Molecular characterization of canine parvovirus strains from domestic dogs in South Africa and Nigeria

Canine parvovirus type 2 (CPV-2), the aetiological agent of haemorrhagic enteritis in dogs emerged in 1978 worldwide. In the mid 1980’s, the original CPV-2 had evolved and was completely replaced by 2 variants, CPV 2a and 2b. In 2000, a new variant of CPV (CPV-2c) was detected in Italy and now circulates in other countries. Haemorrhagic enteritis in dogs is a major disease in South Africa and Nigeria. Both infection rates with CPV-2 and case fatality rates in young dogs are high. CPV-2 is a small, negative-sense, single-stranded DNA virus of 5.2kb long and a member of the Parvoviridae family, which also includes feline panleukopenia virus (FPV) and mink enteritis virus (MEV). The CPV-2 genome is prone to mutations at the VP2-encoding region. As a result we investigated the genetic composition of the VP2 region in the CPV-2 genome using molecular methods (qPCR) to provide information for comparison of field and vaccine strains of the virus. The conventional PCR detection results yielded 137 (97.85%) of the total of 140 feacal samples screened with diarrhoea positive. One hundred-and-six of 108 samples from South Africa (98.15%) tested positive and two (1.85%) were negative, while 30 (96.77%) from 31 faecal samples from Nigeria were positive and 1 (2.23%) was negative. Results obtained from the genotyping of the CPV- 2 strains using CPV-2a/b and CPV-2b/c TaqMan assays employing minor groove binder (MGB) probes, revealed that out of a total of 106 South African samples, 100 (94.34%) were infected with CPV-2b and 6 (5.66%) with CPV-2a, while all the Nigerian samples [n=30 (100%)] contained only CPV2a. There was no reported case of CPV-2c. The VP2 gene of selected DNA samples (n=27), from South Africa (n=19), Nigeria (n=6) and multivalent vaccines (n=2) were amplified and sequenced. These sequences were originally aligned and edited to a total length of 1,750 bp of the CPV-2 VP2 encoding gene. These selected sequences showed 99% maximum identity to the GenBank sequences from the blast results (NCBI BLASThttp:// www.ncbi.nlm.nih.gov/BLAST/) and alignment of all the sequences was performed using ClustalX. Two phylogenetic analyses showed most South African field isolates distant from viruses from other parts of the world. A few clustered with Asian and European strains, while Nigerian CPV-2 strains clustered with USA and some European isolates. The results of the protein analysis showed seven changes of amino acids at positions 265, 297, 324, 424, 426, 440 and 475 for most of the South Africans strains while the Nigerian CPV-2 had only one field isolate with an amino acid change.Dissertation (MSc)--University of Pretoria, 2010.Veterinary Tropical Diseasesunrestricte

Sequencing and Analysis of Lumpy Skin Disease Virus Whole Genomes Reveals a New Viral Subgroup in West and Central Africa

Lumpy skin disease virus (LSDV) is a member of the capripoxvirus (CPPV) genus of the Poxviridae family. LSDV is a rapidly emerging, high-consequence pathogen of cattle, recently spreading from Africa and the Middle East into Europe and Asia. We have sequenced the whole genome of historical LSDV isolates from the Pirbright Institute virus archive, and field isolates from recent disease outbreaks in Sri Lanka, Mongolia, Nigeria and Ethiopia. These genome sequences were compared to published genomes and classified into different subgroups. Two subgroups contained vaccine or vaccine-like samples ("Neethling-like" clade 1.1 and "Kenya-like" subgroup, clade 1.2.2). One subgroup was associated with outbreaks of LSD in the Middle East/Europe (clade 1.2.1) and a previously unreported subgroup originated from cases of LSD in west and central Africa (clade 1.2.3). Isolates were also identified that contained a mix of genes from both wildtype and vaccine samples (vaccine-like recombinants, grouped in clade 2). Whole genome sequencing and analysis of LSDV strains isolated from different regions of Africa, Europe and Asia have provided new knowledge of the drivers of LSDV emergence, and will inform future disease control strategies.</p

Prevalence and characterization of Leptospira spp. in slaughter animals at abattoirs in Gauteng, South Africa and the zoonotic risk posed to abattoir workers

