Prevalence and spatiotemporal distribution of rotavirus diarrhea among children younger than five years old in Lagos, Nigeria.

Data on spatiotemporal distribution of rotavirus diarrhea are limited in many endemic settings. This study determined the prevalence and seasonal distribution of rotavirus among Nigerian children with diarrhea. Here, a total of 406 fecal samples were collected from patients attending six health facilities in Lagos between January - December 2019. Socio-demographic data of each enrolled child were collected. Rotavirus VP6 antigen was detected by enzyme-linked immunoassay (ELISA) and confirmation by VP7 gene detection by reverse transcription polymerase-chain reaction. The overall rotavirus diarrhea prevalence was 16.3% by ELISA with children above 2 years having 29.2% of this prevalence and higher occurrence in females (59.1%) than males (40.9%) (P < .05). Rotavirus diarrhea diagnosis using RT-PCR showed 100% concordance with ELISA. Cases of rotavirus diarrhea were detected from March to July and from September to November with the highest number of cases detected in May and June (22.7% each), followed by July (21.2%). The prevalence of rotavirus diarrhea remains high in Lagos with an emerging higher disease activity in children above 2. A different rotavirus transmission dynamics compared to previous studies from Nigeria and other African countries was found. VP6 ELISA may reliably be used for continuous rotavirus surveillance in Nigeria.

Dynamics of Evolution of Poliovirus Neutralizing Antigenic Sites and Other Capsid Functional Domains during a Large and Prolonged Outbreak.

We followed the dynamics of capsid amino acid replacement among 403 Nigerian outbreak isolates of type 2 circulating vaccine-derived poliovirus (cVDPV2) from 2005 through 2011. Four different functional domains were analyzed: (i) neutralizing antigenic (NAg) sites, (ii) residues binding the poliovirus receptor (PVR), (iii) VP1 residues 1 to 32, and (iv) the capsid structural core. Amino acid replacements mapped to 37 of 43 positions across all 4 NAg sites; the most variable and polymorphic residues were in NAg sites 2 and 3b. The most divergent of the 120 NAg variants had no more than 5 replacements in all NAg sites and were still neutralized at titers similar to those of Sabin 2. PVR-binding residues were less variable (25 different variants; 0 to 2 replacements per isolate; 30/44 invariant positions), with the most variable residues also forming parts of NAg sites 2 and 3a. Residues 1 to 32 of VP1 were highly variable (133 different variants; 0 to 6 replacements per isolate; 5/32 invariant positions), with residues 1 to 18 predicted to form a well-conserved amphipathic helix. Replacement events were dated by mapping them onto the branches of time-scaled phylogenies. Rates of amino acid replacement varied widely across positions and followed no simple substitution model. Replacements in the structural core were the most conservative and were fixed at an overall rate ∼20-fold lower than the rates for the NAg sites and VP1 1 to 32 and ∼5-fold lower than the rate for the PVR-binding sites. Only VP1 143-Ile, a non-NAg site surface residue and known attenuation site, appeared to be under strong negative selection.IMPORTANCE The high rate of poliovirus evolution is offset by strong selection against amino acid replacement at most positions of the capsid. Consequently, poliovirus vaccines developed from strains isolated decades ago have been used worldwide to bring wild polioviruses almost to extinction. The apparent antigenic stability of poliovirus obscures a dynamic of continuous change within the neutralizing antigenic (NAg) sites. During 7 years of a large outbreak in Nigeria, the circulating type 2 vaccine-derived polioviruses generated 120 different NAg site variants via multiple independent pathways. Nonetheless, overall antigenic evolution was constrained, as no isolate had fixed more than 5 amino acid differences from the Sabin 2 NAg sites, and the most divergent isolates were efficiently neutralized by human immune sera. Evolution elsewhere in the capsid was also constrained. Amino acids binding the poliovirus receptor were strongly conserved, and extensive variation in the VP1 amino terminus still conserved a predicted amphipathic helix.

The antiviral activity of leaves of Eucalyptus camaldulensis (Dehn) and Eucalyptus torelliana (R. Muell).

