A possible risk of environmental exposure to HEV in Ibadan, Oyo State, Nigeria.

Hepatitis E virus (HEV) infection is both a major public health concern and emerging global health concern, with a documented incidence of 20 million, 3.4 million clinical cases, 70,000 deaths, and 3,000 stillbirths. The aetiologic agent, HEV is a primarily enterally transmitted hepatotropic virus. Fecal samples were collected from three selected pig farms across Ibadan, South-west Nigeria. Randomly picked samples were pooled per unit pen and fecal suspensions prepared were subjected to HEV Antigen (Ag) enzyme-linked immunosorbent assay. Molecular probing was done by Reverse Transcription and nested polymerase reaction (RT-nPCR) and deep sequencing. Sequencing was done paired-end for 300 cycles using the HiSeq system. Overall farm prevalence of 66.7% (2/3) and prevalence at individual level of 13.2% (9/68) were recorded. All nine samples positive for the ELISA screen were negative when subjected to RT-nPCR assays. Further, on deep sequencing, no HEV genomic fragment was found in the sample using de-novo assembly. Findings suggest possibly inapparent HEV in the pigs studied or a yet to be identified protein with HEV-Ag cross-reactivity ability on ELISA, thus constituting a possible risk of exposure to HEV infection in the population. Consequently, we recommend prompt intervention to unravel the mystery and break the chain of transmission.

Isolation and Genomic Characterization of Echovirus 11 from faeces of a Non-Human Primate in Nigeria.

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, re-suspended in PBS and subjected to RNA extraction, cDNA synthesis, PanEnt 5'-UTR and PanEnt VP1 PCR assays. None of the samples was positive for the PanEnt VP1 assay, but one sample was positive for PanEnt 5'-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.

Patterns of serologic markers of hepatitis B virus infection and the risk of transmission among pregnant women in southwestern Nigeria.

Hepatitis B virus (HBV) infection is a major health concern in developing countries that has a high morbidity and mortality rate. Vertical transmission of HBV from mother to child has been identified as a major factor leading to chronicity with attendant liver conditions, especially in poor socioeconomic settings. This study aims to evaluate the prevalence of serological HBV markers among pregnant women in Ibadan southwestern Nigeria and to determine the implications for perinatal HBV transmission. This study revealed the presence of varied HBV serological patterns of infection or immunity among pregnant women in Ibadan, Nigeria, and thus the risk of mother to child transmission.

Non-polio enteroviruses in faeces of children diagnosed with acute flaccid paralysis in Nigeria.

BACKGROUND: 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. METHODS: Ninety-six isolates, (95 unidentified and one previously confirmed Sabin poliovirus 3) recovered on RD cell 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. CONCLUSIONS: 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.

Preponderance of enterovirus C in RD-L20B-cell-culture-negative stool samples from children diagnosed with acute flaccid paralysis in Nigeria.

Recently, a reverse transcriptase semi-nested polymerase chain reaction (RT-snPCR) assay was recommended by the WHO for direct detection of enteroviruses in clinical specimens. In this study, we use this assay and a modification thereof to screen acute flaccid paralysis (AFP) samples that had previously tested negative for enteroviruses by the RD-L20B algorithm. Thirty paired stool suspensions collected in 2015 as part of the national AFP surveillance program in different states of Nigeria were analyzed in this study. The samples had previously tested negative for enteroviruses in the polio laboratory in accordance with the WHO-recommended RD-L20B-cell-culture-based algorithm. Two samples that had previously been found to contain enteroviruses were included as positive controls. All samples were subjected to RNA extraction, the RT-snPCR assay and a modified version of the RT-snPCR. All amplicons were sequenced, and enteroviruses were identified using the enterovirus genotyping tool and phylogenetic analysis. Amplicons were recovered from the two controls and 50% (15/30) of the samples screened. Fourteen were successfully typed, of which, 7.1% (1/14), 21.4% (3/14), 64.3% (9/14) and 7.1% (1/14) were enterovirus (EV) -A, EV-B, EV-C and a mixture of EV-B and C (EV-C99 and E25), respectively. The two controls were identified as EV-C99 and coxsackievirus (CV) -A1, both of which belong to the species Enterovirus C. In one sample, poliovirus serotype 2 was detected and found to have the VP1 ILE143 variation and was therefore identified as a vaccine strain. The results of this study showed that significant proportion of enterovirus infections (including some with Sabin PV2) are being missed by the RD-L20B-cell-culture-based algorithm, thus highlighting the value of the RT-snPCR assay and its modifications. The circulation and preponderance of EV-C in Nigeria was also confirmed.

