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Masters Degrees (Medical Microbiology)

Permanent URI for this collectionhttps://hdl.handle.net/10413/9620

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Browsing Masters Degrees (Medical Microbiology) by Author "Amoako, Daniel Gyamfi."

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    Molecular characterization of antibiotic-resistant Staphylococcus aureus in an intensive pig production system in KwaZulu-Natal, South Africa.
    (2021) Sineke, Ncomeka.; Amoako, Daniel Gyamfi.; Abia, Akebe Luther King.; Essack, Sabiha Yusuf.; Bester, Linda Antionette.
    The increase in antibiotic resistance in food animals and food of animal origin has been attributed to the extensive use of antibiotics during animal husbandry giving rise to multidrug-resistant bacteria. Staphylococcus aureus is a major threat in veterinary medicine, the agricultural sector and public health because of its zoonotic potential. Despite significant research on S. aureus in food animals in other parts of the world, in-depth studies outside healthcare facilities are limited in South Africa. This study characterized the molecular epidemiology of antibiotic resistant S. aureus from farm-to-fork in an intensive pig production chain in the uMgungundlovu district, Kwa-Zulu Natal, South Africa. A total of 333 samples collected along a pig production chain on the farm (faecal, litter and slurry samples) during transport (truck samples) and at the abattoir (caeca, carcass swabs, carcass rinsate and retail meat samples) were investigated for the presence S. aureus using selective media and biochemical tests. Confirmation was done by using PCR targeting the nucA gene. Antibiotic susceptibility patterns were investigated by the Kirby Bauer disk diffusion according to CLSI guidelines against the WHO-AGISAR recommended panel of antibiotics. Selected resistance and virulence genes were detected using PCR. REPPCR was used to evaluate the molecular relatedness of isolates across the pig production chain. Of the 333 samples, 141 (43%) yielded staphylococci isolates. After molecular confirmation, 97(69%) isolates were confirmed S. aureus and 44(31%) as other staphylococcal species. Isolates displayed resistance to erythromycin (85%), clindamycin (85%), penicillin-G (81%), tetracycline (79%), doxycycline (77%), vancomycin (69%), ampicillin (61%), trimethoprim/sulfamethoxazole (57%), rifampicin (57%), teicoplanin (52%), linezolid (51%), chloramphenicol (51%), nitrofurantoin (47%), moxifloxacin (33%), cefoxitin (20%), ciprofloxacin (15%), tigecycline (10%), levofloxacin (8%), gentamicin (8%), and amikacin (2%). Multidrug resistance (MDR) was recorded in 84% (80/97) of isolates with 56 different antibiograms. Resistance genes ermC, blaZ, tetK, tetM, msrA, aac’6, mecA were evident in 82%, 73%, 58%, 28%, 15%, 5%, and 53% respectively and not all resistance phenotypes were genotypically confirmed. The hla (39%), hld (23%), seb (3%), sed (2%), etb (1%), LukS/F-PV (30%) and tst (11%) virulence genes encoding hemolysin, cytotoxins, staphylococcal enterotoxins (sea and seb), exfoliative toxins, PVL pore-forming toxin and toxic shock syndrome toxin-1 were detected. Genetic fingerprinting revealed the diversity of MRSA isolates in the pig production chain with the major REP-types constituting isolates from different sources within the farm, suggesting transmission within the farm environment with no evidence of transmission across the production chain. This study highlights the phenotypic and genotypic diversity of the virulence and resistance profiles of S. aureus isolated across the pig production chain. Resistance to antibiotics used as growth promoters was evident and the high prevalence of MDR isolates with elevated MAR index values >0.2, specifically at farm level indicates exposure to environments of high antibiotic use, necessitating antibiotic stewardship and proper infection control measures in pig husbandry and intensive pig production.
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    Molecular epidemiology of antibiotic resistant Campylobacter spp. from farm-to-fork in an intensive pig production system in Kwazulu-Natal, South Africa.
    (2021) Sithole, Viwe.; Amoako, Daniel Gyamfi.; Essack, Sabiha Yusuf.; Abia, Akebe Luther King.; Bester, Linda Antionette.
    Background: Campylobacter spp. are among the leading foodborne pathogens, causing Campylobacteriosis, a zoonotic infection that results in bacterial gastroenteritis and diarrhea disease in animals and humans. The emergence and transmission of antibiotic resistance and virulence in Campylobacter spp. is increasingly reported. We investigated the molecular epidemiology of antibiotic resistant Campylobacter spp. isolated across the farm-to-fork-continuum in an intensive pig production system in the uMgungundlovu District, Kwazulu-Natal, South Africa. Methodology: Following ethical approval, samples were collected over a period of sixteen weeks from selected critical points (farm, transport, abattoir and retail) using a farm-to-fork sampling approach according to WHO-AGISAR guidelines. Overall, 520 samples were investigated for the presence of Campylobacter spp. which were putatively identified using selective media with identity and speciation confirmed by polymerase chain reaction (PCR) of specific genes. Resistance profiles were ascertained by the Kirby-Bauer disk diffusion method according to EUCAST and/or CLSI guidelines. Selected