Human enteroviruses are the major cause of aseptic meningitis and are resistant to all known antibiotics and chemotherapeutic agents. Methanolic extracts of Eucalyptus camaldulensis and Eucalyptus torelliana were tested on human enteroviruses: Poliovirus type I, Coxsackievirus B and Echovirus 6. The virucidal tests showed that the crude extracts were active on the test viruses: poliovirus type 1, coxsackievirus B and echovirus 6 giving a neutralization index of one log and above. The cytotoxicity assay of the crude extracts to L20B (a genetically engineered mouse cell line) and human rhabdomyo sarcoma (RD) cells showed that the extract of E. torelliana was more toxic than the extract of E. camaldulensis. The antiviral study showed that the extract of E. torelliana was more active than that of E. camaldulensis.

Multiple independent emergences of type 2 vaccine-derived polioviruses during a large outbreak in northern Nigeria.

Since 2005, a large poliomyelitis outbreak associated with type 2 circulating vaccine-derived poliovirus (cVDPV2) has occurred in northern Nigeria, where immunization coverage with trivalent oral poliovirus vaccine (tOPV) has been low. Phylogenetic analysis of P1/capsid region sequences of isolates from each of the 403 cases reported in 2005 to 2011 resolved the outbreak into 23 independent type 2 vaccine-derived poliovirus (VDPV2) emergences, at least 7 of which established circulating lineage groups. Virus from one emergence (lineage group 2005-8; 361 isolates) was estimated to have circulated for over 6 years. The population of the major cVDPV2 lineage group expanded rapidly in early 2009, fell sharply after two tOPV rounds in mid-2009, and gradually expanded again through 2011. The two major determinants of attenuation of the Sabin 2 oral poliovirus vaccine strain (A481 in the 5'-untranslated region [5'-UTR] and VP1-Ile143) had been replaced in all VDPV2 isolates; most A481 5'-UTR replacements occurred by recombination with other enteroviruses. cVDPV2 isolates representing different lineage groups had biological properties indistinguishable from those of wild polioviruses, including efficient growth in neuron-derived HEK293 cells, the capacity to cause paralytic disease in both humans and PVR-Tg21 transgenic mice, loss of the temperature-sensitive phenotype, and the capacity for sustained person-to-person transmission. We estimate from the poliomyelitis case count and the paralytic case-to-infection ratio for type 2 wild poliovirus infections that ∼700,000 cVDPV2 infections have occurred during the outbreak. The detection of multiple concurrent cVDPV2 outbreaks in northern Nigeria highlights the risks of cVDPV emergence accompanying tOPV use at low rates of coverage in developing countries.

Implications of a circulating vaccine-derived poliovirus in Nigeria.

BACKGROUND: The largest recorded outbreak of a circulating vaccine-derived poliovirus (cVDPV), detected in Nigeria, provides a unique opportunity to analyze the pathogenicity of the virus, the clinical severity of the disease, and the effectiveness of control measures for cVDPVs as compared with wild-type poliovirus (WPV). METHODS: We identified cases of acute flaccid paralysis associated with fecal excretion of type 2 cVDPV, type 1 WPV, or type 3 WPV reported in Nigeria through routine surveillance from January 1, 2005, through June 30, 2009. The clinical characteristics of these cases, the clinical attack rates for each virus, and the effectiveness of oral polio vaccines in preventing paralysis from each virus were compared. RESULTS: No significant differences were found in the clinical severity of paralysis among the 278 cases of type 2 cVDPV, the 2323 cases of type 1 WPV, and the 1059 cases of type 3 WPV. The estimated average annual clinical attack rates of type 1 WPV, type 2 cVDPV, and type 3 WPV per 100,000 susceptible children under 5 years of age were 6.8 (95% confidence interval [CI], 5.9 to 7.7), 2.7 (95% CI, 1.9 to 3.6), and 4.0 (95% CI, 3.4 to 4.7), respectively. The estimated effectiveness of trivalent oral polio vaccine against paralysis from type 2 cVDPV was 38% (95% CI, 15 to 54%) per dose, which was substantially higher than that against paralysis from type 1 WPV (13%; 95% CI, 8 to 18%), or type 3 WPV (20%; 95% CI, 12 to 26%). The more frequent use of serotype 1 and serotype 3 monovalent oral polio vaccines has resulted in improvements in vaccine-induced population immunity against these serotypes and in declines in immunity to type 2 cVDPV. CONCLUSIONS: The attack rate and severity of disease associated with the recent cVDPV identified in Nigeria are similar to those associated with WPV. International planning for the management of the risk of WPV, both before and after eradication, must include scenarios in which equally virulent and pathogenic cVDPVs could emerge.

Determination of measles hemagglutination inhibiting antibody levels among school children in Ibadan, Nigeria.