Comparison of Algorithms for the Detection of Enteroviruses in Stool Specimens from Children Diagnosed with Acute Flaccid Paralysis.

This study was designed to compare both the cell culture dependent and independent enterovirus detection algorithms recommended by the WHO and assess how either might impact our perception of the diversity of enterovirus types present in a sample. Sixteen paired samples (16 isolates from RD cell culture and their corresponding stool suspension, i.e., 32 samples) from AFP cases in Nigeria were analyzed in this study. All the samples were subjected to RNA extraction, cDNA synthesis, the WHO recommended RT-snPCR, and its modification. Amplicons were sequenced and strains identified. Enterovirus diversity was the same between the isolates and fecal suspension for the control and five of the samples. It was, however, different for the remaining 10 (62.5%) samples. Nine (CV-B4, E6, E7, E13, E14, E19, E29, EV-B75, and EV-B77) and five (CV-A1, CV-A11, CV-A13, EV-C99, and PV2) EV-B and EV-C types, respectively, were detected. Particularly, E19 and EV-B75 were only recovered from the isolates while E14, EV-B77, CV-A11, and CV-A13 were only recovered from fecal suspension. Both the cell culture dependent and independent protocols bias our perception of the diversity of enterovirus types present in a sample. Hence, effort should be directed at harmonizing both for increased sensitivity.

Acute Hepatitis E Virus Infection in Two Geographical Regions of Nigeria.

Hepatitis E virus (HEV) remains a major public health concern in resource limited regions of the world. Yet data reporting is suboptimal and surveillance system is inadequate. In Nigeria, there is dearth of information on prevalence of acute HEV infection. This study was therefore designed to describe acute HEV infection among antenatal clinic attendees and community dwellers from two geographical regions in Nigeria. Seven hundred and fifty plasma samples were tested for HEV IgM by Enzyme Linked Immunosorbent Assay (ELISA) technique. The tested samples were randomly selected from a pool of 1,115 blood specimens previously collected for viral hepatitis studies among selected populations (pregnant women, 272; Oyo community dwellers, 438; Anambra community dwellers, 405) between September 2012 and August 2013. One (0.4%) pregnant woman in her 3rd trimester had detectable HEV IgM, while community dwellers from the two study locations had zero prevalence rates of HEV IgM. Detection of HEV IgM in a pregnant woman, especially in her 3rd trimester, is of clinical and epidemiological significance. The need therefore exists for establishment of a robust HEV surveillance system in Nigeria and especially amidst the pregnant population in a bid to improve maternal and child health.

Seroprevalence of infectious bursal disease virus in local chickens in Udu Local Government Area of Delta State, South East Nigeria.

Infectious Bursal Disease Virus (IBDV) poses a great global threat to the poultry industry. Knowledge of the occurrence of the disease is important in the design and implementation of a control program, therefore this study determines the seroprevalence of IBDV in local chickens in Udu Local Government Area of Delta State. 250 chickens were bled by exsanguination and sera obtained were screened using Agar Gel Immunodiffusion (AGID) test. The seropositivity was 51.6%, which is indicates endemicity of the disease. Biosecurity and good sanitary measures are recommended. Molecular characterization of the strains should be carried out for inclusion in generic vaccines.

