Effect of poultry litter soil amendment on antibiotic-resistant Escherichia coli.

Given the high cost and non-renewability of mineral-based fertilizers, there is increasing interest in the innovative use of manure-based materials, such as poultry litter (PL). However, manure-based fertilizers add both nutrients and microbes to the soil, including antibiotic-resistant Escherichia coli (AREc). PL soil amendment impact on AREc in corn fields was evaluated in a randomized field experiment (May-October 2017). Two winter cropping systems (fallow and cover crop) were assigned to whole plots, with three spring-applied fertilizer treatments (untreated control [UC], PL, and commercial fertilizer [CF]) assigned to subplots. Soil was collected from 0 to 15 cm on days 0, 7, 28, 70, 98, and 172 post-treatment applications. Samples were cultured for the enumeration and prevalence of generic, tetracycline-resistant (TETr), third-generation cephalosporin-resistant (3GCr) E. coli isolates, and extended spectrum beta-lactamase (ESBL)-producing Enterobacteriaceae. PL soil amendment significantly (p < 0.05) increased the levels of generic E. coli, TETr E. coli, and 3GCr E. coli on days 7 and 28 compared to UC or CF. Beyond day 28, AREc did not significantly (p > 0.05) differ by fertilizer treatment and returned to baseline on day 70. ESBL-producing Enterobacteriaceae were detected from 16 samples, mostly on day 70. Cover crop significantly decreased TETr E. coli concentration on day 28, with no significant effects on the prevalence of 3GCr E. coli and ESBL-producing Enterobacteriaceae compared to no cover crop. All ESBL-producing Enterobacteriaceae and 79% of the 3GCr E. coli isolates were positive for blaCTX-M gene by polymerase chain reaction. Results show that PL soil amendment transiently increases the levels of AREc compared to mineral fertilizer.

Efficacy of novel staphylococcal surface associated protein vaccines against Staphylococcus aureus and non-aureus staphylococcal mastitis in dairy cows.

Mastitis is an inflammation of the mammary gland commonly caused by bacteria or fungi. Staphylococcus aureus is a major bacterium that causes mastitis in dairy cows. Non-aureus staphylococci are also increasingly reported, with Staphylococcus chromogenes being the most common species. Current staphylococcal mastitis control programs are not fully effective, and treatment with antibiotics is not sustainable. Non-antibiotic sustainable control tools, such as effective vaccines, are critically needed. We previously developed S. aureus surface-associated proteins (SASP) and S. chromogenes surface-associated proteins (SCSP) vaccines that conferred partial protective effects. We hypothesized that vaccination with SASP or SCSP would reduce the incidence of S. aureus mastitis throughout the lactation period. The objective of this study was to evaluate the efficacy of SASP and SCSP vaccines against S. aureus and non-aureus staphylococcal mastitis under natural exposure over 300 days of lactation. Pregnant Holstein dairy cows (n = 45) were enrolled and assigned to receive SASP (n = 15) or SCSP (n = 16) vaccines or unvaccinated control (n = 14). Cows were vaccinated with 1.2 mg of SASP or SCSP with Emulsigen-D adjuvant. Control cows were injected with phosphate-buffered saline with Emulsigen-D adjuvant. Three vaccine injections were given subcutaneously at 60, 40, and 20 days before the expected calving. Booster vaccinations were given at 120 and 240 days in milk. Cows were monitored for mastitis at quarter and cow levels, staphylococcal mastitis incidence, changes in serum and milk anti-SASP and anti-SCSP antibody titers, bacterial counts in milk, adverse reactions, milk yield and milk somatic cells count over 300 days of lactation. The SCSP vaccine conferred a significant reduction in the incidence of staphylococcal mastitis. Milk and serum anti-SASP and anti-SCSP antibody titers were increased in the vaccinated cows compared to unvaccinated control cows. Anti-SASP and anti-SCSP antibody titers decreased at about 120 days in milk, indicating the duration of immunity of about four months. In conclusion, the SASP and SCSP vaccines conferred partial protection from natural infection.

Microbiome Diversity of Anaerobic Digesters Is Enhanced by Microaeration and Low Frequency Sound.

