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Evidence for the presence of African swine fever virus in apparently healthy pigs in South-Kivu Province of the Democratic Republic of Congo.

ABSTRACT: African swine fever (ASF) is the most important disease constraining smallholder pig production in the Democratic Republic of Congo, causing high mortality in domestic pigs with severe impacts on the livelihoods of local populations. This study was conducted with the aim of determining the prevalence of ASF and circulating virus genotypes in asymptomatic pigs raised on smallholder farms in the South Kivu province to understand the transmission dynamics of ASF and ultimately improving disease control. A cross-sectional survey was carried out in 5 districts where 267 pig blood were screened for both antibody and viral genome using indirect Enzyme Linked Immunosorbent Assay (ELISA) and polymerase chain reaction (PCR) respectively. Additionally, amplicons from PCR positive samples were sequenced by Sanger method for genetic analysis of ASF virus (ASFV) based on the C-terminal region of the B646L gene that encodes the major capsid protein p72 and the gene E183L encoding the p54 protein. The overall seroprevalence obtained based on antibody to p30 protein was 37 % and was significantly higher (P < 0.05) in adult (>1 year) animals (44.7 %) than in younger (<1 year) ones (33.5 %). Moreover, the seropositivity varied significantly (P < 0.05) according to the pig husbandry system practiced within the districts investigated with Uvira (55 %) and Mwenga (42.2 %) having the highest ASFV antibodies, while the lowest (10.5 %) were in Kalehe. Free-range pigs exhibited a higher level of seropositivity to ASFV antibody (68.9 %) than pigs kept in the pigsty housing system (21.6 %). However, no statistically significant differences (P > 0.05) were observed when sex of the animal and breed were factored. PCR detection of ASFV amplified a specific band of expected size (257 bp) in 61 out of 267 blood samples, confirming the presence of the viral DNA in 22.8 % of asymptomatic domestic pigs. Statistical analysis revealed that ASFV infection in domestic pigs varied significantly (p < 0.001) according to geographical location and breed, with the highest infection rate found in Walungu district (33.7 %) while the lowest was registered in Kalehe (15.8 %). Local pigs (27.2 %) were more infected than crosses (9.2 %). Phylogenetic analyses based on partial sequences of the p72 and p54 genes revealed that all the ASFV detected belonged to genotype IX, which has previously been reported in other parts of DR Congo, and was clustered together with isolates from Kenya, Uganda and Republic of Congo. This study avails the first evidence of the presence of ASF virus in domestic pigs in the absence of outbreaks in South Kivu province, eastern DR Congo, indicating a need to raise awareness among pig farmers and veterinary authorities on the application of biosecurity measures and good husbandry practices to control the disease.

SUBMITTER: Patrick BN

PROVIDER: S-EPMC7045278 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundAfrican swine fever (ASF) is a highly contagious and severe hemorrhagic viral disease of domestic pigs. The analysis of variable regions of African swine fever virus (ASFV) genome led to more genotypic and serotypic information about circulating strains. The present study aimed at investigating the genetic diversity of ASFV strains in symptomatic pigs in South Kivu province of the Democratic Republic of Congo (DRC).Materials and methodsBlood samples collected from 391 ASF symptomatic domestic pigs in 6 of 8 districts in South Kivu were screened for the presence of ASFV, using a VP73 gene-specific polymerase chain reaction (PCR) with the universal primer set PPA1-PPA2. To genotype the strains, we sequenced and compared the nucleotide sequences of PPA-positive samples at three loci: the C-terminus of B646L gene encoding the p72 protein, the E183L gene encoding the p54 protein, and the central hypervariable region (CVR) of the B602L gene encoding the J9L protein. In addition, to serotype and discriminate between closely related strains, the EP402L (CD2v) gene and the intergenic region between the I73R and I329L genes were analyzed.ResultsASFV was confirmed in 26 of 391 pigs tested. However, only 19 and 15 PPA-positive samples, respectively, were successfully sequenced and phylogenetically analyzed for p72 (B646L) and p54 (E183L). All the ASFV studied were of genotype X. The CVR tetrameric repeat clustered the ASFV strains in two subgroups: the Uvira subgroup (10 TRS repeats, AAAABNAABA) and another subgroup from all other strains (8 TRS repeats, AABNAABA). The phylogenetic analysis of the EP402L gene clustered all the strains into CD2v serogroup 7. Analyzing the intergenic region between I73R and I329L genes revealed that the strains were identical but contained a deletion of a 33-nucleotide internal repeat sequence compared to ASFV strain Kenya 1950.ConclusionASFV genotype X and serogroup 7 was identified in the ASF disease outbreaks in South Kivu province of DRC in 2018-2019. This represents the first report of ASFV genotype X in DRC. CVR tetrameric repeat sequences clustered the ASFV strains studied in two subgroups. Our finding emphasizes the need for improved coordination of the control of ASF.

