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Objectives: Klebsiella michiganensis is an emerging pathogen implicated in nosocomial infections. Here we report on the resistome, virulome and mobilome of a carbapenemase-producing K. michiganensis isolate from urban hospital effluent in Pietermaritzburg, KwaZulu-Natal, South Africa. Klebsiella sp. isolate KP124 was originally isolated from the final effluent of an urban tertiary hospital in Pietermaritzburg, KwaZulu-Natal. Methods: Following phenotypic characterisation and antibiotic susceptibility testing, the genome of carbapenemase-producing isolate KP124 was sequenced using an Illumina MiSeq platform, de novo assembled and analysed using established bioinformatics tools. Results: The draft genome of strain KP124 was 6 544 586 bp in length, comprising 203 contigs >200 bp. Following confirmation of isolate KP124 as K. michiganensis using reference genomes, the blaOXA-181 carbapenemase gene as well as 11 additional genes encoding resistance against β-lactams, aminoglycosides, fluoroquinolones and sulfonamides were detected. Virulence factors enabling iron acquisition and cell adherence, capsule locus type and plasmid replicon types were identified. Conclusion: This study represents the first report of an OXA-181 carbapenemase-producing K. michiganensis isolate from hospital effluent in South Africa. The presence of such a strain in the environment owing to the absence of hospital effluent treatment presents a potential risk to informal communities that may use contaminated surface water domestically.

Vitamin D deficiency remains a global public health issue, particularly in minority ethnic groups. This review investigates the vitamin D status (as measured by 25(OH)D and dietary intake) of the African-Caribbean population globally.A systematic review was conducted by searching key databases (PUBMED, Web of Science, Scopus) from inception until October 2019. Search terms included 'Vitamin D status' and 'African-Caribbean'. A random effects and fixed effects meta-analysis was performed by combining means and standard error of the mean.The search yielded 19 papers that included n = 5670 African-Caribbean participants from six countries. A meta-analysis found this population to have sufficient (>50 nmol/L) 25(OH)D levels at 67.8 nmol/L, 95% CI (57.9, 7.6) but poor dietary intake of vitamin D at only 3.0 µg/day, 95% CI (1.67,4.31). For those living at low latitudes 'insufficient' (as defined by study authors) 25(OH)D levels were found only in participants with type 2 diabetes and in those undergoing haemodialysis. Suboptimal dietary vitamin D intake (according to the UK recommended nutrient intake of 10 µg/day) was reported in all studies at high latitudes. Studies at lower latitudes, with lower recommended dietary intakes (Caribbean recommended dietary intake: 2.5 µg/day) found 'sufficient' intake in two out of three studies.25(OH)D sufficiency was found in African-Caribbean populations at lower latitudes. However, at higher latitudes, 25(OH)D deficiency and low dietary vitamin D intake was prevalent.

The increase in antimicrobial-resistant (AMR) foodborne pathogens, including E. coli and Salmonella in animals, humans, and the environment, is a growing public health concern. Among animals, cattle, pigs, and chicken are reservoirs of these pathogens worldwide. There is a knowledge gap on the prevalence and AMR of foodborne pathogens in small ruminants (i.e., sheep and goats). This study investigates the prevalence and antimicrobial resistance of extended-spectrum beta-lactamase (ESBL) E. coli and Salmonella from sheep and their abattoir environment in North Carolina. We conducted a year-round serial cross-sectional study and collected a total of 1128 samples from sheep (n = 780) and their abattoir environment (n = 348). Sheep samples consisted of feces, cecal contents, carcass swabs, and abattoir resting area feces. Environmental samples consisted of soil samples, lairage swab, animal feed, and drinking water for animals. We used CHROMAgar EEC with 4 μg/ml of Cefotaxime for isolating ESBL E. coli, and ESBL production was confirmed by double-disk diffusion test. Salmonella was isolated and confirmed using standard methods. All of the confirmed isolates were tested against a panel of 14 antimicrobials to elucidate susceptibility profiles. The prevalence of ESBL E. coli and Salmonella was significantly higher in environmental samples (47.7% and 65.5%) compared to the sheep samples (19.5% and 17.9%), respectively (P < 0.0001). We recovered 318 ESBL E. coli and 368 Salmonella isolates from sheep and environmental samples. More than 97% (310/318) of ESBL E. coli were multidrug-resistant (MDR; resistant to ≥3 classes of antimicrobials). Most Salmonella isolates (77.2%, 284/368) were pansusceptible, and 10.1% (37/368) were MDR. We identified a total of 24 different Salmonella serotypes by whole genome sequencing (WGS). The most common serotypes were Agona (19.8%), Typhimurium (16.2%), Cannstatt (13.2%), Reading (13.2%), and Anatum (9.6%). Prevalence and percent resistance of ESBL E. coli and Salmonella isolates varied significantly by season and sample type (P < 0.0001). The co-existence of ESBL E. coli in the same sample was associated with increased percent resistance of Salmonella to Ampicillin, Chloramphenicol, Sulfisoxazole, Streptomycin, and Tetracycline. We presumed that the abattoir environment might have played a great role in the persistence and dissemination of resistant bacteria to sheep as they arrive at the abattoir. In conclusion, our study reaffirms that sheep and their abattoir environment act as important reservoirs of AMR ESBL E. coli and MDR Salmonella in the U.S. Further studies are required to determine associated public health risks.