Leptospirosis is an important global re-emerging, occupational, environmental and zoonotic disease. It is an under-estimated disease of public health and veterinary importance caused by the pathogenic spirochetes belonging to the genus Leptospira. Currently in South Africa, there is limited information on leptospirosis and veterinarians’ beliefs that leptospirosis is not an important disease in the country. The primary aim of the investigation was to determine the prevalence of Leptospira spp. in slaughtered livestock and workers at abattoirs in Gauteng province, South Africa. To achieve this aim, retrospective analysis of laboratory data and cross-sectional serological, bacteriological and molecular studies were conducted on livestock and abattoir workers during the study period. The objective of the retrospective analysis of 11-year (2007 – 2017) data was to determine the seropositivity and infecting serovars of Leptospira in the sera of livestock (suspected or clinical cases of leptospirosis), submitted to the Agricultural Research Council (ARC)-Ondersterpoort Veterinary Research (OVR), Bacteriology serology laboratory. The overall seropositivity for leptospirosis in livestock was, 20.5% (1,425/6,945), using an eight-serovar microscopic agglutination test (MAT) panel. The frequency of seropositivity was 22.0% (1,133/5,168), 16.2% (286/1,763) and 0.0% (0/14) for cattle, pigs and sheep respectively (p<0.00 01). Australis (sv. Bratislava) was the predominant serovar having been detected in 29.4% (333/1,133) and 32.0% (91/286) of seropositive cattle and pigs respectively. The year 2016 of the 11 years retrospective data, had seroprevalence overall of 22.0% (102/466), with 100% (2/2) and 21.6% (101/466) for pigs and cattle respectively. It is important to note that, this was the same period (2016) we conducted the current cross-sectional study. A cross-sectional study was conducted on pigs and cattle slaughtered at Gauteng abattoirs in South Africa: Eighty-five (n=85) sera from slaughtered pigs at 5 consented abattoirs were analysed by MAT. The overall seropositive was, 24.7% (21/85) using 26 antigens panel for pigs in South Africa for the first time; Predominant serogroup was serogroup Australis-Bratislava reported as the predominant in seropositive pigs, 90.5% (19/21), 22.4% (19/85). For the cattle, 199 serum samples were analysed from slaughtered cattle from 11 abattoirs that consent was granted. Seropositive from cattle sera, 27.6% (55/199) with serogroup Sejroe (Hardjo), 10.5% (21/199) as the predominant circulating in the Country. The study demonstrated, for the first time in South Africa, the occurrence of four serovars, namely, Hardjo bovis strain lely 607; Topaz, 3.5% (7/199); Hebdomadis, 2.5% (5/199) and Medanensis, 1.5% (3/199) in slaughtered cattle. The vaccine used to prevent cattle leptospirosis in South Africa does not contain three of the newly detected serovars (Topaz, Hebdomadis and Medensis), an indication that the seropositive cattle acquired infection through natural exposure. There were statistically significant differences (P<0.05) in the detection of the serogroups of Leptospira. Of the five variables analysed, only one variable (abattoir) had statistically significantly (P<0.001) differences in the seroprevalence of leptospirosis in cattle. With the bacteriological culture of 305 kidneys using Ellinghausen McCaullough Johnson Harris (EMJH) media, the isolation rate for Leptospira spp. was 3.9% (12/305), with species-rate being 4.8% (9/186), 4.1% (3/74) and 0.0% (0/45) for cattle, pigs and sheep respectively (P>0.05). The use of quantitative polymerase chain reaction (qPCR) assays detected Leptospira DNA in 27.5% (84/305) of the livestock kidneys tested. Of the animals tested, 26.9% (50/186), 20.3% (15/74) and 42.2% (19/45) of cattle, pigs and sheep kidneys respectively (P=0.03) were positive for Leptospira DNA. It was significant that, although all sheep samples tested for leptospirosis by isolation and serology were negative for Leptospira spp., a high frequency (42.2%) was positive for Leptospira DNA. Sequencing of DNA from isolates of Leptospira spp. and kidney tissues from cattle identified 13 as L. interrogans and 2 as L. borgpetersenii), from pigs 4 were L. interrogans and from sheep kidney tissues, 2 were L. interrogans and 1 was L. borgpetersenii. The phylogenetic tree analyses revealed that all the isolates and the kidney tissue samples grouped together with the pathogenic L. interrogans serovar Icterohaemorrhagiae and L. borgpetersenii serovar Hardjo bovis strain lely 607 from the GenBank retrieved sequences. This study is also the first reported genetic analyses of the pathogenic L. interrogans and L. borgpetersenii, in slaughtered livestock in South Africa. To determine the exposure experience of Leptospira spp. in abattoir workers sampled from six abattoirs, two serological tests (MAT and IgM ELISA) and one molecular method (qPCR) were used. The seroprevalence of Leptospira in 103 workers was 10.7% and 7.8% by IgM ELISA and MAT respectively, and the prevalence of Leptospira DNA in whole blood by qPCR was 16.5% (P>0.05). The overall prevalence (serology and PCR) of Leptospira spp. was 30.1% (31/103). The predominant serovar detected in seropositive workers was Djasiman (50.0%) and the abattoir-related risk factors identified were working in high throughput (HT) abattoirs and exposure to blood and/or water splashes during and after slaughter. Antibodies to Serogroups sejroe (Sv. Wolffi) and Pomona (Sv. Djasiman) were both found in animals and abattoir workers. Although, the main serovars in abattoir workers were different from those in animals. It was concluded that the detection of new serovars Leptospira spp. in South Africa which are not currently in the leptospirosis vaccine used in livestock coupled with the fact that these serovars are not in the diagnostic eight-antigen MAT panel indicate a need to re-assess the status of livestock leptospirosis, as well as to revisit the existing policy and practices on leptospirosis in the country. The use of a diagnostic strategy which included both serological and molecular methods will increase the sensitivity of such an approach. The zoonotic risk of leptospirosis to abattoir workers identified in the study is for the first time in South Africa and it indicates the need to introduce measures to mitigate abattoir-associated risk exposure to leptospirosis in abattoir workers in the country.Thesis (PhD (Veterinary Science))--University of Pretoria, 2020.Veterinary Tropical DiseasesPhD (Veterinary Science)Unrestricte