antibiotic resistance and virulence genes were identified using PCR and DNA sequencing. Clonal relatedness among the isolates was determined using enterobacterial repetitive intergenic consensus polymerase chain reaction (ERIC-PCR). Results: Altogether, 378/520 (72.7%) samples were positive for Campylobacter spp. with C. coli as the most predominant (73.3%), followed by C. jejuni (17.7%) with 9.0% classified as “other”. Relatively high levels of resistance were observed in C. coli and C. jejuni to erythromycin (89% and 99%), streptomycin (87% and 93%), tetracycline (82% and 96%), ampicillin (69% and 85%), and ciprofloxacin (53% and 67%) respectively. The lowest percentage resistance observed was for gentamicin (12%) for both C. coli and C. jejuni, and nalidixic acid (28% and 27%) for C. coli and C. jejuni respectively. Multi-drug resistance (MDR) was noted among 330/378 (87.3%) isolates. The antibiotic resistance genes observed were the tetO (74.6%), the blaOXA-61 (2.9%) and cmeB (11.1%) accounting for the resistance to tetracycline and ampicillin while the membrane efflux pump could confer resistance to ampicillin, tetracycline, ciprofloxacin, and erythromycin. All C. coli and C. jejuni isolates (21) with the gyrA gene exhibited mutation at the Thr-86-Ile region in the quinolone-resistancedetermining region (QRDR) and all C. coli and C. jejuni isolates (18) exhibiting erythromycin resistance showed common transitional mutations A2075G and A2074C in the 23S rRNA gene. Of the virulence genes tested, ciaB, dnaJ, pldA, cdtA, cdtB, cdtC and cadF were detected in 48.6%, 61.1 %, 17.4%, 67.4%, 19.3%, 51% and 5% of all Campylobacter isolates respectively. The ERIC-PCR banding patterns revealed that isolates along the continuum were highly diverse with isolates from the same sampling points belonging to the same major ERIC-types. Conclusion: We showed relatively high levels of resistance to antibiotics commonly used in intensive pig production in South Africa with some evidence, albeit minimal, of transmission across the farm-tofork continuum. This together with the virulence profiles present in Campylobacter spp. presents a challenge to food safety and a potential risk to human health. This is further exacerbated by the reduction in antibiotic treatment options necessitating routine surveillance and monitoring together with antibiotic stewardship, comprehensive biosecurity, and good animal husbandry in intensive pig production.
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    Molecular epidemiology of antibiotic resistant salmonella spp. from farm to fork in an intensive pig production system in KwaZulu-Natal South Africa.
    (2021) Tshakane, Nozipho Pamela.; Essack, Sabiha Yusuf.; Abia Akebe, Luther King.; Amoako, Daniel Gyamfi.
    Antibiotic resistance (ABR) is a worldwide challenge, and, if not resolved, can be a danger to humans, animals and the ecosystem. The inappropriate use and misuse of antibiotics in food animal production creates selection pressure for the development of bacterial resistance. We investigated the molecular epidemiology, antibiotic resistance and virulence of Salmonella spp. from farm-to-fork in an intensive pig production system in KwaZulu-Natal. A herd of pigs was followed from birth to slaughter over a period of 4 months. Following ethical approval, a total of 408 samples were collected, which consisted of feces, litter, slurry, hand and nasal swabs from occupationally exposed workers, carcass swabs and rinsate, caecal samples and pork for retail purposes. Salmonella was putatively identified using selective media, i.e., Brilliance Salmonella Agar and Salmonella Shigella Agar (SS agar). Identification to species and sub-species level was confirmed by polymerase chain reaction (PCR), where the invA gene was used to confirm Salmonella spp. and the iroB gene for Salmonella enterica. Isolates were subjected to antibiotic susceptibility testing using the Kirby-Bauer disk diffusion method against a panel of 14 antibiotics. Isolates were screened for selected virulence genes, misL, spiC, orfL, sopB, pipD, hilA and stn, conferring intracellular survival (misL), type III secretion system (spiC), adhesion and autotransporter (orfL), type III secreted effector protein (sopB), type III secreted effector associated with SPI-1 system (pipD), host cell invasion (hilA), and enterotoxin production (stn) by PCR. Genetic relatedness of the isolates was determined by ERIC-PCR. A total of 399 putative Salmonella spp. were detected by selective media, of which 49% (n= 197) were confirmed by the presence of the invA gene and 45% (n=179) were identified as Salmonella enterica by the presence of the iroB gene. The largest number of Salmonella were isolated from retail meat samples. Antibiotic susceptibility testing showed 10% (n=19) resistance to cefoxitin, 8% (n=16) to amoxicillin and 0.5% (n=1) to gentamicin and chloramphenicol. The isolates carried the hilA (91%), stn (91%), misL (89%), pipD (88%), spiC (87%), orfL (85%) and sopB (72%) virulence genes. The isolates were clonally diverse with 26 ERIC-types and four major ERIC-type groups. The large number of isolates in retail meat samples, their virulence, and, to a lesser extent their antibiotic resistance profiles poses a challenge to the food safety system and requires a comprehensive understanding of molecular epidemiology of the organism so that it’s incidence spread can be reduced and better controlled from the primary source within the food chain.