Positive selection pressure on E2 protein of classical swine fever virus drives variations in virulence, pathogenesis and antigenicity: Implication for epidemiological surveillance in endemic areas.

Classical swine fever (CSF), caused by CSF virus (CSFV), is considered one of the most important infectious diseases with devasting consequences for the pig industry. Recent reports describe the emergence of new CSFV strains resulting from the action of positive selection pressure, due mainly to the bottleneck effect generated by ineffective vaccination. Even though a decrease in the genetic diversity of the positively selected CSFV strains has been observed by several research groups, there is little information about the effect of this selective force on the virulence degree, antigenicity and pathogenicity of this type of strains. Hence, the aim of the current study was to determine the effect of the positive selection pressure on these three parameters of CSFV strains, emerged as result of the bottleneck effects induced by improper vaccination in a CSF-endemic area. Moreover, the effect of the positively selected strains on the epidemiological surveillance system was assessed. By the combination of in vitro, in vivo and immunoinformatic approaches, we revealed that the action of the positive selection pressure induces a decrease in virulence and alteration in pathogenicity and antigenicity. However, we also noted that the evolutionary process of CSFV, especially in segregated microenvironments, could contribute to the gain-fitness event, restoring the highly virulent pattern of the circulating strains. Besides, we denoted that the presence of low virulent strains selected by bottleneck effect after inefficient vaccination can lead to a relevant challenge for the epidemiological surveillance of CSF, contributing to under-reports of the disease, favouring the perpetuation of the virus in the field. In this study, B-cell and CTL epitopes on the E2 3D-structure model were also identified. Thus, the current study provides novel and significant insights into variation in virulence, pathogenesis and antigenicity experienced by CSFV strains after the positive selection pressure effect.

Genetic characterization of the glycoprotein G of Chandipura viruses in India with emphasis on an outbreak of 2015.

Chandipura virus (CHPV) is found to be associated with sporadic encephalitis outbreaks in humans in India since 1965. We report here, the investigation of CHPV activity during the period of June-August 2015 in the state of Gujarat, which revealed 24.44% positivity among 45 referred encephalitis cases. Phylogenetic study of the G gene sequences of strains from Gujarat 2015 along with available sequences of additional strains from different geographical locations and isolation years (1965-2015), indicated the relatedness of the 2015 strain to a group of the CHPV prototype strain of 1965 and the earliest outbreak strains of 2003. Analyses of selection pressure in the G gene revealed positively selected sites within the signal peptide region and a putative CHPV epitope. These results indicate a probable role of G protein-based immune selection and underline the need for continued surveillance to monitor genetic and antigenic variations in the CHPV.

Serotypes of HFMD-associated HEV-B species and genetic characteristics of the VP1 gene in coxsack-ievirus B2 and B5 strains isolated in Anyang area from 2011 to 2015 / 中华微生物学和免疫学杂志

