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The study examines the potential of Zambia’s cattle industry as a solution to food insecurity, while also addressing obstacles such as climate change, disease outbreaks and limited technology adoption. Employing the desktop review study design and drawing on data from the 2022 Livestock Survey Report and the 2017/2018 fisheries and livestock census, augmented by a systematically conducted literature review encompassing 63 peer-reviewed articles, the study provides insights into the geographical dynamics, challenges and opportunities within Zambia’s cattle subsector. It identifies key factors influencing sustainable livestock production, including cattle management systems, environmental characteristics and socio-economic factors. Government policies, market dynamics and infrastructure development are highlighted as moderating factors shaping the viability of cattle enterprises. The study reveals notable variations in cattle populations across regions and identifies obstacles faced by smallholder farmers, including limited financing and regulatory burdens. Despite challenges, the study suggests that fostering sustainable cattle business models in Zambia is achievable through innovative strategies such as enhancing value propositions and improving market access. By addressing these challenges and seizing opportunities, Zambia can enhance its cattle industry, contributing to sustainable development, food security and economic prosperity.

Background and Aim: Inappropriate use of antimicrobials exacerbates antimicrobial resistance (AMR) in the poultry sector. Information on factors driving AMR in the layer poultry sector is scarce in Zambia. This study examined the drivers of AMR in the layer poultry sector in the Lusaka and Copperbelt Provinces of Zambia. Materials and Methods: This cross-sectional study employed a structured questionnaire in 77 layer poultry farms in the provinces of Lusaka and Copperbelt, Zambia, from September 2020 to April 2021. Data analysis was conducted using Stata version 16.1. Antimicrobial resistance was defined as the presence of multidrug resistance (MDR) isolates. Multivariable regression analysis was used to identify drivers of AMR. Results: In total, 365 samples were collected, from which 339 (92.9%) Escherichia coli and 308 (84.4%) Enterococcus spp. were isolated. Multidrug resistance was identified in 39% of the E. coli and 86% of the Enterococcus spp. The overall prevalence of AMR in layer poultry farms was 51.7% (95% confidence interval [CI]: 40.3%–63.5%). Large-scale farmers (Adjusted odds ratio [AOR] = 0.20, 95% CI: 0.04%–0.99%) than small-scale and farmers who were aware of AMR than those who were unaware (AOR = 0.26, 95% CI: 0.08%–0.86%) were less likely to experience AMR problems. Conclusion: This study found a high prevalence of AMR in layer poultry farming linked to the type of farm management practices and lack of AMR awareness. Evidence of high MDR in our study is of public health concern and requires urgent attention. Educational interventions must increase AMR awareness, especially among small- and medium-scale poultry farmers.

Antimicrobial resistance (AMR) among Escherichia coli from food animals is a rising problem, and heavy antimicrobial use in poultry is a contributing factor. In Zambia, studies linking poultry-associated AMR and antibiotic use (AMU) are rare. This study aimed to investigate commercial and medium-/small-scale poultry farmers’ usage of antimicrobials based on a questionnaire survey in ten districts of Zambia. In addition, the study characterized extended-spectrum β-lactamase (ESBL)-producing E. coli isolates obtained from poultry in the same districts. Data regarding knowledge and usage of antimicrobials were collected from commercial and medium-/small-scale poultry farmers using a pre-tested structured questionnaire. At the same time, cloacal samples were collected and analyzed. One hundred and fifty E. coli isolates were tested for antimicrobial susceptibility using eight antibiotic classes. The isolates were further screened for ESBL production by streaking them on cefotaxime (CTX)-supplemented MacConkey agar, then subjecting them to sequencing on a NextSeq. The questionnaire survey showed that more medium-/small-scale than commercial poultry farmers used antimicrobials (OR = 7.70, 95% CI = 2.88–20.61) but less prescriptions (OR = 0.02, 95% CI = 0.00–0.08). Susceptibility testing revealed that resistance was highest to ampicillin (128/148, 86.5%) and tetracycline (101/136, 74.3%) and that the prevalence of multidrug resistance (MDR) (28/30, 93.3%) was high. Whole-genome sequencing (WGS) of eight (8/30, 26.7%) isolates with CTX Minimum Inhibitory Concentration (MIC) ≥ 4 µg/mL revealed the presence of ESBL-encoding genes blaCTX-M-14, blaCTX-M-55, and blaTEM. WGS also detected other AMR genes for quinolones, aminoglycosides, phenicols, tetracycline, macrolides, and folate-pathway antagonists. Altogether, the questionnaire survey results showed a higher proportion of AMU and lower prescription usage among medium-/small-scale farmers. In addition, our results emphasize the circulation of ESBL-producing E. coli strains with associated MDR. It is critical to educate farmers about AMR risks and to encourage responsible usage of antimicrobials. Furthermore, there is a need to strengthen regulations limiting access to antimicrobials. Finally, there is a need to establish a one health system to guide public health response.

