The trivalent oral poliomyelitis vaccine (OPV) contains three different poliovirus serotypes. It use therefore creates particularly favorable conditions for mixed infection of gut cells, and indeed intertypic vaccine-derived recombinants (VdRec) have been frequently found in patients with vaccine-associated paralytic poliomyelitis. Nevertheless, there have not been extensive searches for VdRec in healthy vaccinees following immunization with OPV. To determine the incidence of VdRec and their excretion kinetics in primary vaccinees, and to establish the general genomic features of the corresponding recombinant genomes, we characterized poliovirus isolates excreted by vaccinees following primary immunization with OPV. Isolates were collected from 67 children 2 to 60 days following vaccination. Recombinant strains were identified by multiple restriction fragment length polymorphism assays. The localization of junction sites in recombinant genomes was also determined. VdRec excreted by vaccinees were first detected 2 to 4 days after vaccination. The highest rate of recombinants was on day 14. The frequency of VdRec depends strongly on the serotype of the analyzed isolates (2, 53, and 79% of recombinant strains in the last-excreted type 1, 2, and 3 isolates, respectively). Particular associations of genomic segments were preferred in the recombinant genomes, and recombination junctions were found in the genomic region encoding the nonstructural proteins. Recombination junctions generally clustered in particular subgenomic regions that were dependent on the serotype of the isolate and/or on the associations of genomic segments in recombinants. Thus, VdRec are frequently excreted by vaccinees, and the poliovirus replication machinery requirements or selection factors appear to act in vivo to shape the features of the recombinant genomes.

Echovirus 6 (E6) and echovirus 11 (E11) are common causes of meningitis and other human diseases; they are among the most frequently isolated enteroviruses worldwide. In the present work we have studied genetic variability over the entire VP1 gene of selected isolates representing a wide geographical and temporal range. Fifty new sequences from North Africa were included, together with previously published sequences from different countries. The sequence diversity between strains of the same type was high: 22 and 30 % for E6 and E11, respectively. Phylogenetic analysis revealed five genogroups within each type, the genetic diversity within a genogroup generally being ,20 %. Some genogroups were further subdivided into genotypes, most containing isolates that had circulated over a wide geographical (several countries from different continents) and temporal (up to two decades) range. Several genotypes were also shown to co-circulate in a region during the same period of time. These features differ from other enteroviruses that divide into temporal or geographical clusters. This study reports new sequences from North Africa, updates the molecular epidemiology of E6 and E11, and proposes a new genogroup in each type. INTRODUCTIONHuman enteroviruses (HEVs) are small non-enveloped viruses with a worldwide distribution. Laboratory diagnosis of related infections currently is based on virus detection by isolation in cell culture or by direct PCR amplification from clinical samples. HEV types are identified by seroneutralization or partial sequencing of the VP1 region (Caro et al., 2001;Norder et al., 2001;Oberste et al., 1999a Oberste et al., , b, 2000Palacios et al., 2002). Similar to other RNA viruses, HEVs have a high capacity to evolve genetically. Genotypes are identified based on the genetic variability and phylogenetic relationships among isolates. The circulation of these genotypes throughout the world and over time has been studied to increase our understanding of the dynamics of their transmission, to evaluate their endemicity and to explain the extent of epidemics when they occur. Beyond type identification, sequencing of the VP1 region has also proved to be a reliable method for such molecular epidemiological studies.Echovirus 6 (E6) and echovirus 11 (E11) are among the most commonly isolated HEVs worldwide, and are frequently associated with outbreaks and sporadic cases of aseptic meningitis, as well as with several diseases ranging from mild non-specific illness to encephalitis, paralysis, myocarditis and severe systemic infections in neonates (Abe et al., 2000;Ashwell et al., 1996;Atkinson et al., 1998;Bahri et al., 2005;Belguith et al., 2007a;Boyd et al., 1987;Cabrerizo et al., 2008;Chomel et al., 2003;Druyts-Voets, 1997;El-Sageyer et al., 1998;Joo et al., 2005;Khetsuriani et al., 2006;Mao et al., 2010;Mirand et al., 2008;Miwa & Sawatari, 1994;Somekh et al., 2001;Ventura et al., 2001). However, despite their important impact on human health, studies of the molecular epidemiology of E6 and E11 remain li...

