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

Abstract

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.