AbstractIn the fourth industrial revolution, achieving sustainable agricultural development while feeding a growing world population and maintaining a balanced interrelationship between the economy, society, and the environment has been a significant challenge. However, COVID-19 disrupted agricultural extension and advisory systems, affecting all market participants. The complexity of the pandemic has impacted the private sector, public sector, extension staff, donor funding, education, and research systems, subsequently affecting human nutrition. This paper aims to expertly review COVID-19’s effects on the sustainability of gender-specific agricultural extension service systems. Demand and supply market stakeholders in the extension system are thematically discussed. The findings reveal various consequences of COVID-19 for market actors on both the demand and supply sides of the extension system. Private-sector players often support expansion directly or indirectly, with multi-sectorial support and funding strategies existing for sustainability. In many developing nations, the comprehensive extension system was disrupted by COVID-19, impacting household incomes and nutrition. The pandemic accelerated the adoption of digital technologies in many developing nations, transitioning from a centralized to a decentralized and pluralistic extension model. Multi-sector coordination and execution, especially from health and other sectors, are crucial in managing complex crises that disrupt the market system.

Poultry products in Zambia form an integral part of the human diet in many households, as they are cheap and easy to produce. The burden of poultry diseases has, however, remained a major challenge. Growing consumer demand for poultry products in Zambia has resulted in non-prudent antimicrobial use on farms, intending to prevent and treat poultry diseases for growth optimisation and maximising profits. This cross-sectional study aimed to identify the different types of bacteria causing diseases in chickens in Lusaka and to detect the extended-spectrum lactamase (ESBL)-encoding genes. We collected 215 samples from 91 diseased chickens at three post-mortem facilities and screened them for Gram-negative bacteria. Of these samples, 103 tested positive for various clinically relevant Enterobacteriaceae, including Enterobacter (43/103, 41.7%), Escherichia coli (20/103, 19.4%), Salmonella (10/103, 9.7%), and Shigella (8/103, 7.8%). Other isolated bacteria included Yersinia, Morganella, Proteus, and Klebsiella, which accounted for 21.4%. E. coli, Enterobacter, Salmonella, and Shigella were subjected to antimicrobial susceptibility testing. The results revealed that E. coli, Enterobacter, and Shigella were highly resistant to tetracycline, ampicillin, amoxicillin, and trimethoprim-sulfamethoxazole, while Salmonella showed complete susceptibility to all tested antibiotics. The observed resistance patterns correlated with antimicrobial usage estimated from sales data from a large-scale wholesale and retail company. Six (6/14, 42.9%) E. coli isolates tested positive for blaCTX-M, whilst eight (8/14, 57.1%) Enterobacter samples tested positive for blaTEM. Interestingly, four (4/6, 66.7%) of the E. coli isolates carrying blaCTX-M-positive strains were also positive for blaTEM. Sanger sequencing of the PCR products revealed that five (5/6, 83.3%) of the abovementioned isolates possessed the blaCTX-M-15 allele. The results suggest the presence of potentially pathogenic ESBL-producing Enterobacteriaceae in poultry, threatening public health.

Acinetobacter species have emerged as one of the leading causative agents of Hospital-Acquired Infections (HAIs). Acinetobacter baumannii, the most commonly isolated clinical subspecies among the Acinetobacter species, is known to cause a wide range of HAIs, with an increased risk of mortality ranging from 8% to 40% in hospitalized patients. This study aimed to evaluate the clinical significance of Acinetobacter species isolated from admitted patients, ward environmental surfaces and instruments at the University Teaching Hospital, (UTH) in Lusaka, Zambia.This was a facility-based cross-sectional study that involved collecting clinical samples (cerebrospinal fluid, sputum, pus swabs, blood culture, urine, synovial fluid, pericardial fluid, and pleural fluid) that were submitted to the Microbiology Laboratory at UTH as part of the routine diagnosis and patient care between July and December 2021. Environmental specimens were also collected from surfaces and equipment in contact with patients, such as beds, beddings, suction machines, ventilators, ward gowns and endotracheal tubes.Sixty Acinetobacter isolates were recovered, 40 (66.7%) from clinical specimens and 20 (33.3%) from environmental samples, respectively. Among the clinical isolates, the admission ward had the highest (21.7%; 95% CI: 10.9–32.4) while the main ICU had the least (1.7%; 95% CI: 0.0–5.0%); for environmental samples, the highest isolation was from taps (25%) and sinks (20%). Among the sixty Acinetobacter isolates, A. baumannii was the most prevalent subspecies (n = 43). Highest resistance was observed to tetracycline (98%; 95% CI: 94–100) and co-trimoxazole (70%; 95% CI: 58–82) while the lowest resistance was seen in imipenem (17%; 95% CI: 7.3–28.3), tobramycin (20%; 95%CI: 11.7–36) and cefotaxime (22%; 95% CI: 24–22). The AdeB gene (efflux pump) was detected in 82.5% of the clinical isolates. The frequency of clinical isolates that showed resistance to at least three classes of antibiotics (aminoglycosides, fluoroquinolones and sulphonamides) translated into being Multidrug-resistant (MDR) and Extensive drug resistance (XDR) in the clinical isolates was 75% (30/40) and 52.5% (21/30), respectively. These findings intimate the clinical significance of HAI Acinetobacter infections and need for continuous AMR surveillance to inform treatment guidelines and regular infection prevention processes.

