Virology
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Item Isolation and Genomic Characterization of Echovirus 11 from faeces of a Non-Human Primate in Nigeria(Springer Nature, 2021-05-16) Faleye, T. O. C.; George, U. E.; Klapsa, D.; Majumdar, M.; Oragwa, A. O.; Adewumi, O. M.; Martin, J.; Adeniji, J. A.We recently investigated the presence of enteroviruses (EVs) in non-human primates (NHPs) in Northern Nigeria and documented the presence of EV-A76 of South-East Asian ancestry in an NHP. In this study, we go further to ask if we could also find EVs in NHPs indigenous to the forested South-south Nigeria. Fresh faecal samples were collected from the floor of 10 cages housing NHPs in Cross River Nigeria, resuspended in PBS and subjected to RNA extraction, cDNA synthesis, PanEnt 50-UTR and PanEnt VP1 PCR assays. None of the samples was positive for the PanEnt VP1 assay, but one sample was positive for PanEnt 50- UTR PCR. This sample was subsequently inoculated into RD cell line, produced CPE and the isolate analysed by PCR assays, next-generation whole genome sequencing and passage in four different cell lines showing replication in two of them. Analysis of the complete genome of the isolate identified it as an Echovirus 11 (E11) and revealed a recombinant genomic structure. Phylogenetic analysis showed that the E11 NHP strain was related to human clinical isolates suggesting a zoonotic behaviour. We describe the first isolation and complete genome characterization of an E11 obtained from an NHP in Nigeria having zoonotic potential.Item Extending the utility of the WHO recommended assay for direct detection of enteroviruses from clinical specimen for resolving poliovirus co‑infection(BMC Research Notes, 2018) Faleye, T. O. C; Adewumi, M. O.; Ozegbe, N. P.; Ogunsakin, O. E.; Ariyo, G.; Adeshina, F. W.; Ogunga, O. S.; Oluwadare, S. D.; Adeniji, J. A.Objectives: In a polio-free world there might be reduced funding for poliovirus surveillance. There is therefore the need to ensure that enterovirologist globally, especially those outside the global polio laboratory network, can participate in poliovirus surveillance without neglecting their enterovirus type of interest. To accomplish this, assays are needed that allow such active participation. Results: In this study we describes a sensitive and specific utility extension of the recently recommended WHO RT-snPCR assay that enables independent detection of the three poliovirus types especially in cases of co-infection. More importantly, it piggy-backs on the first round PCR product of the WHO recommended assay and consequently ensures that enterovirologists interested in nonpolio enteroviruses can continue their investigations, and contribute significantly and specifically to poliovirus surveillance, by using the excess of their first round PCR product.Item Isolation and Identification of Enteroviruses from Sewage and Sewage-Contaminated Water Samples from Ibadan, Nigeria, 2012-2013(SciTechnol, 2017) Adeniji, J. A.; Adewale, A. O.; Faleye, T. O. C.; Adewumi, M. O.In 2010, we described sewage contaminated water (SCW) bodies that consistently yielded enteroviruses (EVs) in enterovirus surveillance (ES) sites in Lagos, Nigeria. By 2012, we demonstrated the presence and circulation of Wild Poliovirus 3 (WPV3) in these ES sites. Here we describe ES sites that consistently yield EVs in Ibadan metropolis southwest Nigeria. Twenty-five ES samples were collected by grab method from nine sites between October, 2012 and March, 2013. Samples were concentrated and four (RD, HEp2C, MCF-7 and L20B) different cell lines used for virus isolation from the concentrates. Isolates were subjected to RNA extraction, cDNA synthesis, PanEnterovirus 5l-UTR and VP1 assays. Unidentifiable isolates were further subjected to species-specific RTPCR assays. Amplicons were sequenced, isolates identified and subjected to phylogenetic analysis. Twenty-five isolates were recovered from 8 (32%) of the samples collected. Twenty-three of the isolates were identified as EVs by the PanEntero5l-UTR assay. Thirteen (57%) of the 23 EVs were positive for the VP1 assay, and identified as Coxsackievirus B3 (CVB3) (1 isolate), CVB6 (1 isolate), E6 (2 isolates), E7 (5 isolates), E11 (1 isolate), E12 (1 isolate) and E13 (2 isolates). None and 2 (25%) of the remaining isolates were positive for the EV-B and EV-C assays, respectively. The 2 EV-C positive enteroviruses were isolated on MCF-7. This study describes three very productive ES sites, and documents the presence of CVB3, CVB6, E6, E7, E11, E12 and, E13 in Ibadan, Nigeria. Including other