Characterisation of Milk Microbiota from Subclinical Mastitis and Apparently Healthy Dairy Cattle in Free State Province, South Africa.

Bovine mastitis is an inflammation of the udder tissue of the mammary gland brought on by microbial infections or physical damage. It is characterised by physical, chemical, and biological changes in the udder and milk. While several different bacterial species have been identified as causative agents of mastitis, many subclinical mastitis (SCM) cases remain culture-negative. The aim of this study was to characterise milk microbiota from SCM and apparently healthy dairy cows (non-SCM) by 16S rRNA sequencing. Alpha-diversity metrics showed significant differences between SCM cows and non-SCM counterparts. The beta-diversity metrics in the principal coordinate analysis significantly clustered samples by type (PERMANOVA test, p < 0.05), while non-metric dimensional scaling did not (PERMANOVA test, p = 0.07). The overall analysis indicated a total of 95 phyla, 33 classes, 82 orders, 124 families, 202 genera, and 119 bacterial species. Four phyla, namely Actinobacteriota, Bacteroidota, Firmicutes, and Proteobacteria collectively accounted for more than 97% of all sequencing reads from SCM and non-SCM cow samples. The most abundant bacterial classes were Actinobacteria, Bacilli, Bacteroidia, Clostridia, and Gammaproteobacteria in non-SCM cow samples, whilst SCM cow samples were mainly composed of Actinobacteria, Alphaproteobacteria, Bacilli, Clostridia, and Gammaproteobacteria. Dominant bacterial species in non-SCM cow samples were Anthropi spp., Pseudomonas azotoformans, P. fragi, Acinetobacter guillouiae, Enterococcus italicus, Lactococcus lactis, whilst P. azotoformans, Mycobacterium bovis, P. fragi, Acinetobacter guillouiae, and P. koreensis were dominant in the SCM cow samples. The current study found differences in bacterial species between SCM and non-SCM cow milk; hence, the need for detailed epidemiological studies.

Molecular detection of zoonotic pathogens causing gastroenteritis in humans: Salmonella spp., Shigella spp. and Escherichia coli isolated from Rattus species inhabiting chicken farms in North West Province, South Africa.

Rodents are key carriers and reservoirs of various pathogens of public health importance to both human and animal diseases. This research was carried out in order to identify the selected pathogens, namely, Shigella spp., Salmonella spp. and Escherichia coli from rats that inhabit the poultry houses. A total of 154 samples from captured rats were examined for the zoonotic bacterial pathogens, of which 3.3%, 29.9% and 20.7% were harbouring Shigella spp., Salmonella spp., and E. coli, respectively. A total of 14 Shigella isolates expressed presence of ipaH gene, of which eight and five were positive for S. sonnei and S. boydii, respectively. For Salmonella, 68 isolates were positive for invA and other genes including spy with 26 (38%), sdfI 2 (18%), spvC 14 (20%), hilA 28 (41%), misL 43 (63%), orfL 31 (46%) and spiC 38 (56%). For E. coli, the aggR gene was the most prevalent (62 [42%]), followed by the eae gene, which was only detected in 21 (14%) isolates, while stx gene was not detected in any of the samples. This study shows that zoonotic pathogens with virulence genes are circulating in rodents from selected chicken farms in the North West Province of South Africa. Rodents must therefore be regarded as important carriers of zoonotic pathogens that can potentially infect both humans and animals.

Bacterial profiling of Haemonchus contortus gut microbiome infecting Dohne Merino sheep in South Africa.

A metagenomic approach was used to study the gut microbiome of Haemonchus contortus field strains and that of its predilection site, the abomasum of Dohne Merino sheep. The abomasum contents and H. contortus were collected from 10 naturally infected Dohne Merino sheep. The H. contortus specimens were classified and sexually differentiated using morphometric characters and was further confirmed through molecular identification. We investigated differences and similarities between the bacterial composition of the adult male and female H. contortus gut microbiomes, which were both dominated by bacteria from the Escherichia, Shigella, Vibrio and Halomonas genera. Major abundance variations were identified between the shared adult male and female H. contortus microbiomes. The results also revealed that Succiniclasticum, Rikenellaceae RC9 gut group and Candidatus Saccharimonas were the predominant genera in the Dohne Merino abomasum. This study provides insight into the highly diverse bacterial composition of the H. contortus gut microbiome and the Dohne Merino abomasum which needs to be studied further to explore the complex interactions of different gastrointestinal nematode microbiomes with the host.

Sero-prevalence of Taenia spp. infections in cattle and pigs in rural farming communities in Free State and Gauteng provinces, South Africa.

The aim of this study was to determine sero-prevalence of bovine and porcine cysticercosis in cattle and pigs in rural farming communities in Free State and Gauteng Provinces, Republic of South Africa. Blood samples were collected for a period of twelve months from live cattle (n=1315; 1159) and pigs (n=436; 240) and the serum extracted and stored before analysis by a monoclonal antibody based (HP10) antigen detection ELISA. Results revealed a generally high sero-prevalence and wide distribution throughout the two provinces with Free State having a higher sero-prevalence in both cattle and pigs (23% and 34%) than Gauteng province (15% and 14%). Consumption of infected meat that is either not inspected/missed at meat inspection; poor livestock management practices and limited sanitation in rural communities might have contributed to the occurrence of Taenia spp. infections in the two provinces. It is therefore, recommended that cysticercosis status of animals be established before slaughter. This would assist in ensuring that infected animals are not slaughtered for human consumption or zoonosis preventive measures are taken. Furthermore, public awareness programs on life cycles of T. saginata, T. solium and T. hydatigena and the use of more sensitive diagnostic tools are recommended as part of effective control strategies against taeniid infections.