Abstract Background The food industry is increasingly becoming more scrutinized, given the frequency and intensity with which zoonotic diseases are being reported. Pathogen tracking has become more applicable with regards food safety. It is in this regard that the present study was formulated to track Listeria species. in freshly slaughtered cattle carcasses by utilizing standard and molecular biological techniques. Methods A cross-sectional study design was conducted from March to December 2020 with 200 samples being equally collected in the rainy and dry seasons. A total of 180 and 20 swabs were aseptically collected from carcasses and the environment respectively. Samples were first subjected to pre-enrichment in half-strength Fraser broth followed by enrichment in full strength Fraser broth and subsequent plating on Listeria agar. Listeria growth characteristics were identified up to species level based on their morphological and biochemical characteristics. Further, molecular detection and phylogenetic analysis was conducted. Quantitative proportionate survey data were analyzed using Stata Version 15 software to estimate crude prevalence taking into account complex design at abattoir level. Factors associated with contamination were characterized using logistic regression. Sequences were analyzed using, Genetyyx version 12 and phylogenetic Mega. Results Of the 200 samples, 19 were positive for Listeria species identified as L.innocua 14/19 (73.7%) and L. monocytogenes 5/19 (26.3%). All isolates were from freshly slaughtered carcasses, and none from environment. Siginificant differences in contamination levels were observed based on season: rainy season yielded 14 (73.6%) whilst the dry season 5 (26.3%). The L. monocytogenes strains showed a high degree of homogeneity on phylogenetic analysis and clustered based on abattoir. Seasonality was identified as a major determinant influencing contamination based on the final logistic regression model. Conclusion This study found evidence of L. monocytogenes contamination on traditionally raised beef carcasses across various abattoirs surveyed. The failure to find Listeria contamination on the abattoir environment may to a greater extent intimate cattle carccases as primary sources of contamination. However, a more comprerehnsive study incorporating different geographical regions is needed to conclusively ascertain these present findings.

This study investigated disease outbreaks in farmed Oreochromis niloticus (Nile tilapia) in Siavonga among small-scale cage culture farms on Lake Kariba in order to establish bacterial etiological agents associated with fish mortality and to determine their antibiotic susceptibility. A total of 300 fish samples from 11 farms were aseptically collected and bacteria were isolated from the kidney, liver, brain, and spleen. The isolates were identified using their morphological characteristics and conventional biochemical tests. The antibiotic susceptibility of selected bacteria was determined by the Kirby–Bauer disc diffusion method. The following well-known fish pathogens were identified at a prevalence of Aeromonas spp. (13%), Pseudomonas spp. (10.3%), Micrococcus spp. (9.7%), Klebsiella spp. (8.7%), Lactococcus spp. (7.3%), Streptococcus spp. (7.0%), and Acinetobacter spp. (7.0%). All the isolates tested were susceptible to doxycycline, and complete resistance to ciprofloxacin, co-trimoxazole, and cephalothin was recorded in the Bacillus spp. The observed resistance could be attributed to bacteria from terrestrial sources as fish farmers do not administer antibiotics to fish. To our knowledge, this is the first study to establish the occurrence of several bacterial species infecting tilapia in Zambia and the first to determine the antibiotic susceptibility of fish bacteria among small-scale farms on Lake Kariba. The current study provides baseline information for future reference and fish disease management on Lake Kariba and in Zambia.

The emergence of pre-extensively drug-resistant tuberculosis (pre-XDR-TB) is a threat to TB control programs in developing countries such as Zambia. Studies in Zambia have applied molecular techniques to understand drug-resistance-associated mutations, circulating lineages and transmission patterns of multi-drug-resistant (MDR) Mycobacterium tuberculosis. However, none has reported genotypes and mutations associated with pre-XDR TB. This study characterized 63 drug-resistant M. tuberculosis strains from the University Teaching Hospital between 2018 and 2019 using targeted gene sequencing and conveniently selected 50 strains for whole genome sequencing. Sixty strains had resistance mutations associated to MDR, one polyresistant, and two rifampicin resistant. Among MDR strains, seven percent (4/60) had mutations associated with pre-XDR-TB. While four, one and nine strains had mutations associated with ethionamide, para-amino-salicylic acid and streptomycin resistances, respectively. All 50 strains belonged to lineage 4 with the predominant sub-lineage 4.3.4.2.1 (38%). Three of four pre-XDR strains belonged to sub-lineage 4.3.4.2.1. Sub-lineage 4.3.4.2.1 strains were less clustered when compared to sub-lineages L4.9.1 and L4.3.4.1 based on single nucleotide polymorphism differences. The finding that resistances to second-line drugs have emerged among MDR-TB is a threat to TB control. Hence, the study recommends a strengthened routine drug susceptibility testing for second-line TB drugs to stop the progression of pre-XDR to XDR-TB and improve patient treatment outcomes.