Vitamin D is a fundamentally critical nutrient that the human body requires to function properly. It plays an important role in musculoskeletal health due to its involvement in the regulation of calcium and phosphorus. Having a low level of vitamin D in the body may be detrimental for a wide range of health outcomes, including risk of osteoporotic and stress fractures, risk of CVD and some cancers, and lowering of the capability of the immune system. Vitamin D is an unusual nutrient; it is not a vitamin, in the true sense of the word but a pro-hormone. The main source of vitamin D is UV exposure, not dietary intake. Interestingly, there are two forms of vitamin D, vitamin D 2 and vitamin D 3 , both of which are metabolised into 25-hydroxyvitamin D (25(OH)D) in the liver, the biomarker of vitamin D status. Vitamin D deficiency is a global public health problem, especially amongst older people and ethnic minority groups. The newest publication from the UK Government's Public Health England Department recommends that vitamin D intake should be 10 μg daily and this recommendation compares well (albeit lower) with other guidelines such as the Institute of Medicine recommendation of 15 μg for those aged 1–70 years and 20 μg for those 70 years or over. Few countries, however, have a specific vitamin D policy to prevent deficiency in populations. Finland leads the way, demonstrating impressive results in reducing population-level vitamin D deficiency through mandatory food fortification programmes. Collaboration between academia, government and industry, including countries from varying latitudes, is essential to identify long-term solutions to the global issue of vitamin D deficiency. This paper provides a narrative review of the evidence related to the role of vitamin D deficiency in health outcomes, outlines controversies regarding setting levels of adequacy, identifies the prevalence of vitamin D deficiency across the globe, and identifies population-level strategies adopted by countries to prevent vitamin D deficiency.

The power of citizen science to contribute to both science and society is gaining increased recognition, particularly in physics and biology. Although there is a long history of public engagement in agriculture and food science, the term ‘citizen science’ has rarely been applied to these efforts. Similarly, in the emerging field of citizen science, most new citizen science projects do not focus on food or agriculture. Here, we convened thought leaders from a broad range of fields related to citizen science, agriculture, and food science to highlight key opportunities for bridging these overlapping yet disconnected communities/fields and identify ways to leverage their respective strengths. Specifically, we show that (i) citizen science projects are addressing many grand challenges facing our food systems, as outlined by the United States National Institute of Food and Agriculture, as well as broader Sustainable Development Goals set by the United Nations Development Programme, (ii) there exist emerging opportunities and unique challenges for citizen science in agriculture/food research, and (iii) the greatest opportunities for the development of citizen science projects in agriculture and food science will be gained by using the existing infrastructure and tools of Extension programmes and through the engagement of urban communities. Further, we argue there is no better time to foster greater collaboration between these fields given the trend of shrinking Extension programmes, the increasing need to apply innovative solutions to address rising demands on agricultural systems, and the exponential growth of the field of citizen science.

