A new species of parvovirus tentatively named human bocavirus 4 (HBoV4) was genetically characterized. Among 641 feces samples from children and adults the most commonly detected bocaviruses species were HBoV2>HBoV3>HBoV4>HBoV1 with HBoV2 prevalence of 21% and 26% in Nigerian and Tunisian children. HBoV3 and HBoV4 species combined were found in 12/192 cases of non-polio acute flaccid paralysis (AFP) from Tunisia and Nigeria and 0/96 healthy Tunisian contacts (p=0.01). Evidence of extensive recombination at the NP1 and VP1 gene boundary between and within species was found. The multiple species and high degree of genetic diversity seen among the human bocaviruses found in feces relative to the highly homogeneous HBoV1 suggest that this world-wide distributed respiratory pathogen may have recently evolved from an enteric bocavirus, perhaps after acquiring an expanded tropism favoring the respiratory track. Elucidating the possible role of the newly identified enteric bocaviruses in human diseases including AFP and diarrhea will require further epidemiological studies.

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A novel picornavirus genome was sequenced, showing 42.6%, 35.2%, and 44.6% of deduced amino acid identities corresponding to the P1, P2, and P3 regions, respectively, of the Aichi virus. Divergent strains of this new virus, which we named salivirus, were detected in 18 stool samples from Nigeria, Tunisia, Nepal, and the United States. A statistical association was seen between virus shedding and unexplained cases of gastroenteritis in Nepal (P ‫؍‬ 0.0056). Viruses with approximately 90% nucleotide similarity, named klassevirus, were also recently reported in three cases of unexplained diarrhea from the United States and Australia and in sewage from Spain, reflecting a global distribution and supporting a pathogenic role for this new group of picornaviruses.The falling cost of DNA sequencing has led to a recent surge in human and animal virus discoveries (1-3, 5-12, 14, 16-17, 19-24, 27, 30, 31, 33, 39, 43, 44). While the pathogenicity of some newly characterized human viruses has been demonstrated, it remains unknown or controversial for other viruses, which may be commensal or pathogenic in only a very small fraction of infections (25,32,40,42,45). Genetic characterization of previously unknown viruses allows the rapid design of nucleic acid tests needed to determine their association with different medical conditions, their presence in different populations, and the design of antibody tests for determining seroprevalence (25, 28, 34, 35, 47).Using sequence-independent PCR amplification, pyrosequencing, and sequence similarity searches (46) (see the text in the supplemental material), we analyzed the virus sequences present in 95 stool samples from Nigerian children suffering from nonpolio acute flaccid paralysis (AFP). Sequences derived from a 10-month-old female child exhibiting right-side asymmetric sudden flaccid paralysis (patient no. NG-J1) formed a 6,981-bp contig consisting of 2,903 individual sequence reads, which was distantly related to sequence of the Aichi virus species in the Kobuvirus genus of the Picornaviridae family (48, 49). Similar sequences were also observed in a second, 24-month-old patient with right-side asymmetric sudden flaccid paralysis (patient no. NG-F1). Gaps between sequenced viral fragments were connected by nested reverse transcription-PCR (RT-PCR), while the 5Ј and 3Ј extremity sequences were acquired using primers designed over conserved regions of bovine, porcine, and human kobuviruses. We temporarily named these viruses saliviruses (stool Aichi-like viruses).The resulting salivirus genome, NG-J1, was 7,124 bp in length with a GC content of 57%, excluding a poly(A) tail. NG-J1 contained a large open reading frame of 7,125 bp encoding a putative polyprotein precursor of 2,374 amino acids (aa), a 5Ј untranslated region (UTR) of 709 bp, and a 3ЈUTR of 148 bp (Fig. 1).NG-J1 and NG-F1 were highly similar, with nucleotide similarities of 94% and 95% in the P1 and P3 regions, respectively. Salivirus NG-J1 had approximately 90% nucleotide similarity to the recently described klasse...

