Diversity and emergence of new variants of African swine fever virus Genotype I circulating in domestic pigs in Nigeria (2016-2018).

BACKGROUND: African swine fever (ASF) is the most lethal disease of pigs caused by ASF virus (ASFV) with severe economic implications and threat to the swine industry in endemic countries. Between 2016 and 2018, several ASF outbreaks were reported throughout pig producing states in Nigeria. OBJECTIVES: Thereafter, this study was designed to identify the ASFV genotypes responsible for these outbreaks within the study period (2016-2018). METHODS: Twenty-two ASFV-positive samples by polymerase chain reaction were selected. The samples were collected during passive surveillance in eight states of Nigeria were characterised using 3 partial genes sequences of the virus namely, p72 capsid protein of the B646L, p54 envelope protein of E183L and the central variable region (CVR) within B602L of ASFV. RESULTS: Phylogenetic and sequences analysis based on p72 and p54 revealed ASFV genotype I as the circulating virus. Sequence analysis of the CVR of B602L revealed genetic variations with six ASFV tandem repeat sequence (TRS) variants namely, Tet-15, Tet-20a, Tet-21b, Tet-27, Tet-31 and Tet-34, thus increasing the overall genetic diversity of ASFV in Nigeria. Three of the TRS variants, Tet-21b, Tet-31 and Tet-34, were identified for the first time in Nigeria. The new TRS variants of ASFV genotype I were identified in Enugu, Imo, Plateau and Taraba states, while co-circulation of multiple variants of ASFV genotype I was recorded in Plateau and Benue states. CONCLUSIONS: The high genetic diversity, emergence and increasing recovery of new variants of genotype I in Nigeria should be a concern given that ASFV is a relatively stable DNA virus. The epidemiological implications of these findings require further investigation.

Infectious bursal disease in Nigeria: continuous circulation of reassortant viruses.

Outbreaks of infectious bursal disease (IBD), a highly contagious immunosuppressive disease of young chickens, are still reported globally despite vaccination efforts. This study investigated the genetic characteristics of infectious bursal disease virus (IBDV) from 26 reported outbreaks in 2019 in Nigeria. Nucleotide sequences of VP2 hypervariable (hvVP2) region (n=26) and VP1 (n=23) of Nigerian IBDVs were determined. Our results revealed the detection of reassortant strains with segment A related to very virulent IBDV (vvIBDV) having virulence marker (222A, 242I, 256I, 294I and 299S), whereas their segment B were closely related to previously detected IBDV strains having QEG substitution at positions 145-147. Phylogenetic analysis of the hvVP2 region revealed that all the Nigerian IBDV clustered with vvIBDV (genogroup 3) and were independent of the Asian/European lineage. Interestingly, in the hvVP2, all the viruses had a G-S substitution at residue 254. Additionally, one isolate had an A321T substitution at the PHI loop, which has been suggested to play a key role in antigenicity. Four of the viruses (Bauchi=3 and Plateau=1) had a unique A-T substitution at residue 144 on the VP1 region. We also observed a T174S substitution in nine of the Nigerian viruses from Bauchi and Plateau state that were not found in any outbreak viruses from Oyo and Akwa Ibom. This report demonstrates the circulation of reassortant strains in commercial and backyard poultry farms in Nigeria despite sustained vaccination efforts. Our data suggest that the Nigerian outbreak viruses have mutations that may affect antigenicity and contribute to antigenic drift.

Genetic analysis and emergence of canine parvovirus type 2c in South Eastern Nigeria.

BACKGROUND: Genetic analysis of canine parvovirus type 2 (CPV-2) variants circulating in South Eastern Nigeria was investigated. The original strain of CPV-2 emerged in 1978, mutated later to CPV-2a and has continued to be evolved. AIMS: To genetically characterize CPV-2 strains detected in dogs in South Eastern Nigeria and to phylogenetically group the viruses with existing sequencing data. METHODS: A total number of 82 rectal swabs were collected and stored in virus transport medium (VTM) from suspected cases of CPV-2 within the study area and were tested with polymerase chain reaction (PCR). RESULTS: Seventy-nine samples (96.3%) were positive for CPV-2 and sequence analysis of partial VP2 gene of 20 amplicons revealed circulation of CPV-2a (n=4) and CPV-2c (n=16) in the region. The obtained strains clustered together. However, the group was further divided into two clear clusters comprising of 2a and 2c strains. The vaccine strain and the CPV-2 reference strains from USA formed a monophyletic cluster. CONCLUSION: Canine parvovirus types 2a and 2c are co-circulating in South Eastern region of Nigeria and therefore, there is an urgent need for an improved vaccine to cover for the emerging strain (CPV-2c) in Nigeria.

Molecular detection of rabies virus strain with N-gene that clustered with China lineage 2 co-circulating with Africa lineages in Monrovia, Liberia: first reported case in Africa.

