Epidemiology of peste-des-petits-ruminants virus in sheep goats and camels in Nigeria

Abstract

Peste-des-petits-ruminants (PPR), caused by the peste-des-petits ruminants virus (PPRV) is endemic in Nigeria and is a major constraint to the improvement of small ruminant production, thereby affecting the rural poor who depends on these livestock species as their source of livelihoods. The major objectives of this work were: 1) to conduct a seroprevalence of PPRV in camels, sheep and goats in Nigeria; 2) to isolate PPRV from naturally infected animals; and 3) to characterize the isolates using molecular techniques and compare the profile of the current isolates with the worldwide vaccine (Nig75/1) and other existing strains from other parts of the world. The major contribution of this work to current knowledge were: finding that more than one lineage of PPRV was circulating in Nigeria, the emerging Asian lineage IV gradually replacing the traditional lineage II in the country; the isolation in cell cultures and full genome sequencing of current field viruses which was last done in 1976; the unique amino acid residues in the individual proteins of the various lineages of PPRV were identified and used to develop a lineage-specific diagnostic test for the virus; and the current seroprevalence report of PPR in camels, sheep and goats from all the agro-ecological zones of the country. This work has also increased the sequences of PPRV available in GenBank. A total of 6,065 field serum samples from camels (1,517), goats (3,489) and sheep (1,059) were collected from all the six agro-ecological zones of Nigeria. The study subjects cut across all ages and sexes. The samples were subjected to both the N and H protein based c-ELISA. In addition, 140 clinical samples from 16 sheep and 63 goats with symptoms suggestive of PPRV infection were collected from different (15) states of Nigeria during a four year period (2010 2013). Published primers were used to amplify a 351 bp segment of the PPRV nucleoprotein (N) gene and a 370 bp segment of the fusion (F) gene. Monkey CV 1 cell line expressing the sheep-goat SLAM protein was used to isolate PPRV from RT-PCR positive samples. Thirty primer pairs were used to amplify and sequence two full genome isolates of the current circulating PPRV in Nigeria. Primer express software was used to develop a lineage-specific real time PCR for PPRV. The overall prevalence estimate of serum positive results for PPRV in sheep and goats was 23.16 % (n = 1,018/4,548, 95 % confidence interval [CI]: 21.79 - 24.57. There were significant differences in prevalences between the states (p = 0.001), zones (p = 0.058) and age (p = 0.032). Taraba State had the highest seroprevalence rate of 27.97 % while the lowest rate of 14.76 % was observed in Cross River State. There were no significant differences in the PPRV prevalences between male and female animals (p = 0.571) and between species (p = 0.639). The overall prevalence in camels was 3.36 % (51/1517, 95 % CI: 2.51 4.39). There were no significant differences in prevalences between states (p = 0.892) and between male and female camels (p = 0.742). The prevalence differed significantly (p < 0.001) by body condition scores - camels with poor body condition score have a higher (16.67 %) antibody seroprevalences to PPRV compared to those with fair and good body condition scores. There was a statistically significant difference between camels aged ? 5 years and those > 5 years (p = 0.004). The clinical samples subjected to RT-PCR showed that 33 (42 %) animals were positive for PPRV nucleic acid, which included two sheep (13 %) and 31 goats (49 %). The amplicons were sequenced and phylogenetic analysis using the neighbour joining, maximum parsimony and Bayesian analysis of the sequences with those available in GenBank showed that isolates from the current study belonged to lineage II and lineage IV. The lineage II viruses from this study grouped into two clades, one closely related to the vaccine virus (Nigeria75/1) and the other clade group with other wild-type viruses from Mali, Senegal and Sierra Leone. The lineage IV isolates also grouped into two sub-clades, one closely related to a 2011 strain from Gabon and the other closely related to a 1997 strain from Cameroon. Analysis of two full genomes of PPRV from Nigeria representing the two lineages of the virus circulating currently in the country showed that the lineage II representative isolated in 2012 has an identity of 96 % at the nucleotide level with the lineage II Nigeria75/1 virus isolated in 1975 and used as a vaccine. The lineage IV representative isolated in 2013 revealed an identity of 96 % with Sungri/96 from India. The unique amino acid residues in the individual proteins of the various lineages of PPRV were identified and used to develop a one-step TaqMan® real-time RT-PCR assay targeting the H gene of PPRV. This study reports for the first time, the presence of PPRV lineage IV, the Asian lineage in Nigeria. As part of the study, a lineage-specific assay was also developed, which once validated, could be included in the panel of assays recommended by the World Organization for Animal Health (OIE). The seroprevalence studies showed occasional transient PPRV infection of camels and the current antibody seroprevalence to PPRV in the small ruminants population in Nigeria. There is the need to include camels among species to be studied in elucidating the epidemiology of the disease in sheep and goats. The seroprevalence studies may be helpful in reaching a decision on the vaccination strategy for the control of the disease in the country.