Biogas is produced by a consortium of bacteria and archaea. We studied how the microbiome of poultry litter digestate was affected by time and treatments that enhanced biogas production. The microbiome was analyzed at six, 23, and 42 weeks of incubation. Starting at week seven, the digesters underwent four treatments: control, microaeration with 6 mL air L-1 digestate per day, treatment with a 1000 Hz sine wave, or treatment with the sound wave and microaeration. Both microaeration and sound enhanced biogas production relative to the control, while their combination was not as effective as microaeration alone. At week six, over 80% of the microbiome of the four digesters was composed of the three phyla Actinobacteria, Proteobacteria, and Firmicutes, with less than 10% Euryarchaeota and Bacteroidetes. At week 23, the digester microbiomes were more diverse with the phyla Spirochaetes, Synergistetes, and Verrucomicrobia increasing in proportion and the abundance of Actinobacteria decreasing. At week 42, Firmicutes, Bacteroidetes, Euryarchaeota, and Actinobacteria were the most dominant phyla, comprising 27.8%, 21.4%, 17.6%, and 12.3% of the microbiome. Other than the relative proportions of Firmicutes being increased and proportions of Bacteroidetes being decreased by the treatments, no systematic shifts in the microbiomes were observed due to treatment. Rather, microbial diversity was enhanced relative to the control. Given that both air and sound treatment increased biogas production, it is likely that they improved poultry litter breakdown to promote microbial growth.

Dynamics of Extended-spectrum Beta-lactamase-producing, Third-generation Cephalosporin-resistant and Tetracycline-resistant Escherichia coli in Feedlot Cattle With or Without Tylosin Administration.

The impact of in-feed use of tylosin in feedlot cattle on Gram-negative foodborne bacteria is unknown. We evaluated the effect of continuous in-feed tylosin use on the concentration and prevalence of tetracycline-resistant (TETr)-, third-generation cephalosporin-resistant (3GCr)-, and extended-spectrum ß-lactamase-producing (ESBLs) E. coli in feedlot cattle. A cohort of weaned calves (10 animals/group) were randomized to receive a feed ration with or without tylosin. Fecal samples, regularly collected over the entire feeding period, and pen surface and feed samples, collected at the end of the feeding period, were cultured on selective media. Enumeration and binary outcomes were analyzed by mixed effects linear regression or logistic regression, respectively, using treatment and days on feed as fixed factors, and animal ID as a random variable. Tylosin supplementation did not affect the fecal concentrations of TETrE. coli or fecal prevalence of 3GCrE. coli. However, cattle in the tylosin group were 1.5 times more likely (Odds ratio = 1.5: 95% confidence interval: 1.1-2.0) to harbor ESBLs E. coli than the control cattle. Regardless of tylosin treatment, fecal concentrations of TETrE. coli and the prevalence of 3GCr- and ESBLs-E. coli increased over time. Tylosin-supplemented feed did not affect the prevalence of TETrE. coli; 3GCr and ESBLs-E. coli were not detected from the feed samples. Most of the 3GCr- and ESBLs-E. coli isolates carried the blaCTX-M-15 gene, widely detected among ESBLs-E. coli human isolates. In summary, although in-feed tylosin use in feedlot cattle did not select for TETr- and 3GCr-E. coli, it increased the likelihood of detecting ESBL-producing E. coli. Furthermore, the study indicated that the feedlot production setting gradually increases the levels of E. coli resistant to the critically and/or important antibiotics for public health, indicating an increased risk of their dissemination beyond the feedlot environment.

Effect of continuous in-feed administration of tylosin to feedlot cattle on macrolide and tetracycline resistant enterococci in a randomized field trial.

