal set was defined: ms 25 and ms37, ms17, ms19, ms33, ms39, and ms40 including two universal markers. The selection was based on MLVA markers with a Diversity Index >0.8 and a selection of others depending of the population tested and the aim of the study. This set of 7 MLVA markers yields strain clusters similar to those obtained by MLST.ConclusionsMLVA can discriminate relevant subgroups among strains belonging to the same ST. MLVA offers the possibility to deduce the ST from the MLVA Type. It permits to investigate local outbreaks or to track the worldwide spread of clones and the emergence of variants.

Hepatitis E virus (HEV) is globally distributed, transmitted enterically and between humans and animals. Phylogenetic analysis has identified five distinct HEV genotypes. The first full-length sequence of an African strain (Chad) is presented and compared to 31 complete HEV genomes available, including the fulminant hepatitis strain from India, swine strains and a strain from Morocco. The two African strains are more closely related to genotype 1 than to any other genotypes and together they possibly form a sub-genotype or sixth genotype. The first evidence for recombination between divergent HEV strains is presented.

Genomic analysis of Salmonella enterica revealed the existence of a variable number of tandem repeats (VNTR) at multiple loci. Some S. enterica strains are considered as references (Typhi Ty2, Typhi CT18, Typhimurium LT2, Enteritidis LK5, PT4, and Enteritidis 07-2642, and Newport). These allowed the selection of markers to develop the genotyping technique, multiple-locus VNTR analysis (MLVA). These markers were used to discriminate S. enterica isolated from humans, food, or the environment. In this report, the characteristics and specifications of 58 salmonella markers described from 2003 to 2009 are analyzed. Some VNTR loci were used as markers. The markers were used to discriminate S. enterica isolates from different sources and geographical localizations. Among the VNTR loci described in the published reports, eight presented with a high diversity index (DI) of polymorphism of more than 0.80. The selection of several markers within a single locus validated their polymorphism characteristic. Despite unequal DI values, the use of a panel of markers is a powerful discriminatory tool for the surveillance and identification of the source of salmonella outbreak. Depending on the markers selected, MLVA should be used either for macro- or microepidemiological purposes. The main challenge in the future for this technique is standardization.

BackgroundSalmonella (S.) enterica is the main cause of salmonellosis in humans and animals. The epidemiology of this infection involves large geographical distances, and strains related to an episode of salmonellosis therefore need to be reliably discriminated. Due to the limitations of serotyping, molecular genotyping methods have been developed, including multiple loci variable number of tandem repeats (VNTR) analysis (MLVA). In our study, 11 variable number tandem-repeats markers were selected from the S. enterica Typhimurium LT2 genome to evaluate the genetic diversity of 206 S. enterica strains collected in Cambodia between 2001 and 2007.FindingsThirty one serovars were identified from three sources: humans, animals and food. The markers were able to discriminate all strains from 2 to 17 alleles. Using the genotype phylogeny repartition, MLVA distinguished 107 genotypes clustered into two main groups: S. enterica Typhi and other serovars. Four serovars (Derby, Schwarzengrund, Stanley, and Weltevreden) were dispersed in 2 to 5 phylogenic branches. Allelic variations within S. enterica serovars was represented using the minimum spanning tree. For several genotypes, we identified clonal complexes within the serovars. This finding supports the notion of endemo-epidemic diffusion within animals, food, or humans. Furthermore, a clonal transmission from one source to another was reported. Four markers (STTR3, STTR5, STTR8, and Sal20) presented a high diversity index (DI > 0.80).ConclusionsIn summary, MLVA can be used in the typing and genetic profiling of a large diversity of S. enterica serovars, as well as determining the epidemiological relationships of the strains with the geography of the area.