The presence of antimicrobial-resistant Enterococci in poultry is a growing public health concern worldwide due to its potential for transmission to humans. The aim of this study was to determine the prevalence and patterns of antimicrobial resistance and to detect drug-resistant genes in Enterococcus faecalis and E. faecium in poultry from four districts in Zambia. Identification of Enterococci was conducted using phenotypic methods. Antimicrobial resistance was determined using the disc diffusion method and antimicrobial resistance genes were detected using polymerase chain reaction and gene-specific primers. The overall prevalence of Enterococci was 31.1% (153/492, 95% CI: 27.1–35.4). Enterococcus faecalis had a significantly higher prevalence at 37.9% (58/153, 95% CI: 30.3–46.1) compared with E. faecium, which had a prevalence of 10.5% (16/153, 95% CI: 6.3–16.7). Most of the E. faecalis and E. faecium isolates were resistant to tetracycline (66/74, 89.2%) and ampicillin and erythromycin (51/74, 68.9%). The majority of isolates were susceptible to vancomycin (72/74, 97.3%). The results show that poultry are a potential source of multidrug-resistant E. faecalis and E. faecium strains, which can be transmitted to humans. Resistance genes in the Enterococcus species can also be transmitted to pathogenic bacteria if they colonize the same poultry, thus threatening the safety of poultry production, leading to significant public health concerns.

Background. Salmonella infections are a public health problem across the globe. In South Sudan, there is little information regarding the prevalence and antibiotic resistance patterns of Salmonella. Therefore, this study assessed the prevalence and antimicrobial susceptibility of Salmonella isolates from chickens and humans in South Sudan. Fecal samples were collected and cultured on Xylose Lysine Deoxycholate Agar for the isolation of Salmonella and confirmed using biochemical tests and PCR through the amplification of the invA gene. A total of 417 fecal samples were examined, of which 270 (64.7%) were chicken cloacal swabs while 147 (35.3%) were humans’ stool specimens. Results. Eleven (11) Salmonella isolates were isolated from humans while nine were from chickens. All 11 isolates from humans were susceptible to sulfamethoxazole-trimethoprim, chloramphenicol, streptomycin, cefotaxime, nalidixic acid, and gentamicin. However, 4 (36.7%) isolates showed resistance to ciprofloxacin, 2 (18.9%) to ampicillin, and 1 (9.1%) to tetracycline. All chicken isolates were susceptible to chloramphenicol, streptomycin, sulfamethoxazole-trimethoprim, ciprofloxacin, cefotaxime, nalidixic acid, and gentamicin but showed resistance to tetracycline 2 (22.2%) and ampicillin 1 (11.1%). Conclusion. Antimicrobial resistant isolates were isolated in both chickens and humans. Further, MDR isolates were found in both chicken and human samples, and this is a public health concern. This, therefore, calls for concerted efforts to educate producers and consumers on public health, food safety, food hygiene in food production, and enhancement of surveillance programmes on zoonotic bacteria and antimicrobial susceptibility.

Relapsing fever (RF) is an arthropod-borne disease caused by Borrelia spirochete, which is one of the major public health concerns in endemic regions including Africa. However, information on Borrelia spirochetes is limited in Zambia. Here, we investigate the Borrelia spirochetes harbored by Ornithodoros ticks in Zambian National Parks. We analyzed 182 DNA samples pooled from 886 Ornithodoros ticks. Of these, 43 tested positive, and their sequence revealed that the ticks harbored both Old and New World RF borreliae. This research presents the first evidence of Old-World RF borreliae in Zambia. The New World RF borreliae detected herein differed from the Candidatus Borrelia fainii previously reported in Zambia and were closely related to the pathogenic Borrelia sp. VS4 identified in Tanzania. Additionally, Borrelia theileri was recently reported in Zambia. Hence, at least four different Borrelia species occur in Zambia, and the organisms causing relapsing fever there might be more complex than previously thought. We empirically confirmed that real-time PCR with TaqMan minor groove binder probes accurately and simultaneously detected both Old and New World RF. In this manner, they could facilitate quantitative analyses of both types of RF borreliae. Subsequent investigations should endeavor to isolate the aforementioned Borrelia spp. and perform serosurveys on patients with RF.

The study investigated the management practices that contribute to disease outbreaks in farmed tilapia in Lusaka and central and southern provinces in Zambia. It was a cross-sectional qualitative study undertaken from January to March 2021 in which questionnaires were administered to 49 farmers to assess their fish health management and biosecurity competence. Data were analysed using means, percentages, ratios, and logistical regression. The results showed that the majority of the farms had high stocking densities (>8 fish/m2, 44.4%), reared Nile tilapia (67.7%), and sourced water for farming from rivers and streams (45.7%). A few farmers measured water quality parameters daily (16.7%) and removed dead fish from ponds daily (20.8%). The stocking density (p = 0.013), fish species (p = 0.031), dead fish disposal methods (p = 0.023), and control of predator birds (p = 0.016) influenced the total mortality recorded on farms, while pond type (p = 0.031 and p = 0.045), water source (p = 0.023), and stocking density (p = 0.027) influenced the duration of a mortality episode. It is evident that some fish health management practices and biosecurity concepts among tilapia farmers in the study area are inadequate and may not contain disease outbreaks or the spread of pathogens.