Background: Cronobacter sakazakii (formerly Enterobacter sakazakii) causes serious infections in infants with high mortality. Though contaminated powdered infant formula is the most common implicated vehicle, the organism has been isolated from other foods and environmental sources and infections have occurred among persons who did not consume or handle formula milk. There is significant underreporting of C. sakazakii infections in most countries and very little data is available from India. There is no information on human carriage, environmental sources and potential transmission routes of this pathogen from India. The aim of this study was to study the prevalence of this organism in North India (Haryana, Punjab and Chandigarh) and study the antibiotic resistance. Methods & Materials: A wide range of samples (n = 887) were collected from poultry farms, meat abattoirs (pork and mutton), dairy farms (pig stool, goat stool), human diarrhea stool samples and domestic environmental samples (water and soil) and processed for Cronobacter sakazakii. Presumptive C. sakazakii colonies were confirmed by MALDI-TOF. Antibiotic sensitivity was performed. Results: Incidence of Cronobacter sakazakii was found to be 7.1% with individual incidence of 3.37% in humans, 9.4% in goat meat, 11.1% goat feces, 5.9% in pork, 7% pig stool, 6.7% in chicken stool, 11.5% environment. Of the 63 isolates, maximum resistance was observed against ampicillin (50.8%), followed by ciprofloxacin (20.6%) and them meropenem (17.2%). Carbapenem resistance was found in both animal food source and human strains. Conclusion: Cronobacter sakazakii is part of fecal flora of humans as well as is carried by food production animals and is also present in domestic environment with potential risk of cross-contamination. The overall antibiotic resistance in human isolates was low in comparison to food animals and their fecal samples. A higher resistance in animal isolates could be due to rampant use of antibiotics as growth promoters in animals.

Increasing evidence supports the concept that early-life environmental influences, including nutrition and stress, have an impact on long-term health outcomes and disease susceptibility. The objective of the present study was to determine whether dietary spray-dried plasma (SDP), fed during the first 2 weeks post-weaning (PW), influences subsequent immunological and intestinal injury responses to Salmonella typhimurium challenge. A total of thirty-two piglets (age 16-17 d) were weaned onto nursery diets containing 0, 2·5 % SDP (fed for 7 d PW) or 5 % SDP (fed for 14 d PW), and were then fed control diets (without SDP), for the remainder of the experiment. At 34 d PW (age 50 d), pigs were challenged with 3 × 10⁹ colony-forming units of S. typhimurium. A control group (non-challenged) that was fed 0 % SDP in the nursery was included. At 2 d post-challenge, the distal ileum was harvested for the measurement of inflammatory, histological and intestinal physiological parameters. S. typhimurium challenge induced elevated ileal histological scores, myeloperoxidase (MPO), IL-8 and TNF, and increased intestinal permeability (indicated by reduced transepithelial voltage (potential difference) and elevated 4 kDa fluorescein isothiocyanate dextran (FD4) flux rates). Compared with S. typhimurium-challenged controls (0 % SDP), pigs fed the 5 % SDP-14 d diet exhibited reduced ileal histological scores, MPO levels, IL-8 levels and FD4 flux rates. Pigs fed the 5 % SDP-14 d nursery diet exhibited increased levels of plasma and ileal TNF-α in response to the challenge, compared with the other treatments. These results indicate that inclusion of SDP in PW diets can have an influence on subsequent immunological and intestinal injury responses induced by later-life S. typhimurium challenge.

The objective of this study was to determine the prevalence of Campylobacter and Salmonella in broilers and their distribution in the indoor and outdoor farm environment. Nine hundred samples (400 faecal; 300 indoor environment; 200 outdoor environment), were collected from 10 individual broiler houses on 10 farms. Campylobacter jejuni prevalence was significantly higher (P=0.003) in faeces (29.5%; 118/400) than the environment (0.8%; 4/500) in contrast to Salmonella Typhimurium from faecal (8.8%; 35/400) and environmental (8.4%; n=42/500) sources (P=0.217). S. Typhimurium predominantly exhibited antimicrobial resistance (AR) to streptomycin (46%) and tetracycline (31.5%). C. jejuni isolates exhibited AR only to tetracycline (55.5%). The PFGE profile revealed 100% similarity between S. Typhimurium isolates from faecal and environmental sources. No relationship was detected between C. jejuni isolates. The low prevalence of Campylobacter and Salmonella in the outdoor environment indicates that it may not be a significant reservoir for transmission of these pathogens on broiler farms.