Evaluation of larviposition site selection of Glossina brevipalpis.

Tsetse species (Diptera: Glossinidae) are vectors of trypanosome parasites which cause disease in both humans and livestock. In South Africa Glossina austeni Newstead, 1912 and G. brevipalpis Newstead, 1911 are responsible for the cyclical transmission of animal trypanosomes causing African animal trypanosomiasis also referred to as nagana. Gravid tsetse females deposit a single larva in specific sites but little information is available on biotic and abiotic factors that govern site selection. This study therefore aimed to characterize some of the substrate conditions that may influence selection of larviposition sites. Colonised, gravid female G. brevipalpis were presented with a choice of four larviposition sites. Sites differed in qualities of pH (5, 7, 9), salinity (0, 1.3, 4g/L) and the presence of other tsetse pupae (G. brevipalpis or G. austeni). These trials indicated no significant selection by gravid females with regard to pH and salinity. Females selected significantly more often for sites with pupae (P<0.05), but also favored sites containing conspecific over heterospecific pupae (P<0.05). These results present the first indication of an aggregation effect of tsetse pupae in G. brevipalpis. This may imply that G. brevipalpis larvae produce a pheromone during pupation as seen in G. morsitans morsitans. Isolation of such semio-chemicals would allow the development of larviposition traps to attract gravid females.

Prevalence of Theileria equi and Babesia caballi infections in horses belonging to resource-poor farmers in the north-eastern Free State Province, South Africa.

The prevalence of Theileria equi and Babesia caballi infections in the north-eastern Free State Province of South Africa was determined by examination of thin and thick Giemsa-stained blood smears, IFAT and PCR. No parasites were detected by microscopy from any blood samples collected at five study sites, Qwaqwa, Kestell, Harrismith, Vrede and Warden. Of the tested serum samples, 28/29 (96.5%), 20/21 (95.2%) and 42/42 (100%) were positive by IFAT for T. equi infections in Harrismith, Kestell and Qwaqwa, respectively, and 5/29 (17.2%), 13/21 (61.9%) and 30/42 (71.4%) were sero-positive for B. caballi infections in Harrismith, Kestell and Qwaqwa, respectively. All DNA samples from the study sites were negative for B. caballi infections by PCR, but five samples, two from each of Kestell and Warden and one from Vrede, were PCR positive for T. equi infections. The high prevalence of antibodies against T. equi and B. caballi in the sampled horses indicates that the animals had been exposed to T. equi and B. caballi infections but the absence of parasitaemia and very low number of positive PCR samples, however, imply that T. equi and B. caballi are endemically stable in the north-eastern Free State Province.

A trypanosome species isolated from naturally infected Haemaphysalis hystricis ticks in Kagoshima Prefecture, Japan.

Common arthropod vectors for trypanosomes are flies, fleas and bugs. This study reports on an unknown trypanosome species isolated from naturally infected Haemaphysalis hystricis ticks, hereby, referred to as Trypanosoma KG1 isolate. The parasite has been successfully cultured in vitro with L929 or HEK 293T cell line as feeder cells. This trypanosome cannot survive in vitro without feeder cells. Following experimental infections of ticks, the trypomastigote-like and the epimastigote-like forms of this trypanosome could be detected by Giemsa-stained smears of the midgut and salivary glands of Ornithodoros moubata ticks which were made to feed on a culturing medium containing Trypanosoma KG1 isolate through an artificial membrane. Trypanosoma KG1 isolate could also be detected from Giemsa-stained smears of the haemolymph up to 30 days post-inoculation into the O. moubata haemocoel. Trypanosoma KG1 isolate cannot be propagated in laboratory animals including mice, rats, rabbits and sheep. A phylogenetic tree constructed with the 18S rRNA gene indicates that Trypanosoma KG1 is a member of the stercorarian trypanosomes.

Preliminary application and evaluation of loop-mediated isothermal amplification (LAMP) for detection of bovine theileriosis and trypanosomosis in Tanzania.

The sensitivity of LAMP, PCR and microscopy to detect Theileria spp. and Trypanosoma congolense in field-derived bovine blood samples from Tanzania was evaluated and compared. No parasites were detected by microscopy. Furthermore, no bovine Theileria spp. were detected by LAMP and PCR from all the 24 samples collected from Arusha. Four and one out of 24 samples were positive for Theileria congolense infection by LAMP and PCR respectively while, 18 and nine out of 40 samples from Dar es Salaam were positive by LAMP and PCR for Theileria spp. Infection, respectively. Although all samples from Dar es Salaam were negative for Trypanosoma congolense infections by PCR, 12 out of 40 samples were LAMP positive. Whilst PCR is an established gene amplification method for the detection of Theileria and trypanosome parasites, this study introduces LAMP as an alternative molecular diagnostic tool that could be used in large-scale epidemiological surveys.