Despite a long history of dog-transmitted human rabies outbreaks in Liberia, West Africa, no reports exist of molecular characterisation of the causative lyssaviruses. This study investigated Rabies lyssavirus (RABV) strains isolated at the dog-human interface in Monrovia, Liberia 2016 and 2017, by reverse transcription polymerase chain reaction, using primers specific for the nucleoprotein (N) gene. Out of 20 specimens (19 dog brain samples and one human saliva) tested as suspected rabies cases, three (15%) were positive. Purified amplicons from all three positive specimens were sequenced in both forward and reverse directions. Phylogenetic analysis was conducted in MEGA7 and PhyML3 to determine their relationship with RABV sequences accessioned in NCBI GenBank. The first of three RABV strains detected clustered with China lineage 2 RABVs of dogs (99% homology to KU963489 and DQ666322). The second strain segregated with Africa lineage 2 RABVs also of dog origin, and the third strain segregated with Africa lineage 3 RABVs of Southern Africa viverrids. Our results show a transcontinental strain of rabies virus co-circulating with Africa lineages in post-conflict Liberia. This finding should stimulate more effective sub-regional planning and execution of one-health actions, towards stepwise surveillance and elimination of rabies in West Africa by 2030.

Application of loop-mediated isothermal amplification assay in the detection of herpesvirus of turkey (FC 126 strain) from chicken samples in Nigeria.

AIM: This study was designed to optimize and apply the use of loop-mediated isothermal amplification (LAMP) as an alternative to conventional polymerase chain reaction (PCR) for the detection of herpesvirus of turkeys (HVT) (FC 126 strain) in vaccinated and non-vaccinated poultry in Nigeria. MATERIALS AND METHODS: HVT positive control (vaccine) was used for optimization of LAMP using six primers that target the HVT070 gene sequence of the virus. These primers can differentiate HVT, a Marek's disease virus (MDV) serotype 3 from MDV serotypes 1 and 2. Samples were collected from clinical cases of Marek's disease (MD) in chickens, processed and subjected to LAMP and PCR. RESULTS: LAMP assay for HVT was optimized. HVT was detected in 60% (3/5) and 100% (5/5) of the samples analyzed by PCR and LAMP, respectively. HVT was detected in the feathers, liver, skin, and spleen with average DNA purity of 3.05-4.52 µg DNA/mg (A260/A280) using LAMP. Conventional PCR detected HVT in two vaccinated and one unvaccinated chicken samples, while LAMP detected HVT in two vaccinated and three unvaccinated corresponding chicken samples. However, LAMP was a faster and simpler technique to carry out than PCR. CONCLUSION: LAMP assay for the detection of HVT was optimized. LAMP and PCR detected HVT in clinical samples collected. LAMP assay can be a very good alternative to PCR for detection of HVT and other viruses. This is the first report of the use of LAMP for the detection of viruses of veterinary importance in Nigeria. LAMP should be optimized as a diagnostic and research tool for investigation of poultry diseases such as MD in Nigeria.

Genetic Characterization of Circulating African Swine Fever Viruses in Nigeria (2007-2015).

Sequencing and analysis of three discrete genome regions of African swine fever viruses (ASFV) from archival samples collected in 2007-2011 and active and passive surveillance between 2012 and 2015 in Nigeria were carried out. Analysis was conducted by genotyping of three single-copy African swine fever (ASF) genes. The E183L and B646L genes that encode structural proteins p54 and p72, respectively, were utilized to delineate genotypes before intragenotypic resolution by characterization of the tetrameric amino acid repeat region within the hypervariable central variable region of the B602L gene. The results showed no variation in the p72 and p54 gene regions sequenced. Phylogeny of p72 sequences revealed that all the Nigerian isolates belonged to genotype I, while that of the p54 recovered the Ia genotype. Analysis of B602L gene revealed the differences in the number of tetrameric repeats. Four new variants (Tet-15, Tet-17a, Tet-17b and Tet-48) were recovered, while a fifth variant (Tet-20) was the most widely distributed in the country displacing Tet-36 reported previously in 2003-2006. The viruses responsible for ASF outbreaks in Nigeria are from very closely related but mutated variants of the virus that have been circulating since 1997. A practical implication of the genetic variability of the Nigerian viral isolates in this study is the need for continuous sampling and analysis of circulating viruses, which will provide epidemiological information on the evolution of ASFV in the field versus new incursion for informed strategic control of the disease in the country.

Spatio-temporal dynamics of African swine fever outbreaks in Nigeria, 2002-2007.

African swine fever (ASF) was first introduced into Nigeria through Lagos state in 1997. The disease rapidly spread to Ogun state in 1998 and extended to the Niger Delta (Delta, Rivers and Akwa Ibom states) in the same year. In 1998, Kaduna, Plateau and Benue states all north of the country experienced ASF for the first time. Poor farm biosecurity, bad abattoir practices and extensive/free range pig farming systems led to extensive spread of the diseases to about 16 Nigerian states excluding the far northwest and north east. A total of 1036 field samples collected over a 6-year period covering 19 Nigerian states were analysed during the period under review; 805 samples were PCR positive and 231 negative. Positive samples were detected in all three surveillance phases and from all agroecological zones across the country. For the first time since its incursion, ASF was identified in some states; Bauchi, Adamawa Taraba and Gombe with chances of control very slim and further spread of the virus northward envisaged. Outbreaks of the disease are now a perennial problem with an increasing disease burden in areas where high numbers of pigs are produced in the country. The National Veterinary Research Institute (NVRI), Vom, since 2002 investigated ASF based on tissue submissions and reports made by individuals, private & commercial farms and agricultural bodies. We present an analysis of geographical and temporal distribution of ASF in the country from 2002 to 2007 and a review of historic outbreaks since the first incursion. Risk factors and prospects for control are discussed.