Determinants of mortality among patients with drug-resistant tuberculosis in northern Nigeria.

BACKGROUND:Drug-Resistant tuberculosis (DR-TB) is estimated to cause about 10% of all TB related deaths. There is dearth of data on determinants of DR-TB mortality in Nigeria. Death among DR-TB treated cohorts in Nigeria from 2010 to 2013 was 30%, 29%, 15% and 13% respectively. Our objective was to identify factors affecting survival among DR-TB patients in northern Nigeria. METHODS:Demographic and clinical data of all DR-TB patients enrolled in Kano, Katsina and Bauchi states of Nigeria between 1st February 2015 and 30th November 2016 was used. Survival analysis was done using Kaplan-Meier and multiple regression with Cox proportional hazard modeling. RESULTS:Mean time to death during treatment is 19.2 weeks and 3.9 weeks among those awaiting treatment. Death was recorded among 38 of the 147 DR-TB patients assessed. HIV co-infection significantly increased probability of mortality, with an adjusted hazard ratio (aHR) of 2.35, 95% CI: 1.05-5.29, p = 0.038. Treatment delay showed significant negative association with survival (p = 0.000), not starting treatment significantly reduced probability of survival with an aHR of 7.98, 95% CI: 2.83-22.51, p = 0.000. Adjusted hazard ratios for patients started on treatment more than eight weeks after detection or within two to four weeks after detection, was beneficial though not statistically significant with respective p-values of 0.056 and 0.092. The model of care (facility vs. community-based) did not significantly influence survival. CONCLUSION:Both HIV co-infected DR-TB patients and DR-TB patients that fail to start treatment immediately after diagnosis are at significant risk of mortality. Our study showed no significant difference in mortality based on models of care. The study highlights the need to address programmatic and operational issues pertaining to treatment delays and strengthening DR-TB/HIV co-management as key strategies to reduce mortality

Seroprevalence and characterization of Brucella species in cattle slaughtered at Gauteng abattoirs, South Africa