Clostridium difficile is the leading cause of nosocomial diarrhea in humans and a major cause of enteritis in neonatal piglets, foals and calves. The aim of this longitudinal study was to determine and compare the prevalence, antimicrobial susceptibility, and toxinotype profiles of C. difficile isolated from pigs and their environment in the indoor conventional and outdoor antimicrobial free (ABF) production systems. Ten conventional and eight ABF cohorts of 35 pigs each and their environment were sampled at different stages of production at farm and slaughter. C. difficile prevalence in pigs was highest at the farrowing stage in both conventional (34%, 120/350) and ABF (23%, 56/244) systems, and decreased with age. This reduction in C. difficile prevalence in pigs at later stages of production mirrored the decreased prevalence in the farm environment. At slaughter, C. difficile was isolated at a low frequency from the carcasses and processing environment in both production systems. All but three isolates were resistant to ciprofloxacin (99%, 505/508), while 1.0% (5/508) and 6.0% (23/508) of isolates exhibited resistance to tetracycline and erythromycin, respectively. Toxinotype V (tcdA+tcdB+) was the predominant strain identified in both systems (conventional: 94%, 376/401; ABF: 82%, 88/107), while the rest were toxinotype XIII (tcdA+tcdB+). To conclude, we isolated antimicrobial resistant C. difficile regardless of antimicrobial use on the farm. Based on the phenotypic and genotypic similarity of C. difficile isolated in this study, we conclude that the unique production practices employed in conventional and ABF production systems have no impact on the pathogen population.

This chapter discusses the various phenotyping methods that are currently used to characterize foodborne pathogens. The methods that are predominantly used and described in this chapter include serotyping, phage typing, biotyping, antibiotyping and multi-locus enzyme electrophoresis. In this chapter, the principles of each of these methods, technical aspects, applications, interpretation of findings, strengths and weaknesses are described. Further reading on less common phenotypic methods is included as a concluding remark.

SummaryThe aim of this study was to determine the phenotypic and genotypic diversity of multiple Campylobacter isolates (n = 3 per sample) present within individual (heterogeneity) pig faecal and carcass samples at farm and slaughter, respectively. We isolated 1459 Campylobacter coli (1110 on farm and 349 from slaughter) from 908 pigs and 757 carcasses and characterized them for their antimicrobial susceptibility profile to a panel of six antimicrobials using the agar dilution method. Overall, we detected a significantly higher Campylobacter prevalence at the farm (54.7%) than at slaughter (19%) level (P < 0.05). C. coli isolates were resistant most commonly to tetracycline (66.2%) and erythromycin (53.6%) while fluoroquinolone resistance was detected in isolates (n = 17) only from the farm level. Phenotypic diversity of C. coli isolates at the 4‐fold minimum inhibitory concentration levels within the same sample was detected in 38.6% (n = 192) pigs and 40.2% (n = 58) carcass swabs with no significant difference between the two sources (P = 0.72). Phenotypic heterogeneity based on the antimicrobial resistance patterns was observed in 32.5% (n = 162) of the farm samples and in 30.5% (n = 44) carcass swabs at slaughter (P = 0.64). A subset of 40 isolates representing ten pigs and eight carcass samples (originating from separate pigs) were further genotyped by multi locus sequence typing. The observation of phenotypic diversity was replicated at the genotypic level, as it was highlighted by the 22 sequence types which represented the 40 isolates. In conclusion, we detected multiple C. coli subtypes from individual pig or carcass samples indicating unprecedented level of heterogeneity. Our study clearly signifies the importance of testing multiple colonies to make appropriate and valid conclusions in epidemiological‐based studies.

To determine the antimicrobial resistant profiles and clonality of Campylobacter coli isolated from clinically ill humans and retail meats.A total of 98 C. coli isolates (20 from humans and 78 from retail meats) were phenotypically characterized. Antimicrobial susceptibility testing was done using agar dilution method for ciprofloxacin, gentamicin, erythromycin and doxycycline. Seventy C. coli isolates including humans (n = 20) and retail meats (n = 50) were genotyped by multilocus sequence typing (MLST) and pulsed-field gel electrophoresis (PFGE). Resistance to ciprofloxacin was found in 29% and 15% of isolates from retail meats and humans. We observed 61 PFGE profiles using two enzymes (SmaI, KpnI) with an Index of discrimination of 0.99, whereas MLST generated 37 sequence types. Two clonal complexes were identified with 58 (82%) C. coli isolates clustered in the ST-828 complex.Resistance to ciprofloxacin and erythromycin was identified in C. coli obtained from retail meats and ill humans. PFGE typing of C. coli isolates was more discriminatory than MLST. Grouping of C. coli isolates (82%) by MLST in ST-828 clonal complex indicates a common ancestry.A high frequency of resistance found to ciprofloxacin and erythromycin is concerning from food safety perspective. PFGE using single or double restriction enzymes was found to be more discriminatory than MLST for genotyping C. coli. Overall, the C. coli populations recovered from humans and retail meats were genotypically diverse.