cell lines in EV isolation protocols can broaden the diversity of EV types recoverable.Item Non-polio enteroviruses in faeces of children diagnosed with acute flaccid paralysis in Nigeria(Springer Nature, 2017) Faleye, T. O. C.; Adewumi, M. O.; Japhet, M. O.; David, O. M.; Oluyege, A. O.; Adeniji, J. A.; Famurewa, O.The need to investigate the contribution of non-polio enteroviruses to acute flaccid paralysis (AFP) cannot be over emphasized as we move towards a poliovirus free world. Hence, we aim to identify non-polio enteroviruses recovered from the faeces of children diagnosed with AFP in Nigeria.Ninety-six isolates, (95 unidentified and one previously confirmed Sabin poliovirus 3) recovered on RDcell culture from the stool of children <15 years old diagnosed with AFP in 2014 were analyzed. All isolates were subjected to RNA extraction, cDNA synthesis and three different PCR reactions (one panenterovirus 5′-UTR and two different VP1 amplification assays). VP1 amplicons were then sequenced and isolates identified. Results: 92.71% (89/96) of the isolates were detected by at least one of the three assays as an enterovirus. Precisely, 79.17% (76/96), 6.25% (6/96), 7.30% (7/96) and 7.30% (7/96) of the isolates were positive for both, positive and negative, negative and positive, as well as negative for both the 5′-UTR and VP1 assays, respectively. In this study, sixty-nine (69) of the 83 VP1 amplicons sequenced were identified as 27 different enterovirus types. The most commonly detected were CV-B3 (10 isolates) and EV-B75 (5 isolates). Specifically, one, twenty-four and two of the enterovirus types identified in this study belong to EV-A, EV-B and EV-C respectively. This study reports the circulating strains of 27 non-polio enterovirus types in Nigerian children with AFP in 2014 and Nigerian strains of CV-B2, CV-B4, E17, EV-B80, EV-B73, EV-B97, EV-B93, EV-C99 and EV-A120 were reported for the first time. Furthermore, it shows that being positive for the 5′-UTR assay should not be the basis for subjecting isolates to the VP1 assays.Item Enterovirus A119 in A Child with Acute Flaccid Paralysis, Nigeria(MedCrave Group, 2016) Adeniji, J. A.; Oragwa, A. O.; George, U. E.; Ibok, U. I.; Faleye, T. O. C.; Adewumi, M. O.The oldest EV-A119 record was in 2008 in a chimpanzee in Cameroon and subsequently in more non-human primates and healthy children. Here we report for the first time the detection of EV-A119 in a child with Acute Flaccid Paralysis, thus suggesting possible association with a clinical condition in humansItem "Defining the Enterovirus Diversity Landscape of a Fecal Sample: A Methodological Challenge?"(MDPI, 2015) Faleye, T. O. C.; Adewumi, M. O.; Adeniji, J. A.Enteroviruses are a group of over 250 naked icosahedral virus serotypes that have been associated with clinical conditions that range from intrauterine enterovirus transmission withfataloutcome through encephalitis and meningitis, to paralysis. Classically, enterovirus detection was done by assaying for the development of the classic enterovirus-specific cytopathic effect in cell culture. Subsequently, the isolates were historically identified by a neutralization assay.More recently, identification has been done by reverse transcriptase-polymerase chain reaction (RT-PCR). However, in recent times, there is a move towards direct detection and identification of enteroviruses from clinical samples using the cell culture-independent RT semi-nested PCR (RT-snPCR) assay. This RT-snPCR procedure amplifies the VP1 gene, which is then sequenced and used for identification. However, while cell culture-based strategies tend to show a preponderance of certain enterovirus species depending on the cell lines included in the isolation protocol, the RT-snPCR strategies tilt in a different direction. Consequently, it is becoming apparent that the diversity observed in certain enterovirus species, e.g., enterovirus species B(EV-B), might not be because they are the most evolutionarily successful. Rather, it might stem from cell line-specific bias accumulated over several years of use of the cell culture-dependent isolation protocols. Furthermore, it might also be a reflection of the impact of the relative genome concentration on the result of pan-enterovirus VP1 RT-snPCR screens used during the identification of cell culture isolates. This review highlights the impact of these two processes on the current diversity landscape of enteroviruses and the need to re-assess enterovirus detection and identification algorithms in a bid to better balance our understanding of the enterovirus diversity landscape.