Background: Pseudomonas aeruginosa (P. aeruginosa) and Acinetobacter species and are among the leading causes of hospital-acquired infections (HAI). Treating patients with HAI caused by these pathogens is usually a challenge due to intrinsic and acquired resistance to the commonly used and affordable antibiotics, which has been heightened by the emergence of metallo-β lactamases thereby exhibiting resistance to carbapenems. Currently, carbapenem resistance represents an increasing problem in the health care settings globally but there is still a paucity of data on carbapenem resistance in our setting. Objective: To detect carbapenem resistance and the susceptibility to other antimicrobial agents in P. aeruginosa and Acinetobacter species isolated from blood, sputum, skin and soft tissue specimens between March, 2018 and June, 2019 at the University Teaching Hospital (UTH), Lusaka, Zambia. Method: This was a retrospective hospital based study, involving routine blood, sputum, skin and soft tissue samples submitted to the microbiology laboratory. Conventional microbiology methods were used to identify the organisms, while Kirby-Bauer disc diffusion method was used to determine the antimicrobial susceptibility profiles of the isolates. In order to detect carbapenemase production in P. aeruginosa isolates that were resistant to imipenem by disc diffusion, the isolates were subjected to the modified carbapenem inactivation method (mCIM) method. Results: A total of 384 samples were analyzed, of which 84 P. aeruginosa and 11 Acinetobacter species were isolated. The P. aeruginosa isolates were susceptible to imipenem (94%), piperacillin-tazobactum (95.2%), amikacin (91%), ciprofloxacin (69%), gentamicin (63.1%) and ceftazidime (51.2%). The Acinetobacter species isolates were most susceptible to amikacin (90.9%), imipenem (82.9%) and piperacillin-tazobactum (82.8%), ciprofloxacin, tetracycline and cefepime all at 27.3% each but had the lowest susceptibility at 9.1% to both gentamicin and cefotaxime. Carbapenem resistance in P. aeruginosa was 5/84 (6%) and 2/11 (18.2%) in Acinetobacter species. Carbapenemase production was confirmed in all five P. aeruginosa isolates. Conclusion: Carbapenem resistance was low in both organisms demonstrating that imipenem is still an effective treatment choice for invasive infections caused by these organisms in our setting. In order to reduce HAI and improve patient outcome, there is need to strengthen antimicrobial surveillance/stewardship and infection control at the UTH.

Water is a global need for the health and survival of all life forms and a habitat to some. However, approximately one billion people in developing countries do not have access to safe drinking water in which Namibia is not an exception. Rural communities of Ohangwena and Omusati regions of Namibia construct hand-dug wells as a source of water. However, these hand-dug wells are not monitored for water quality and are generally poorly constructed. In addition, these hand-dug wells are easily accessed by both humans and animals making them a potential reservoir for zoonotic bacteria partly known to aid antimicrobial resistance development, a global health concern. A metagenomics analysis targeting the 16S rRNA gene was used to detect the bacterial communities in a total of 40 hand-dug well water samples, with 20 being from the wet season and the rest from the dry season. A total of 57 bacterial pathogens were detected with a diverse of zoonotic assemblage. Analysis revealed significant differences in diversity and evenness, and no significant difference in richness of zoonotic bacterial pathogens in hand-dug wells between the wet and dry seasons (P < 0.05) with the dry season having a higher diversity and evenness compared to the wet season. The analysis showed that the hand-dug well water was not definitely safe for human and livestock consumption and their exposure to zoonotic pathogens was pronounced in the dry season.

Bat trypanosomes consist of more than 30 trypanosome species from over 70 species of bats. Recent studies suggest that bats play a role in disseminating trypanosomes from African continent to the terrestrial mammals both in the Afrotropic-Palearctic Ecozones and Nearctic Ecozone. However, the diversity, distribution, and evolution of bat trypanosomes are still unclear. To better understand their evolution, more genetic data of bat trypanosomes from a variety of locations are required. During a survey of Borrelia spp. of bats inhabiting a cave in Zambia, we observed flagellate parasites from 5 of 43 hemocultures. Sequence and phylogenetic analyses of the glycosomal glyceraldehyde 3-phosphate dehydrogenase gene (gGAPDH; 572 bp) and the 18S ribosomal RNA gene (18S rRNA gene; 1,079–1,091 bp) revealed that all were Trypanosoma spp. belonged to the Trypanosoma cruzi clade. Three and two of them exhibited the similarity with T. conorhini and T. dionisii, respectively. The present study provides the first genetic data on Trypanosoma spp. of bats inhabiting Zambia. Keywords: Zambia, Bat, Trypanosoma cruzi clade, Trypanosoma conorhini, Trypanosoma dionisii