SUMMARY For epidemiological investigations of the most common and non-host-adapted Salmonella serotypes, such as Typhimurium, highly discriminatory approaches are essential. In the present study, we evaluated three genotyping methods; amplified fragment length polymorphism (AFLP), pulsed-field gel electrophoresis (PFGE) and repetitive palindromic extragenic–PCR (Rep–PCR) using 40 isolates. AFLP showed the highest discriminatory index (0·939), resolution and throughput. To determine clonality of Salmonella Typhimurium isolates and epidemiological relatedness in different commercial pig production units, we employed AFLP in combination with antimicrobial resistance pattern and phage typing. Salmonella serovar Typhimurium isolates (n=196) obtained from a longitudinal study of 18 pig farms over a 3-year period were studied. Using this approach, 16 distinct clonal types were identified. We found two common multidrug- resistant patterns including AmCmStSuTe and AmKmStSuTe. Two commonly multidrug- resistant phage types that are of known public health importance, DT104 and DT193, were also common. AFLP differentiated distinct clones within DT104, a phage type previously reported to be clonal. Fourteen of the clonal types were unique to one of the two production systems, showing diversity between independent commercial pig production systems located in the same geographical area. Clonal types obtained from nursery farms and corresponding finishing units were, however, similar.

Conventional swine production evolved to routinely use antimicrobials, and common occurrence of antimicrobial-resistant Salmonella has been reported. There is a paucity of information on the antimicrobial resistance of Salmonella in swine production in the absence of antimicrobial selective pressure. Therefore, we compared the prevalence and antimicrobial resistance of Salmonella isolated from antimicrobial-free and conventional production systems. A total of 889 pigs and 743 carcasses were sampled in the study. Salmonella prevalence was significantly higher among the antimicrobial-free systems (15.2%) than the conventional systems (4.2%) (odds ratio [OR] = 4.23; P < 0.05). Antimicrobial resistance was detected against 10 of the 12 antimicrobials tested. The highest frequency of resistance was found against tetracycline (80%), followed by streptomycin (43.4%) and sulfamethoxazole (36%). Frequency of resistance to most classes of antimicrobials (except tetracycline) was significantly higher among conventional farms than antimicrobial-free farms, with ORs ranging from 2.84 for chloramphenicol to 23.22 for kanamycin at the on-farm level. A total of 28 antimicrobial resistance patterns were detected. A resistance pattern with streptomycin, sulfamethoxazole, and tetracycline (n = 130) was the most common multidrug resistance pattern. There was no significant difference in the proportion of isolates with this pattern between the conventional (19.5%) and the antimicrobial-free systems (18%) (OR = 1.8; P > 0.05). A pentaresistance pattern with ampicillin, chloramphenicol, streptomycin, sulfamethoxazole, and tetracycline was strongly associated with antimicrobial-free groups (OR = 5.4; P = 0.01). While showing the higher likelihood of finding antimicrobial resistance among conventional herds, this study also implies that specific multidrug-resistant strains may occur on antimicrobial-free farms. A longitudinal study with a representative sample size is needed to reach more conclusive results of the associations detected in this study.