Among Coxsackie B viruses, Coxsckievirus B5 is one of the most predominant serotypes in human, it is frequently associated with cases of neurological diseases, epidemics of meningitis and is a common cause of cardiomyopathy and diabetes. In the present study 27 isolates of Coxsackievirus B5 from North Africa, obtained from cerebrospinal fluid and stool samples of healthy individuals, patients with acute flaccid paralysis or aseptic meningitis were investigated by partial sequencing in the 5' half of the VP1 region and compared to the up-to-date published Coxsackievirus B5 sequences in the same genomic region. Four distinct genomic groups and ten different clusters were individualized. Most of the isolates from Algeria and Tunisia belonged to two clusters. For both, the sequences from North Africa clustered mainly with sequences from European countries, the majority isolated recently during the 2000s. The analysis of the alignment of amino-acids sequences in the VP1 gene revealed four major substitutions in strains from different clusters, we also noticed changes in the BC-loop region; this region is associated with viral antigenicity. This study permit to better identify circulating Coxsackievirus B5 strains throughout the world and their genetic relationship. The protein analysis showed changes that could imply some antigenic significance. J. Med. Virol. 83:1247-1254, 2011. © 2011 Wiley-Liss, Inc.

Coxsackievirus type B1 (CVB1) has emerged globally as the predominant enterovirus serotype and is associated with epidemics of meningitis and chronic diseases. In this report, the phylogeny of CVB1 was studied based on the VP1 sequences of 11 North African isolates and 81 published sequences. All CVB1 isolates segregated into four distinct genogroups and 10 genotypes. Most of the identified genotypes of circulating CVB1 strains appear to have a strict geographical specificity. The North African strains were of a single genotype and probably evolved distinctly. Using a relaxed molecular clock model and three different population models (constant population, exponential growth and Bayesian skyline demographic models) in coalescent analysis using the BEAST program, the substitution rate in CVB1 varied between 6.95 × 10(-3) and 7.37 × 10(-3) substitutions/site/year in the VP1 region. This study permits better identification of circulating CVB1, which has become one of the most predominant enterovirus serotypes in humans.

Enterovirus A71 (EV-A71) is a leading cause of hand-foot-and-mouth disease (HFMD) and can be associated with severe neurological complications. EV-A71 strains can be classified into seven genogroups, A-H, on the basis of the VP1 capsid protein gene sequence. Genogroup A includes the prototype strain; genogroups B and C are responsible of major outbreaks worldwide, but little is known about the others, particularly genogroups E and F, which have been recently identified in Africa and Madagascar, respectively. The circulation of EV-A71 in the African region is poorly known and probably underestimated. A rapid and specific assay for detecting all genogroups of EV-A71 is required. In this study, we developed a real-time RT-PCR assay with a competitive internal control (IC). The primers and TaqMan probe specifically target the genomic region encoding the VP1 capsid protein. Diverse EV-A71 RNAs were successfully amplified from the genogroups A, B, C, D, E, and F, with similar sensitivity and robust reproducibility. Neither cross reaction with other EVs nor major interference with the competitive IC was detected. Experimentally spiked stool and plasma specimens provided consistent and reproducible results, and validated the usefulness of the IC for demonstrating the presence of PCR inhibitors in samples. The analysis in an African laboratories network of 1889 untyped enterovirus isolates detected 15 EV-A71 of different genogroups. This specific real-time RT-PCR assay provides a robust and sensitive method for the detection of EV-A71 in biological specimens and for the epidemiological monitoring of EV-A71 including its recently discovered genogroups.