Namibia is a large country with a population slightly above 2 million of which 73% live in rural areas. It is the driest country in sub-Saharan Africa with unpredictable rainfall patterns which occur between October and May. The Cuvelai Etosha Basin is a rural setting in which most people depend on groundwater to circumvent water scarcity by the construction of hand-dug wells. Communities consume hand-dug well water throughout the year without; knowledge of its safety, and treatment prior to use. Hence, this study aimed at evaluating the seasonal safety and quality of hand-dug well water for human consumption. A 16S rRNA metagenomics study was conducted to explore the influence of season on the human pathogenic bacterial species richness, diversity, evenness and abundance in the hand-dug wells. Bacterial pathogens were identified and analysis revealed no significant difference in species richness and diversity, and a significant difference in species evenness and abundance between the wet and dry seasons. Hence, seasonality had an influence on human bacterial pathogens evenness and abundance but had no influence on species richness and diversity. In addition, the water in hand-dug wells of the Cuvelai Etosha Basin is not safe for consumption and domestic use unless sanitized. It is recommended that efforts should be made to treat the water prior to use, and hand-dug wells should be properly constructed to reduce contamination.

Increased antimicrobial resistance (AMR) has been reported for pathogenic and commensal Escherichia coli (E. coli), hampering the treatment, and increasing the burden of infectious diarrhoeal diseases in children in developing countries. This study focused on exploring the occurrence, patterns, and possible drivers of AMR E. coli isolated from children under-five years in Zambia. A hospital-based cross-sectional study was conducted in the Lusaka and Ndola districts. Rectal swabs were collected from 565 and 455 diarrhoeic and healthy children, respectively, from which 1020 E. coli were cultured and subjected to antibiotic susceptibility testing. Nearly all E. coli (96.9%) were resistant to at least one antimicrobial agent tested. Further, 700 isolates were Multi-Drug Resistant, 136 were possibly Extensively-Drug Resistant and nine were Pan-Drug-Resistant. Forty percent of the isolates were imipenem-resistant, mostly from healthy children. A questionnaire survey documented a complex pattern of associations between and within the subgroups of the levels of MDR and socio-demographic characteristics, antibiotic stewardship, and guardians’ knowledge of AMR. This study has revealed the severity of AMR in children and the need for a community-specific-risk-based approach to implementing measures to curb the problem.

Pigs have been shown to be a reservoir for recently emerging livestock-associated Staphylococcus aureus (LA-SA), including methicillin resistant strains in many countries worldwide. However, there is sparse information about LA-SA strains circulating in Zambia. This study investigated the prevalence, phenotypic and genotypic characteristics of S. aureus from pigs and workers at farms and abattoirs handling pigs in Lusaka Province of Zambia. A total of 492 nasal pig swabs, 53 hand and 53 nasal human swabs were collected from farms and abattoirs in selected districts. Standard microbiological methods were used to isolate and determine antimicrobial susceptibility patterns of S. aureus. Polymerase Chain Reaction was used to confirm the species identity and detect antimicrobial resistance and virulence genes of isolates, whereas genetic diversity was evaluated using spa typing. Overall prevalence of S. aureus was 33.1%, 37.8% for pigs and 11.8% for humans. The isolates were resistant to several antibiotics with resistance ranging from 18% to 98% but were all susceptible to vancomycin. Typical LA-SA spa types were detected. The presence of plasmid mediated resistance genes such as tetM (12.8%), other resistance determinants and immune evasion cluster genes among the isolates is of great public health concern. Thus, continuous surveillance of S. aureus using a “One health” approach is warranted to monitor S.aureus infections and spread of antimicrobial resistance.

Antimicrobial resistance (AMR) is a global public health problem affecting animal and human medicine. Poultry production is among the primary sources of income for many Zambians. However, the increased demand for poultry products has led to a subsequent increase in antimicrobial use. This study assessed the awareness of AMR and associated factors among layer poultry farmers in Zambia. A cross-sectional study was conducted among 77 participants from September 2020 to April 2021. Data was analysed using Stata version 16.1. The overall awareness of AMR among the farmers was 47% (n = 36). The usage of antibiotics in layer poultry production was high at 86% (n = 66). Most antibiotics were accessed from agrovets (31%, n = 24) and pharmacies (21%, n = 16) without prescriptions. Commercial farmers were more likely to be aware of AMR compared to medium-scale farmers (OR = 14.07, 95% CI: 2.09–94.70), as were farmers who used prescriptions to access antibiotics compared to those who did not (OR = 99.66, 95% CI: 7.14–1391.65), and farmers who did not treat market-ready birds with antibiotics compared to those who did (OR = 41.92, 95% CI: 1.26–1396.36). The awareness of AMR among some layer farmers was low. Therefore, policies that promote the rational use of antibiotics need to be implemented together with heightened surveillance activities aimed at curbing AMR.