Immune status of school children aged from 10-23 years against measles virus was determined by the hemagglutination-inhibition (HI) test with a view of assessing herd immunity. Blood samples from 500 schoolchildren were collected by finger-pricking in Ropacco filter papers. Sera were extracted in 1 mL of cold phosphate buffered saline and treated with 25% (w/v) kaolin and 50.0% monkey red blood cell (RBC) to a final concentration of 1:10. The measles hemagglutinating antigen used for the test was prepared from measles vaccine. Results showed that 62 (12.4%) were positive for measles HI antibody at a titer of 1:640, and 78 (15.6%) had a titer <1:10. There was no significant relationship (P > 0.05) between antibody titer and the schools, while a significant relationship (P < 0.05) existed between antibody titer and age. Although the majority of the schoolchildren had the measles antibody in their sera, titers were, however, beneath the threshold of protection in 33.4% of them. The significant association between age of the schoolchildren and HI titers showed that those antibodies were waning according to age. The study has shown a considerable high level of protection against measles among schoolchildren. To prevent future outbreak of measles among these schoolchildren, it will be advocated that a second dose of measles vaccine be administered.

Effect of temperature and distance on the viral outcome of clinical specimens from acute flaccid paralysis cases in Nigeria.

AIM: The effect of cold box temperature and distance on virus titers, poliovirus isolation rate, and appearance of orphan polioviruses was investigated. METHODS: Cold boxes with stools were randomly selected and examined for internal temperature over a 7-month period. After virus isolation in Rhabdomyosarcoma (RD) cell line RDand L20B, titer calculations and intratypic differentiation were done on isolates. Sequencing and molecular studies were done on the isolates periodically in the order of arrival in the laboratory for a period of 30 months. RESULTS: Seventy-one (51.1%) boxes had the temperature range of 1 -4°C, 53 (38.1%) had 4.5 -8°C,. while 15 (10.8%) had temperature between 8.5°C and 17.0°C. Poliovirus was isolated from 24 (8.6%) specimens made up of 13 wild 1 and 2 and 11 Sabins 1, 2, 3 with titers between 10(1.8) and 10(5.4) TCID(50) /100 µl. Temperature and titer were inversely proportional and statistically significant. (r = -0.83, P < 0.05). Distance to laboratory was not significantly related (r = -0.025) to temperature when appropriate cold box temperature was maintained. Of the 18,188 acute flaccid paralysis (AFP) specimens received in the laboratory between June 2008 and December 2010, 1,752 poliovirus isolates (9.6%) consisting of 480 wild and 82 orphans were found. A positive correlation between the distance and orphan viruses (r = 0.425; P = 0.027) was observed. CONCLUSION: While poliovirus titer depends on the inside temperature of the cold box, distance to the laboratory was found to be a predisposing factor to the appearance of orphan viruses.

Outbreak of type 2 vaccine-derived poliovirus in Nigeria: emergence and widespread circulation in an underimmunized population.

Wild poliovirus has remained endemic in northern Nigeria because of low coverage achieved in the routine immunization program and in supplementary immunization activities (SIAs). An outbreak of infection involving 315 cases of type 2 circulating vaccine-derived poliovirus (cVDPV2; >1% divergent from Sabin 2) occurred during July 2005-June 2010, a period when 23 of 34 SIAs used monovalent or bivalent oral poliovirus vaccine (OPV) lacking Sabin 2. In addition, 21 "pre-VDPV2" (0.5%-1.0% divergent) cases occurred during this period. Both cVDPV and pre-VDPV cases were clinically indistinguishable from cases due to wild poliovirus. The monthly incidence of cases increased sharply in early 2009, as more children aged without trivalent OPV SIAs. Cumulative state incidence of pre-VDPV2/cVDPV2 was correlated with low childhood immunization against poliovirus type 2 assessed by various means. Strengthened routine immunization programs in countries with suboptimal coverage and balanced use of OPV formulations in SIAs are necessary to minimize risks of VDPV emergence and circulation.

Measles virus strain diversity, Nigeria and Democratic Republic of the Congo.