To assess the dynamics of genetic reversion of live poliovirus vaccine in humans, we studied molecular evolution in Sabin-like poliovirus isolates from Nigerian acute flaccid paralysis cases obtained from routine surveillance. We employed a novel modeling approach to infer substitution and recombination rates from whole-genome sequences and information about poliovirus infection dynamics and the individual vaccination history. We confirmed observations from a recent vaccine trial that VP1 substitution rates are increased for Sabin-like isolates relative to the rate for the wild type due to increased nonsynonymous substitution rates. We also inferred substitution rates for attenuating nucleotides and confirmed that reversion can occur in days to weeks after vaccination. We combine our observations for Sabin-like virus evolution with the molecular clock for VP1 of circulating wild-type strains to infer that the mean time from the initiating vaccine dose to the earliest detection of circulating vaccine-derived poliovirus (cVDPV) is 300 days for Sabin-like virus type 1, 210 days for Sabin-like virus type 2, and 390 days for Sabin-like virus type 3. Phylogenetic relationships indicated transient local transmission of Sabin-like virus type 3 and, possibly, Sabin-like virus type 1 during periods of low wild polio incidence. Comparison of Sabin-like virus recombinants with known Nigerian vaccine-derived poliovirus recombinants shows that while recombination with non-Sabin enteroviruses is associated with cVDPV, the recombination rates are similar for Sabin isolate-Sabin isolate and Sabin isolate-non-Sabin enterovirus recombination after accounting for the time from dosing to the time of detection. Our study provides a comprehensive picture of the evolutionary dynamics of the oral polio vaccine in the field. IMPORTANCEThe global polio eradication effort has completed its 26th year. Despite success in eliminating wild poliovirus from most of the world, polio persists in populations where logistical, social, and political factors have not allowed vaccination programs of sustained high quality. One issue of critical importance is eliminating circulating vaccine-derived polioviruses (cVDPVs) that have properties indistinguishable from those of wild poliovirus and can cause paralytic disease. cVDPV emerges due to the genetic instability of the Sabin viruses used in the oral polio vaccine (OPV) in populations that have low levels of immunity to poliovirus. However, the dynamics responsible are incompletely understood because it has historically been difficult to gather and interpret data about evolution of the Sabin viruses used in OPV in regions where cVDPV has occurred. This study is the first to combine whole-genome sequencing of poliovirus isolates collected during routine surveillance with knowledge about the intrahost dynamics of poliovirus to provide quantitative insight into polio vaccine evolution in the field. O ver the last 50 years, mass vaccination campaigns with oral polio vaccine (OPV) have been a key fa...

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BackgroundDespite several studies on the seroprevalence of antibodies against Crimean-Congo Haemorrhagic Fever virus (CCHFV) from humans and cattle in Nigeria, detailed investigation looking at IgG and IgM have not been reported. Additionally, there have been no confirmed cases of human CCHFV infection reported from Nigeria.Principal FindingsSamples from sera (n = 1189) collected from four Local Government Areas in Borno State (Askira/Uba, Damboa, Jere and Maiduguri) were assessed for the presence of IgG and IgM antibodies. The positivity rates for IgG and IgM were 10.6% and 3.5%, respectively. Additionally, sera from undiagnosed febrile patients (n = 380) were assessed by RT-PCR assay for the presence of CCHFV RNA. One positive sample was characterised by further by next generation sequencing (NGS) resulting in complete S, M and L segment sequences.ConclusionsThis article provides evidence for the continued exposure of the human population of Nigeria to CCHFV. The genomic analysis provides the first published evidence of a human case of CCHFV in Nigeria and its phylogenetic context.

Laboratory diagnosis has played a critical role in the Global Polio Eradication Initiative since 1988, by isolating and identifying poliovirus (PV) from stool specimens by using cell culture as a highly sensitive system to detect PV. In the present study, we aimed to develop a molecular method to detect PV directly from stool extracts, with a high efficiency comparable to that of cell culture. We developed a method to efficiently amplify the entire capsid coding region of human enteroviruses (EVs) including PV. cDNAs of the entire capsid coding region (3.9 kb) were obtained from as few as 50 copies of PV genomes. PV was detected from the cDNAs with an improved PV-specific real-time reverse transcription-PCR system and nucleotide sequence analysis of the VP1 coding region. For assay validation, we analyzed 84 stool extracts that were positive for PV in cell culture and detected PV genomes from 100% of the extracts (84/84 samples) with this method in combination with a PV-specific extraction method. PV could be detected in 2/4 stool extract samples that were negative for PV in cell culture. In PV-positive samples, EV species C viruses were also detected with high frequency (27% [23/86 samples]). This method would be useful for direct detection of PV from stool extracts without using cell culture. Laboratory diagnosis has played a critical role in the Global Polio Eradication Initiative since 1988, by detecting and identifying poliovirus (PV). To date, PV has been detected by isolating the virus, using cell culture systems, from stool specimens from acute flaccid paralysis (AFP) cases, including polio and other paralysis cases, in the World Health Organization (WHO) Global Polio Laboratory Network (1, 2). The advantages of the cell culture-based procedure are (i) minimal equipment requirements, (ii) high sensitivity (detection limit of 1 infectious unit containing 50 to 1,000 virions) (3), and, importantly, (iii) biological amplification of PV to high titers (about 10 6 50% cell culture infectious dose [CCID 50 ] or 10 8 to 10 9 viral genome copies per l of cell lysate) for subsequent nucleotide sequence analysis of the capsid protein VP1 coding region, which is required for final identification and molecular epidemiological analysis of PV strains. Sequencing is essential for classifying PV isolates into vaccine strains, wild-type strains, and circulating vaccine-derived PV (VDPV) strains and for determining the necessity of additional vaccination plans by identifying virus reservoirs, along with molecular epidemiological methods (4). A major disadvantage of the cell culture system is the timeliness of reporting; it takes 10 days to confirm that a sample is negative for PV (2). To improve the efficiency of PV detection and identification, including the timeliness, the development of methods for direct detection of PV has been encouraged by WHO (http://www.polioeradication.org /Research/Grantsandcollaboration.aspx). Such direct detection methods must provide efficient PV detection, comparable to that of cell cultur...

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