Objective To investigate the serotypes of human enterovirus B ( HEV-B) species cau-sing hand, foot and mouth disease ( HFMD) and to analyze the genetic characteristics of VP1 region in cox-sackievirus B2 ( CVB2 ) and coxsackievirus B5 ( CVB5 ) strains circulating in Anyang area during 2011 to 2015. Methods Real-time RT-PCR and semi-nested RT-PCR were performed to identify coxsackievirus A16 (CVA16), enterovirus 71 (EV71) and other serotypes of enterovirus in order to obtain the complete etiologic composition of HFMD. The numbers of HEV-B serotypes and the percentages of specimens positive for every serotype in all enterovirus-positive specimens were calculated. As CVB2 and CVB5 were the pre-dominant serotypes of HEV-B species, five pairs of primers targeting the VP1 regions of CVB2 and CVB5 were designed to obtain the complete nucleotide sequences of CVB2 and CVB5 VP1 regions. The phylogenet-ic trees were constructed based on the VP1 sequences obtained in this study and those submitted to GenBank by using MEGA7. 0 and BioEdit7. 2. The selection pressures on VP1 regions of CVB2 and CVB5 strains cir-culating in China in recent years were evaluated with the online program of DataMonkey. Results A total of 57 specimens that belonged to 14 serotypes of HEV-B species were detected in Anyang area from 2011 to 2015. The 14 serotypes of HEV-B species accounted for 56% of all serotypes of enterovirus and the speci-mens positive for HEV-B species accounted for 3. 06% of all enterovirus-positive specimens. The HFMD ca-ses caused by most of the HEV-B serotypes were sporadic cases. Small outbreaks of HFMD could also be caused by some serotypes of HEV-B such as CVB2 and CVB5. The complete sequences of VP1 region were obtained from 8 CVB2 strains and 9 CVB5 strains. The phylogenetic trees based on the VP1 sequences dem-onstrated that the CVB2 strains were classified into four genotypes ( A-D) . The mean evolutionary distances between different genotypes ranged from 0. 191 to 0. 208 and the similarities in nucleotide sequences ranged from 79. 7% to 85. 8%. The CVB5 strains were classified into 6 genotypes (A-F). The mean evolutionary distances and the similarities in nucleotide sequences between different genotypes of CVB5 strains ranged from 0. 170 to 0. 285 and 76. 0% to 86. 8%, respectively. Strains of different genotypes varied significantly in the residues on positons 157 and 263 in the VP1 region of CVB2 strains and on positions 75, 90 and 95 in the VP1 region of CVB5 strains. All of the CVB2 strains isolated in Anyang area belonged to D genotype and located intensively in one lineage. The CVB5 strains circulated in Anyang area belonged to F genotype and located in two lineages. The selection pressures on CVB2 strains of D genotype and CVB5 strains of F geno-type circulating in China in recent years were 0. 037 and 0. 036, respectively. Six positively selected amino acid sites were found in the VP1 region of CVB5 strains, but no positively selected amino acid site was found in the VP1 region of CVB2 strains. Conclusion HEV-B species was an essential component of the etiologic spectrum of HFMD in Anyang area during 2011 to 2015, of which CVB5 and CVB2 were the predominant se-rotypes. The VP1 region of CVB5 was more complex and active than that of CVB2 over the course of evolution.

RHOXF2 gene, a new candidate gene for spermatogenesis failure.

INTRODUCTION: Genes involved in testicular differentiation, spermatogenesis, proliferation and apoptosis of germ cells have been shown to evolve rapidly and display rapid DNA changes. These genes are therefore good candidates for explaining impairments in spermatogenesis. Initial studies of some of these genes appear to confirm this hypothesis. The RHOXF2 candidate gene belongs to the RHOX family clustered in Xq24 and is specifically expressed in the testis. It contains four exons and codes for a 288 amino acid (aa) transcription factor. It has a high degree of homology (>99.9%) with its paralogue RHOXF2B, which is also preferentially expressed in the testis. OBJECTIVES: To sequence RHOXF2 and RHOXF2B in intracytoplasmic sperm injection (ICSI) patients and identify any single-nucleotide polymorphisms (SNPs) associated with impaired spermatogenesis. MATERIALS: A cohort of 327 patients in ICSI programmes at Poissy and Bichat hospitals. All patients gave their written, informed consent to participation. One hundred patients had unaffected spermatogenesis and 227 displayed impaired spermatogenesis. METHODS: The four exons in each of RHOXF2 and RHOXF2B were sequenced in 47 patients with oligospermia or non-obstructive azoospermia. Given that exons 2 and 3 were found to harbour most of the SNPs, only these two exons were sequenced in the remaining 280 subjects. RESULTS: Due to the extremely high degree of sequence identity between RHOXF2 and RHOXF2B, we were not able to distinguish between the sequences of these two genes. Although 9 SNPs were identified, there were no significant frequency differences between ICSI patients with normal vs. impaired spermatogenesis. Two insertions were identified: a 21-nucleotide insertion was retrieved in both groups and a guanine insertion (inducing a premature stop codon) only found in two patients with impaired spermatogenesis. CONCLUSION/OUTLOOK: RHOXF2 is a good candidate for rapid evolution by positive selection. Analysis of the polymorphism frequency in exons 2 and 3 did not allow us to correlate the identified SNPs with male infertility. However, a single nucleotide insertion was identified only in men with impaired spermatogenesis. Further work will be needed to establish whether genetic changes in RHOXF2 can give rise to defects in spermatogenesis.