Project description:IntroductionIn the field of tuberculosis (TB), and particularly in regard to paediatric TB (PedTB), clinical skills of health professionals play an important role in determining quality of care. In an era where novel diagnostic technologies and efficient treatment regimens are being made available for the poorest, we must not divert our attention from the importance of clinical skills, as this deliverable remains the cornerstone of individualized patient care and ultimately the best assurance for optimal use of resources. The aim of our work was to study the epidemiology of PedTB and the determinants of PedTB under-detection in the South-Kivu Province of the Democratic Republic of Congo (DRC), a setting with nearly no technical resources allowing to support the clinical diagnosis of PedTB, i.e. chest X-rays, rapid molecular tests or culture laboratories.MethodsWe collected TB notification data from 2010 to 2015 and analysed the space-time variations in notification for the different forms of TB among the 113 health facilities (HF) the South-Kivu Province, a region with a low HIV incidence. The different forms of TB notified were: smear positive pulmonary TB (SS+PTB), smear negative pulmonary TB (SS-PTB) and extra-pulmonary TB (EPTB). We further analysed the distribution of these different forms of the disease per age group and explored the possibility to predict the detection of PedTB.ResultsSignificant differences were observed between HF in regard to the proportion of paediatric TB and the proportion of SS-TB among adults. We found a strong correlation between the proportion of PedTB and three major factors: the proportion of TB cases with no bacteriological confirmation (SS-TB) among adults, the number of TB cases notified by the HF and the fact that the HF was supported or not by Médecins Sans Frontières (MSF). The proportion of SS-TB among adults was found to be a valid indicator for predicting the level of detection of PedTB at the same HF.ConclusionOur observations strongly suggest that under-detection of PedTB is associated with insufficient clinical skills and technical resources at the HF level which similarly affects other forms of the disease, in particular SS-TB. We demonstrated that, in the specific context of South-Kivu, under-detection of PedTB can be predicted by a low SS-TB/SS+PTB ratio in the adult population. In the context of severely under-resourced settings, this ratio could be used to rapidly identify HF that should benefit in priority from deeper evaluation, and eventually targeted interventions.

Project description:BackgroundAfrican swine fever (ASF), a highly contagious hemorrhagic disease, affects domestic pigs in the Democratic Republic of Congo (DRC) where regular outbreaks are reported leading to high mortality rates approaching 100% in the affected regions. No study on the characteristics of the complete genome of strains responsible for ASF outbreaks in the South Kivu province of DRC is available, limited a better understanding of molecular evolution and spread of this virus within the country. The present study aimed at determining the complete genome sequence of ASFV strains genotype X involved in 2018-2019 ASF disease outbreaks in South Kivu province of DRC.Materials and methodsGenomic DNA of a spleen sample from an ASFV genotype X-positive domestic pig in Uvira, during the 2018-2019 outbreaks in South Kivu, was sequenced using the Illumina HiSeq X platform. Obtained trimmed reads using Geneious Prime 2020.0.4 were blasted against a pig reference genome then contigs were generated from the unmapped reads enriched in ASFV DNA using Spades implemented in Geneious 2020.0.4. The assembly of the complete genome sequence of ASFV was achieved from the longest overlapping contigs. The new genome was annotated with the genome annotation transfer utility (GATU) software and the CLC Genomics Workbench 8 software was further used to search for any ORFs that failed to be identified by GATU. Subsequent analyses of the newly determined Uvira ASFV genotype X genome were done using BLAST for databases search, CLUSTAL W for multiple sequences alignments and MEGA X for phylogeny.Results42 Gbp paired-end reads of 150 bp long were obtained containing about 0.1% of ASFV DNA. The assembled Uvira ASFV genome, termed Uvira B53, was 180,916 bp long that could be assembled in 2 contigs. The Uvira B53genome had a GC content of 38.5%, encoded 168 open reading frames (ORFs) and had 98.8% nucleotide identity with the reference ASFV genotype X Kenya 1950. The phylogenetic relationship with selected representative genomes clustered the Uvira B53 strain together with ASFV genotype X reported to date (Kenya 1950 and Ken05/Tk1). Multiple genome sequences comparison with the two reference ASFV genotype X strains showed that 130 of the 168 ORFs were fully conserved in the Uvira B53. The other 38 ORFs were divergent mainly due to SNPs and indels (deletions and insertions). Most of 46 multigene family (MGF) genes identified were affected by various genetic variations. However, 8 MGF ORFs present in Kenya 1950 and Ken05/Tk1 were absent from the Uvira B53 genome including three members of MGF 360, four of MGF 110 and one of MGF 100 while one MGF ORF (MGF 360-1L) at the left end of the genome was truncated in Uvira B53. Moreover, ORFs DP96R and p285L were also absent in the Uvira B53 genome. In contrast, the ORF MGF 110-5L present in Uvira B53 and Ken05/Tk1 was missing in Kenya 1950. The analysis of the intergenic region between the I73R and I329L genes also revealed sequence variations between the three genotype X strains mainly characterized by a deletion of 69 bp in Uvira B53 and 36 bp in Kenya 1950, compared to Ken05/Tk1. Assessment of the CD2v (EP402R) antigen unveiled the presence of SNPs and indels particularly in the PPPKPY tandem repeat region between selected variants representing the eight serogroups reported to date. Uvira B53 had identical CD2v variable region to the Uganda (KM609361) strain, the only other ASFV serogroup 7 reported to date.ConclusionWe report the first complete genome sequence of an African swine fever virus (ASFV) p72 genotype X and CD2v serogroup 7, termed Uvira B53. This study provides additional insights on genetic characteristics and evolution of ASFV useful for tracing the geographical spread of ASF and essential for improved design of control and management strategies against ASF.

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Evidence for the presence of African swine fever virus in apparently healthy pigs in South-Kivu Province of the Democratic Republic of Congo.

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