BACKGROUND: Brucellosis is an infectious and contagious zoonotic bacterial disease of both humans and animals. In developing countries where brucellosis is endemic, baseline data on the prevalence of brucellosis, using abattoir facilities, is important. OBJECTIVES: The aim of this study was to determine the seroprevalence of antibodies against Brucella in slaughter cattle at Gauteng province, South Africa and to charac-terize isolates of Brucella spp. METHODS: In this cross-sectional study, un-clotted blood samples with correspond-ing organ tissue samples were collected from slaughtered cattle. Serological [Rose Bengal test (RBT), complement fixation test (CFT) and indirect ELISA (iELISA)], mo-lecular (PCR) and bacteriological methods were used to detect Brucella antibodies and Brucella spp. from 200 slaughtered cattle in 14 abattoirs. RESULTS: The RBT revealed a seroprevalence of brucellosis as 11.0% (22 of 200) and iELISA confirmed 5.5% (11 of 200). The estimated seroprevalence from RBT and iELISA was 5.5% while RBT and CFT was 2.0% (4 of 200). Brucella melitensis (n = 6) and B. abortus (n = 5) were isolated from 11 cattle tissues (5.5%) as confirmed to spe-cies level with AMOS PCR and differentiated from vaccine strains with Bruce-ladder PCR. Seven of the 11 isolates originated from seropositive cattle of which five were biotyped as B. abortus bv 1 (n = 2) and B. melitensis bv 2 (n = 1) and B. melitensis bv 3 (n = 2). CONCLUSIONS: This is the first documentation of B. melitensis in cattle in South Africa. The zoonotic risk of brucellosis posed by Brucella-infected slaughter cattle to abattoir workers and consumers of improperly cooked beef cannot be ignored.Gauteng Department of Agriculture and Rural Development (GDARD)http://www.wileyonlinelibrary.com/journal/vms3pm2020Production Animal StudiesVeterinary Tropical Disease

Sequencing and Analysis of Lumpy Skin Disease Virus Whole Genomes Reveals a New Viral Subgroup in West and Central Africa

Lumpy skin disease virus (LSDV) is a member of the capripoxvirus (CPPV) genus of the Poxviridae family. LSDV is a rapidly emerging, high-consequence pathogen of cattle, recently spreading from Africa and the Middle East into Europe and Asia. We have sequenced the whole genome of historical LSDV isolates from the Pirbright Institute virus archive, and field isolates from recent disease outbreaks in Sri Lanka, Mongolia, Nigeria and Ethiopia. These genome sequences were compared to published genomes and classified into different subgroups. Two subgroups contained vaccine or vaccine-like samples (“Neethling-like” clade 1.1 and “Kenya-like” subgroup, clade 1.2.2). One subgroup was associated with outbreaks of LSD in the Middle East/Europe (clade 1.2.1) and a previously unreported subgroup originated from cases of LSD in west and central Africa (clade 1.2.3). Isolates were also identified that contained a mix of genes from both wildtype and vaccine samples (vaccine-like recombinants, grouped in clade 2). Whole genome sequencing and analysis of LSDV strains isolated from different regions of Africa, Europe and Asia have provided new knowledge of the drivers of LSDV emergence, and will inform future disease control strategies

Sequencing and Analysis of Lumpy Skin Disease Virus Whole Genomes Reveals a New Viral Subgroup in West and Central Africa

Lumpy skin disease virus (LSDV) is a member of the capripoxvirus (CPPV) genus of the Poxviridae family. LSDV is a rapidly emerging, high-consequence pathogen of cattle, recently spreading from Africa and the Middle East into Europe and Asia. We have sequenced the whole genome of historical LSDV isolates from the Pirbright Institute virus archive, and field isolates from recent disease outbreaks in Sri Lanka, Mongolia, Nigeria and Ethiopia. These genome sequences were compared to published genomes and classified into different subgroups. Two subgroups contained vaccine or vaccine-like samples (“Neethling-like” clade 1.1 and “Kenya-like” subgroup, clade 1.2.2). One subgroup was associated with outbreaks of LSD in the Middle East/Europe (clade 1.2.1) and a previously unreported subgroup originated from cases of LSD in west and central Africa (clade 1.2.3). Isolates were also identified that contained a mix of genes from both wildtype and vaccine samples (vaccine-like recombinants, grouped in clade 2). Whole genome sequencing and analysis of LSDV strains isolated from different regions of Africa, Europe and Asia have provided new knowledge of the drivers of LSDV emergence, and will inform future disease control strategies