We investigated the genetic diversity of measles virus (MV) in Nigeria (2004-2005) and the Democratic Republic of the Congo (DRC) (2002-2006). Genotype B3 strains circulating in Kinshasa, DRC, in 2002-2003 were fully replaced by genotype B2 in 2004 at the end of the second Congo war. In Nigeria (2004-2005), two genetic clusters of genotype B3, both of which were most closely related to 1 variant from 1998, were identified. Longitudinal analysis of MV strain diversity in Nigeria suggested that only a few of the previously described 1997-1998 variants had continued to circulate, but this finding was concomitant with a rapid restoration of genetic diversity, probably caused by low vaccination coverage and high birth rates. In contrast, the relatively low genetic diversity of MV in DRC and the genotype replacement in Kinshasa reflect a notable improvement in local measles control.

Phylogenetics and pathogenesis of early avian influenza viruses (H5N1), Nigeria.

Three highly pathogenic avian influenza subtype H5N1 and 4 Newcastle disease viruses were isolated from sick or dead chickens in southwestern Nigeria. Sequencing and phylogenetic analysis placed them within H5N1 subclade 2.2.2. Intravenous and intranasal pathogenicity tests produced systemic disease with vascular endothelial cell tropism in chickens.

Isolation of recombinant type 2 vaccine-derived poliovirus (VDPV) from a Nigerian child.

A type 2 vaccine-derived poliovirus (VDPV), differing from Sabin 2 at 2.5% (22/903) of VP1 nucleotide (nt) positions, was isolated from an incompletely immunized 21-month-old Nigerian child who developed acute flaccid paralysis in 2002. Sequences upstream of nt position 620 (within the 5'-untranslated region [5'-UTR]) and downstream of nt position 5840 (in the 3C(pro) region) were derived from species C enteroviruses unrelated to the oral poliovirus vaccine (OPV) strains. The two substitutions associated with the attenuated phenotype had either recombined out (A(481)-->G in the 5'-UTR) or reverted (Ile(143)-->Thr in VP1). The VDPV isolate had lost the temperature sensitive phenotype of Sabin 2 and it was antigenically distinct from the parental OPV strain, having amino acid substitutions in or near neutralizing antigenic sites 1 and 3. The date of the initiating OPV dose, calculated from the number of synonymous substitutions in the capsid region, was estimated to be approximately 16 to 18 months before onset of paralysis, a finding inconsistent with the most recent mass OPV campaign (conducted 12 days before onset of paralysis) as being the source of infection. Although no related type 2 VDPVs were detected in Nigeria or elsewhere, the VDPV was found in an area where conditions favor VDPV emergence and spread.

Molecular characterization of diverse species enterovirus-B types from children with acute flaccid paralysis and asymptomatic children in Nigeria.

Non-polio enteroviruses (NPEVs) have often been identified in association with acute flaccid paralysis (AFP) in most polio surveillance studies worldwide. In a polio endemic country like Nigeria, there is need for distinction of AFP due to poliovirus and those potentially due to NPEVs. This study was undertaken to characterize the enterovirus (EV) types circulating in both children with and without AFP in Nigeria. Of fecal sample from 966 children with AFP, 96 (10%) were positive for NPEVs in RD cells, while 42 (5.5%) of 756 samples from non-AFP children was positive. Genotyping of all NPEV isolates was done by partial VP1 gene sequencing and phylogenetic analysis. EV-B was the predominant species detected (84%) and infection was common in children with AFP with CVB3, E6, and E11 constituting the predominant types detected. The CVB3 isolates cluster with Chinese CVB3 isolate recently detected in a newborn with AFP. There was also a remarkable clustering of isolates such as E6, E12, E13, E24, E30 and E33 to types previous detected in West Africa suggesting a probable circulation of these lineages in the region. Taken together, this study reveals a diverse species EV-B types in AFP cases and highlights the fact that other neurotropic EVs circulate in asymptomatic persons. Improved continuous surveillance of NPEV is warranted as in the likely attainment of polio eradication, other neurotropic EVs may emerge causing similar paralytic diseases.

Specific viruses detected in nigerian children in association with acute respiratory disease.

Occurrence of different viruses in acute respiratory tract infections of Nigerian children was examined. Respiratory swabs were collected from 246 children referred to hospital clinics because of acute respiratory symptoms from February through May 2009. Validated real-time RT-PCR techniques revealed nucleic acids of at least one virus group in 189 specimens (77%). Human rhinoviruses and parainfluenza viruses were present each in one third of the children. Adenoviruses, enteroviruses, human metapneumovirus, human bocavirus, and influenza C virus were also relatively common. Possibly due to their seasonal occurrence, influenza A and B virus, and respiratory syncytial virus were detected rarely. We conclude that all major groups of respiratory tract viruses are causing illness in Nigerian children.