Reliability of testing and potential impact on HIV prevention in Nigeria.

Several factors including variability of human immunodeficiency virus (HIV), laboratory facilities, cost and competence of personnel handling the tests are some of the important factors that affect accuracy and reliability of HIV testing in most parts of Africa. Recently investigators in Africa have observed that antibody detection assays based on antigens derived from HIV-1 subtype B show moderate to significantly lower sensitivity for detection of infection by various non-B subtypes. In this study, we evaluated the reliability of two EIA and 12 rapid HIV-1/2 test kits that are commercially available in Nigeria using the Western immunoblotting technique as reference. A panel of 100 sera from Western blot confirmed symptomatic or asymptomatic HIV-1 infected persons and 90 seronegative patients from those referred for testing in our laboratory were used for this study. Each sample was tested with two HIV-1/2 EIA, and 12 HIV-1/2 rapid test kits commercially available at one time or the other for HIV-1/2 testing in Nigeria. Overall, the sensitivity of the two EIA kits were 100% and 91.0% with specificity of 96.7% and 91.1% respectively. The sensitivity of the rapid test kits ranged from 88% to 98.0% with specificity of 92.2% to 100%. Further analysis showed significant variation in the sensitivity and specificity of the same kit based on whether an individual had asymptomatic or symptomatic infection The results of this study highlight the problem of diagnosis of HIV infections in Africa. It shows that the sensitivity of most of the rapid assays shall not be adequate for detection of early infection. The implications of possible misdiagnosis on the various intervention strategies that rely predominantly on correct HIV status of an individual are enormous. Thus, there is an urgent need for review of the current HIV testing assays or algorithms in Nigeria and other parts of Africa.

Neutralizing antibodies against poliovirus serotypes among children in southwest Nigeria.

In May 1988, the World Health Assembly resolved to eradicate poliomyelitis globally by the year 2000. Despite the reported success in national immunization days, acute flaccid paralysis surveillance and accelerated efforts to meet the deadline including 'mopping-up' were executed in 1999 and subsequent years. Nigeria remains one of the major reservoirs for wild poliovirus transmission. Neutralizing antibody titre to the three poliovirus serotypes was determined among children from different communities in southwest of Nigeria, and analysed by age, gender and location. About 0.5-2 ml of blood sample was collected by venepuncture from each child. Aliquot of serum from each blood sample was inactivated prior to neutralization test by the beta method for poliovirus antibodies. A total of 347 (59.6 per cent) out of 500 and 82 children enrolled for the study had at least antibody titre of 1:8 against each of the three poliovirus serotypes. Immunity level to the three poliovirus serotypes increased with age and peaked in children aged 4-6 years. Seven (53.8 per cent) out of 13 unvaccinated children tested in the study had detectable neutralizing antibody to the three serotypes. Immunity pattern of P2 > P1 and P3 was observed but no correlation between gender and antibody to the poliovirus serotypes. The populations had 59.6 per cent herd immunity for the three poliovirus serotypes. In a country with high incidence of poliomyelitis this situation leaves a high number of non-immunized children at the risk of infection with one or more poliovirus serotypes.

Time-dependent inhibition of theophylline elimination by erythromycin stearate.

The effect of erythromycin stearate at a dose of 1 g daily on theophylline disposition was studied in six healthy adult males. The mean value of 5.46 h for the theophylline half-life after 48 h pretreatment with erythromycin stearate was not significantly different from the control value of 5.24 h. The half-life value of 8.48 h after a 1-week course of the antibiotic, however, represented a significant increase (p less than 0.005). Theophylline disposition 1 week after the last dose of the antibiotic was not significantly different from the control. It is suggested that therapeutic doses of erythromycin stearate for a week or more could impair the disposition of concurrently administered theophylline. The impairment however, is reversible, being absent a week after the last antibiotic dose.