Liver abscess causes substantial economic loss to the beef cattle industry through liver condemnation, reduced animal performance, and carcass yield. Continuous in-feed use of tylosin is the most effective and a commonly used practice in beef cattle production to prevent liver abscess. However, such mass medication can increase the level of antimicrobial resistant bacteria. We investigated the effect of continuous in-feed use of tylosin in feedlot cattle on (i) concentrations and prevalence of erythromycin-resistant (ERYr) and tetracycline-resistant (TETr) enterococci; (ii) associated antimicrobial resistance genes (ARGs) for resistance; (iii) species distribution; iv) macrolide and tetracycline resistance gene concentrations; and (v) tylosin concentration. A cohort of weaned calves were randomized to receive tylosin-medicated feed (Tylosin; n = 10) or nonmedicated feed (Control; n = 10) for a full feedlot cycle. Feces, feed and pen-surface samples were collected and processed by culture, droplet digital PCR, and liquid chromatography/mass spectroscopy for bacterial enumeration, detection and characterization, ARG quantification, and tylosin concentration, respectively. Data were analyzed by mixed effects linear- or binary-regression models depending on the outcomes. Tylosin administration significantly increased fecal concentration (P < 0.001) and prevalence (P = 0.021) of ERYr enterococci and erm(B) gene concentration (P < 0.001), compared to the control group. Interestingly, tylosin administration significantly reduced (P = 0.037) fecal TETr enterococci concentration compared to the control group, with no significant effect (P = 0.758) on fecal tet(M) concentration. In both treatment groups, enterococci concentrations increased over time, peaking on 174 days in feed before returning to the baseline. ERYr enterococci concentration was significantly (P = 0.012) higher in tylosin medicated feeds, with no significant effect (P = 0.321) on TETr enterococci concentration. Pen-surface concentration of ermB was significantly (P = 0.024) higher in the tylosin group, with no significant effect (P > 0.05) on bacterial concentrations. Increased diversity and a shift in the composition of enterococcal species and ARGs were observed over time, although tylosin use did not significantly affect (P > 0.05) their prevalence. Tylosin concentration was significantly higher in the feces of tylosin administered cattle (P < 0.001) and medicated feed (P = 0.027), with numerically higher pen-surface concentration (P = 0.065) in the tylosin group. In conclusion, continuous in-feed use of tylosin in feedlot cattle increases macrolide resistant enterococci and its fecal excretion, while decreasing tetracycline resistance. Two medically important species, E. faecium and E. faecalis, were predominant regardless of resistance status or sample source. Risk-based approaches including label changes to limit tylosin use such as withdrawal period, and development of effective manure treatments are potential areas of research to reduce environmental and public health impacts.

Occurrence and prevalence of antimicrobial resistance in urban karst groundwater systems based on targeted resistome analysis.

Antimicrobial resistance (AMR) is a global crisis threatening human, animal, and environmental health. The natural environment, specifically water resources, has been recognized as a reservoir and dissemination pathway for AMR; however, urban karst aquifer systems have been overlooked. This is a concern as these aquifer systems provide drinking water to about 10 % of the global population; yet, the urban influence on the resistome in these vulnerable aquifers is sparingly explored. This study used high-throughput qPCR to determine the occurrence and relative abundance of antimicrobial resistant genes (ARG) in a developing urban karst groundwater system in Bowling Green, KY. Ten sites throughout the city were sampled weekly and analyzed for 85 ARGs, as well as seven microbial source tracking (MST) genes for human and animal sources, providing a spatiotemporal understanding of the resistome in urban karst groundwater. To further understand ARGs in this environment, potential drivers (landuse, karst feature type, season, source of fecal pollution) were considered in relation to the resistome relative abundance. The MST markers highlighted a prominent human influence to the resistome in this karst setting. The concentration of targeted genes varied between the sample weeks, but all targeted ARGs were prevalent throughout the aquifer regardless of karst feature type or season, with high concentrations captured for sulfonamide (sul1), quaternary ammonium compound (qacE), and aminoglycoside (strB) antimicrobial classes. Higher prevalence and relative abundance were detected during the summer and fall seasons, as well as at the spring features. Linear discriminant analysis suggested that karst feature type had higher influence on ARGs in the aquifer compared to season and the source of fecal pollution had the least influence. These findings can contribute to the development of effective management and mitigation strategies for AMR.

Prevalence and whole-genome sequencing characterization of Salmonella in urban karst groundwater and predominantly groundwater-fed surface waters for serotypes and antimicrobial resistance.

Karst aquifers provide a significant source of drinking water around the world; however, they are prone to bacterial contamination. We investigated the prevalence, serotypes, and antimicrobial resistance genes (ARGs) of Salmonella in urban karst groundwater and groundwater-fed surface waters. Weekly water samples (n = 443) were collected from nine groundwater sites and the Barren River in Bowling Green, KY. Additionally, cross-sectional samples were collected from 45 sites, including groundwaters and two rivers, in Tampa, FL. Salmonella was detected in 14.5% of Bowling Green samples resulting in 176 isolates representing 18 serotypes. Two (4.4%) of the Tampa samples were positive yielding eight isolates representing two serotypes. In Bowling Green, Salmonella prevalence varied by sampling site, week, month, and season, with the highest prevalence in surface water in the month of November and during summer. Salmonella Newport was the most common serotype detected. Nine of the serotypes detected in the study were among the top 20 serotypes commonly associated with human infections in the United States. Isolates were clustered with human clinical isolates, or isolates obtained from food animals, suggesting the importance of humans and agricultural practices as a source of water contamination. ARGs were detected in 12.5% (n = 176) of the Bowling Green isolates; overall, 81.8% (n = 22) of these were multidrug-resistant (resistance to ≥3 antimicrobial classes). Notably, the four Salmonella Agona isolates were resistant to four antimicrobial classes, mercury, and quaternary ammonium compounds. Seasonality and the widespread occurrence of Salmonella both in the groundwater and groundwater-fed surface waters is a potential risk to public health.