Antimicrobial resistant strains of bacteria are an increasing threat to animal and human health. Resistance mechanisms to circumvent the toxic action of antimicrobials have been identified and described for all known antimicrobials currently available for clinical use in human and veterinary medicine. Acquired bacterial antibiotic resistance can result from the mutation of normal cellular genes, the acquisition of foreign resistance genes, or a combination of these two mechanisms. The most common resistance mechanisms employed by bacteria include enzymatic degradation or alteration of the antimicrobial, mutation in the antimicrobial target site, decreased cell wall permeability to antimicrobials, and active efflux of the antimicrobial across the cell membrane. The spread of mobile genetic elements such as plasmids, transposons, and integrons has greatly contributed to the rapid dissemination of antimicrobial resistance among several bacterial genera of human and veterinary importance. Antimicrobial resistance genes have been shown to accumulate on mobile elements, leading to a situation where multidrug resistance phenotypes can be transferred to a susceptible recipient via a single genetic event. The increasing prevalence of antimicrobial resistant bacterial pathogens has severe implications for the future treatment and prevention of infectious diseases in both animals and humans. The versatility with which bacteria adapt to their environment and exchange DNA between different genera highlights the need to implement effective antimicrobial stewardship and infection control programs in both human and veterinary medicine.

The objectives of this study were to determine and compare the prevalence and antimicrobial resistance of Campylobacter species in swine reared in conventional and antimicrobial-free (ABF) production systems. Campylobacter coli was the predominant species, with 1,459 isolates (99%) in the study. We found significantly higher prevalence of C. coli on the ABF farms (77.3%) than on the conventional farms (27.6%) among pigs at the nursery stage (P < 0.001). At slaughter, we found significantly higher prevalence at the postevisceration than at the preevisceration stage (P < 0.001) in both production systems. The 1,459 C. coli isolates were tested with the agar dilution method for their susceptibility to six antimicrobials: chloramphenicol, ciprofloxacin, erythromycin, gentamicin, nalidixic acid, and tetracycline. Resistance was most prevalent against tetracycline (66.2% of isolates) followed by erythromycin (53.6% of isolates). Frequency of resistance to these two antimicrobials was significantly higher among conventional herds (83.4% for tetracycline and 77% for erythromycin) than among ABF herds (56.2% for tetracycline and 34.5% for erythromycin). Resistance to ciprofloxacin at the MIC (> 4 mg/liter) was also found on farms in both systems. Multidrug-resistant C. coli strains were detected in both the conventional (7%) and ABF (4%) herds. This is the first report of ciprofloxacin-resistant strains of C. coli in ABF pigs in the United States. These findings highlight the high prevalence of antimicrobial-resistant C. coli in both conventional and ABF pig production systems and have significant implications for the persistence of antimicrobial-resistant Campylobacter in the pig production environment regardless of levels of antimicrobial use.

OBJECTIVES:To determine the prevalence and antimicrobial resistance of Campylobacter species in swine reared in the intensive and extensive antimicrobial-free (ABF) production systems at farm and slaughter. In the ABF system, antimicrobials are neither used for growth promotion nor therapeutic purposes. METHODS:Swine faecal and carcass swabs were collected from 10 groups of pigs (five each from intensive and extensive ABF farms) at the finishing farm and the slaughter plant. A total of 292 pigs at farm (extensive 118; intensive 174) and 254 carcass swabs (extensive 134; intensive 120) were collected during the study. Campylobacter species were isolated under microaerobic conditions and confirmed by biochemical testing. Up to three presumptive Campylobacter colonies per positive pig/carcass were further characterized. Speciation was done by PCR, targeting ceuE and hipO genes for Campylobacter coli and Campylobacter jejuni, respectively. The isolates were tested for their antimicrobial resistance profile using the agar dilution method against six antimicrobials. RESULTS:A total of 526 Campylobacter isolates were cultured from 292 pigs and 254 carcasses sampled. All the isolates were found to be C. coli. Overall prevalence of C. coli was 55.8% on farm (55% extensive and 56.3% intensive) and 26% at slaughter (32.8% extensive and 18.3% intensive). There was no significant difference in C. coli between the intensive and extensive systems on the finishing farms (P = 0.83). At post-chill stage, C. coli were isolated only from the extensively reared ABF pigs. Antimicrobial resistance against ciprofloxacin (MIC > 4 mg/L) was found at the farm level in both the intensive- and extensive-reared groups. The erythromycin/nalidixic acid/tetracycline resistance pattern (3%) was the most common pattern in multidrug-resistant C. coli. CONCLUSIONS:This study highlights the high prevalence of diverse and antimicrobial-resistant C. coli in the ABF production systems of swine. This is the first study reporting the isolation of ciprofloxacin-resistant strains from ABF pigs in the USA and warrants concern.