INTRODUCTION: Les gènes impliqués dans la différenciation des testicules, la spermatogenèse, la prolifération et l'apoptose des cellules germinales ont été montrés comme ayant une évolution rapide de la séquence d'ADN. Ces gènes sont donc de bons candidats pour expliquer les déficiences de la spermatogenèse. Les premières études semblent confirmer cette hypothèse. Le gène RHOXF2, appartenant à la famille des gènes RHOX avec un cluster dans Xq24, est un bon candidat car spécifiquement exprimé dans les testicules. Ce gène a un degré élevé d'homologie (> 99,9%), avec son paralogue RHOXF2B , qui est également exprimé préférentiellement dans les testicules. OBJECTIFS: Séquencer RHOXF2 chez des patients infertiles bénéficiant d'une injection intracytoplasmique de spermatozoïdes (ICSI) afin d'identifier des polymorphismes associés à une déficience de la spermatogenèse. MATÉRIELS: Une cohorte de 327 patients inclus dans un programme d'ICSI. Tous les patients ont donné leur consentement écrit et éclairé à la participation de cette étude. Cent patients n'avaient pas d'altération de la spermatogenèse et 227 avaient une déficience. MÉTHODES: Les quatre exons de RHOXF2 ont été séquencés chez 47 patients présentant une oligospermie ou une azoospermie non obstructive. Étant donné que les exons 2 et 3 ont été trouvés comme ayant le plus de SNPs, seuls ces deux exons ont été séquencés dans les 280 sujets restants. RÉSULTATS: Bien que 9 SNPs aient été identifiés, il n'y avait pas de différence de fréquences significatives entre les patients ayant une altération, ou non de la spermatogenèse. Deux insertions ont été identifiées: une insertion de 21 nucléotides retrouvées dans les deux groupes et une insertion d'une guanine (induisant un codon stop prématuré) chez deux patients présentant une altération de la spermatogenèse. CONCLUSION: RHOXF2 est un bon candidat pour une évolution rapide par sélection positive. L'analyse de la fréquence des polymorphismes dans les exons 2 et 3 ne nous permet pas actuellement de corréler les SNP identifiés avec l'infertilité masculine. Cependant, une insertion d'un seul nucléotide a été identifiée uniquement chez des hommes avec une déficience de la spermatogenèse. Des travaux complémentaires seront nécessaires pour déterminer l'impact du gène RHOXF2 sur la spermatogenèse.

Positive Selection Analysis of VP1 Genes of Worldwide Human Enterovirus 71 Viruses / 中国病毒学·英文版

Human enterovirus 71 viruses have been long circulating throughout the world. In this study, we performed a positive selection analysis of the VP1 genes of capsid proteins from Enterovirus 71 viruses. Our results showed that although most sites were under negative or neutral evolution, four positions of the VP1 genes were under positive selection pressure. This might account for the spread and frequent outbreaks of the viruses and the enhanced neurovirulence. In particular, position 98 might be involved in neutralizing antibodies, modulating the virus-receptor interaction and enhancing the virulence of the viruses. Moreover, both positions 145 and 241 might correlate to determine the receptor specificity. However, these positions did not display much difference in amino acid polymorphism. In addition, no position in the VP1 genes of viruses isolated from China was under positive selection.