Background: Staphylococcus aureus is a frequent cause of hospital-acquired infections worldwide. The carriage of S. aureus among healthcare workers has been associated with transmission of the bacterium to susceptible patients. Objectives: The aim of this study was to determine the carriage rate, antimicrobial susceptibility profile and spa type of S. aureus among healthcare workers at a large tertiary referral hospital in Lusaka, Zambia. Methods: This was a prospectivecross-sectional study that involved the collection of nasal and hand swabs from 140 healthcare workers from May to July 2017 at the University Teaching Hospital in Lusaka. In this study, we recruited 103 nurses, 28 doctors and 9 laboratory scientists. Conventional microbiological methods were used to isolate and identify S. aureus. Antimicrobial susceptibility testing, including methicillin resistance was determined using the disc diffusion method, while molecular analysis of the isolates was achieved by spa typing. Results: The overall S. aureus carriage among the health care workers was 17.1% (24/140). Of these, 13.6% (19/140) and 8.6% (12/140) were nasal and hand carriers, respectively. Carriage rate was highest among doctors (17.9%, 5/28), followed by nurses (17.5%, 18/103) and laboratory scientists (11.1%, 1/9). About 25.8% (8/31) of the S. aureus isolates were methicillin-resistant and showed resistance to more than four antibiotics. About 25.8% (8/31) of the S. aureus isolates were positive for the spa gene. Of these, seven were typeable. Two spa types, t015 (42.8%, 3/7) and t069 (14.3%, 1/7) were detected, whereas the spa type for 42.8% (3/7) of the isolates were unknown. Conclusion: The carriage of S. aureus and prevalence of Methicillin-resistant S. aureus among healthcare workers examined was high. This calls for regular intervention measures such as screening and decolonizing of healthcare workers to reduce the carriage and spread of this pathogen in healthcare centres in Zambia. Keywords: Antimicrobial resistance, Carriage rate, Healthcare workers, Methicillin-resistant Staphylococcus aureus, Spa typing

Ticks are obligate ectoparasites as they require to feed on their host blood during some or all stages of their life cycle. In addition to the pathogens that ticks harbor and transmit to vertebrate hosts, they also harbor other seemingly nonpathogenic microorganisms including nutritional mutualistic symbionts. Tick nutritional mutualistic symbionts play important roles in the physiology of the host ticks as they are involved in tick reproduction and growth through the supply of B vitamins as well as in pathogen maintenance and propagation. Coxiella-like endosymbionts (CLEs) are the most widespread endosymbionts exclusively reported in ticks. Although CLEs have been investigated in ticks in other parts of the world, there is no report of their investigation in ticks in Zambia. To investigate the occurrence of CLEs, their maintenance, and association with host ticks in Zambia, 175 ticks belonging to six genera, namely Amblyomma, Argas, Haemaphysalis, Hyalomma, Ornithodoros, and Rhipicephalus, were screened for CLEs, followed by characterization of CLEs by multi-locus sequence typing of the five Coxiella housekeeping genes (dnaK, groEL, rpoB, 16S rRNA, and 23S rRNA). The results showed that 45.7% (n = 80) were positive for CLEs. The comparison of the tick 16S rDNA phylogenetic tree with that of the CLEs concatenated sequences showed that there was a strong correlation between the topology of the trees. The results suggest that most of the CLEs have evolved within tick species, supporting the vertical transmission phenomenon. However, the negative results for CLE in some ticks warrants further investigations of other endosymbionts that the ticks in Zambia may also harbor.

A cross-sectional study was used to identify and assess prevalence and phenotypic antimicrobial resistance (AMR) profiles of Escherichia coli and other enterobacteria isolated from healthy wildlife and livestock cohabiting at a 10,000 acres game ranch near Lusaka, Zambia. Purposive sampling was used to select wildlife and livestock based on similarities in behavior, grazing habits and close interactions with humans. Isolates (n = 66) from fecal samples collected between April and August 2018 (n = 84) were examined following modified protocols for bacteria isolation, biochemical identification, molecular detection, phylogenetic analysis, and antimicrobial susceptibility testing by disc diffusion method. Data were analyzed using R software, Genetyx ver.12 and Mega 6. Using Applied Profile Index 20E kit for biochemical identification, polymerase chain reaction assay and sequencing, sixty-six isolates were identified to species level, of which Escherichia coli (72.7%, 48/66), E. fergusonii (1.5%, 1/66), Shigella sonnei (22.7%, 14/66), Sh. flexinerri (1.5%, 1/66) and Enterobacteriaceae bacterium (1.5%, 1/66), and their relationships were illustrated in a phylogenetic tree. Phenotypic antimicrobial resistance or intermediate sensitivity expression to at least one antimicrobial agent was detected in 89.6% of the E. coli, and 73.3% of the Shigella isolates. The E. coli isolates exhibited the highest resistance rates to ampicillin (27%), ceftazidime (14.3%), cefotaxime (9.5%), and kanamycin (9.5%). Multidrug resistance (MDR) was detected in 18.8% of E. coli isolates while only 13.3% Shigella isolates showed MDR. The MDR was detected among isolates from impala and ostrich (wild animals in which no antimicrobial treatment was used), and in isolates from cattle, pigs, and goats (domesticated animals). This study indicates the possible transmission of drug-resistant microorganisms between animals cohabiting at the wildlife–livestock interface. It emphasizes the need for further investigation of the role of wildlife in the development and transmission of AMR, which is an issue of global concern.

Bat-associated bartonellae, including Bartonella mayotimonensis and Candidatus Bartonella rousetti, were recently identified as emerging and potential zoonotic agents, respectively. However, there is no report of bat-associated bartonellae in Zambia. Thus, we aimed to isolate and characterize Bartonella spp. from bats and bat flies captured in Zambia by culturing and PCR. Overall, Bartonella spp. were isolated from six out of 36 bats (16.7%), while Bartonella DNA was detected in nine out of 19 bat flies (47.3%). Subsequent characterization using a sequence of five different genes revealed that three isolates obtained from Egyptian fruit bats (Rousettus aegyptiacus) were Ca. B. rousetti. The isolates obtained from insectivorous bats (Macronycteris vittatus) were divided into two previously unclassified bat-associated bartonellae. A phylogenetic analysis of the six genotypes of Bartonella gltA sequences from nine pathogen-positive bat flies revealed that three genotypes belonged to the same clades as bat-associated bartonellae, including Ca. B. rousetti. The other three genotypes represented arthropod-associated bartonellae, which have previously been isolated only from ectoparasites. We demonstrated that Ca. B. rousetti is maintained between bats (R. aegyptiacus) and bat flies in Zambia. Continuous surveillance of Bartonella spp. in bats and serological surveys in humans in Africa are warranted to evaluate the public health importance of bat-associated bartonellae.

[This corrects the article DOI: 10.1371/journal.pone.0205986.].

ABSTRACT Bacillus anthracis is a Gram-positive endospore-forming bacterial species that causes anthrax in both humans and animals. In Zambia, anthrax cases are frequently reported in both livestock and wildlife, with occasional transmission to humans, causing serious public health problems in the country. To understand the genetic diversity of B. anthracis strains in Zambia, we sequenced and compared the genomic DNA of B. anthracis strains isolated across the country. Single nucleotide polymorphisms clustered these strains into three groups. Genome sequence comparisons revealed a large deletion in strains belonging to one of the groups, possibly due to unequal crossing over between a pair of rRNA operons. The deleted genomic region included genes conferring resistance to bacitracin, and the strains with the deletion were confirmed with loss of bacitracin resistance. Similar deletions between rRNA operons were also observed in a few B. anthracis strains phylogenetically distant from Zambian strains. The structure of bacitracin resistance genes flanked by rRNA operons was conserved only in members of the Bacillus cereus group. The diversity and genomic characteristics of B. anthracis strains determined in this study would help in the development of genetic markers and treatment of anthrax in Zambia. IMPORTANCE Anthrax is caused by Bacillus anthracis, an endospore-forming soil bacterium. The genetic diversity of B. anthracis is known to be low compared with that of Bacillus species. In this study, we performed whole-genome sequencing of Zambian isolates of B. anthracis to understand the genetic diversity between closely related strains. Comparison of genomic sequences revealed that closely related strains were separated into three groups based on single nucleotide polymorphisms distributed throughout the genome. A large genomic deletion was detected in the region containing a bacitracin resistance gene cluster flanked by rRNA operons, resulting in the loss of bacitracin resistance. The structure of the deleted region, which was also conserved among species of the Bacillus cereus group, has the potential for both deletion and amplification and thus might be enabling the species to flexibly control the level of bacitracin resistance for adaptive evolution.

In Zambia, anthrax outbreaks among cattle are reported on nearly an annual basis. Presently, there is a lack of serological assays and information to develop an anthrax management and control strategy. In this study, an indirect enzyme-linked immunosorbent assay (ELISA) based on recombinant protective antigen domain 1 (rPA-D1) of Bacillus anthracis was developed and used to detect anti-PA antibodies in cattle in Zambia. An antigen coating of 10 ng/well and a serum dilution of 1:100 were determined to be the optimal rPA-D1 ELISA titration conditions. The intra- and inter-assay % coefficients of variation were less than 10% and 15%, respectively. The rPA-D1 ELISA could detect seroconversion in the cattle 1 month after anthrax vaccination. In a cross-sectional study conducted in the Western Province, Zambia, 187 serum samples from 8 herds of cattle were screened for anti-PA antibodies using the rPA-D1 ELISA. The seropositive rate of the serum samples was 8%, and the mean anti-PA antibody was 0.358 ELISA units. Additionally, we screened 131 cattle serum samples from Lusaka, which is a nonendemic area, and found no significant association between the antibody levels and sampling area (endemic versus nonendemic area). Conversely, significant differences were observed between the anti-PA antibody levels and herds, anti-PA antibody levels and vaccination status and anti-PA antibody levels and vaccination timing. Collectively, these findings suggest that the rPA-D1 ELISA is a useful tool for the detection of anti-PA antibodies in cattle in Zambia. The low proportion of seropositive sera indicates that there is inadequate cattle vaccination in the Western Province and, in addition to other epidemiological factors, this may precipitate the anthrax outbreak recurrence.