Prevalence and Antimicrobial Resistance of Escherichia coli O157:H7 and Salmonella, and the Prevalence of Staphylococcus aureus in Dairy Cattle and Camels under Pastoral Production System.

Escherichia coli O157:H7, Salmonella and Staphylococcus aureus are common foodborne pathogens. We determined the prevalence of E. coli O157:H7 and Salmonella in feces and milk and the prevalence of S. aureus in milk from dairy cattle and camels in the Borana pastoral community in the Southern Oromia Region of Ethiopia. Paired individual cow composite (pooled from all quarters in equal proportions) milk and fecal samples were collected from cows (n = 154) and camels (n = 158). Samples were cultured on bacterial isolation and identification media. E. coli O157:H7 and Salmonella isolates were further tested for susceptibility against nine antimicrobial drugs. Different risk factors associated with hygienic milking practices were recorded and analyzed for their influence on the prevalence of these bacteria in milk and feces. The prevalence of E. coli O157:H7 and Salmonella in feces were 3.9% and 8.4%, respectively, in cows, and 0.6% and 2.5%, respectively, in camels. E. coli O157:H7 and Salmonella were detected in the composite milk samples of 2.6% and 3.9% of the cows, respectively, and 0% and 1.3% of the camels, respectively. S. aureus was detected in composite milk samples of 33.4% of the cows and 41.7% of the camels. All E. coli O157:H7 (n = 11) and Salmonella (n = 25) isolates from both animal species and sample types were resistant to at least one antimicrobial drug. Multidrug resistance was observed in 70% (7/10) of the E. coli O157:H7 fecal and milk isolates from cows and 33.3% (2/6) of the Salmonella fecal and milk isolates from camels. The prevalence of these bacteria in feces and milk was not affected by risk factors associated with milking practices. Given the very close contact between herders and their animals and the limited availability of water for hand washing and udder cleaning, these bacteria are most likely present in all niches in the community. Improving community awareness of the need to boil milk before consumption is a realistic public health approach to reducing the risk of these bacteria.

Hygienic assessment of fish handling practices along production and supply chain and its public health implications in Central Oromia, Ethiopia.

Fishborne diseases are among the major causes of morbidity and mortality worldwide. Contamination of the aquatic ecosystem and unhygienic handling practices along the fish supply chain can lead to a contaminated fish. Consumption of raw or under cooked fish and fish products is a major source of fishborne infections in humans. Despite reports of fish contamination with foodborne pathogens in Ethiopia, information regarding the hygienic status of fish handling practices is limited. We assessed fish hygienic handling practices at production sites and along the fish supply chain in three towns in east Shewa zone of Oromia. Data were collected using a semi-structured questionnaire interviews and personal observations. The study consisted of purposively selected respondents comprising of 50 fishermen, 10 retailers, 20 food establishments serving fish, and 120 consumers. Descriptive statistics and Chi-square test were used to present the proportion of various actors along the fish production and supply chain and to compare the proportions of observations among the different categories respectively. We observed that the lakes were accessible to animals and exposed to chemical and microbial contaminations through rainwater run-off. Fish were processed under unhygienic practices like washing of filleted fish with lake water, indiscriminate processing at unhygienic landing sites, use of a single knife for processing all fish with infrequent washing and with no disinfection in between. Majority (70%; n = 10) of the retailers and all the food establishments transported fish in vehicles with no cold chain facilities. Good hygienic practices we observed were the use of refrigerators for storage in all retailers and 70% (n = 20) of the food establishments; 30% of retailers used vehicles with a cold chain facility for the transportation of fish. Over three-fourths (77%; n = 120) of the consumers preferred consuming raw fish; 80% of them lacked the knowledge of fishborne diseases. The study revealed a wide range of unhygienic handling practices along fish production and supply chain; lack of infrastructure for post-harvest fish handling and processing, lack of appropriate transportation facilities and presence of knowledge gaps regarding fish borne diseases.

Tetracycline-Resistant, Third-Generation Cephalosporin-Resistant, and Extended-Spectrum ß-Lactamase-Producing Escherichia coli in a Beef Cow-Calf Production System.

ABSTRACT: Cow-calf production plays a significant role in the beef production chain. However, bacteria in these systems are not typically monitored for antimicrobial resistance (AMR). We determined the baseline level of AMR in fecal bacteria collected from preweaned calves prior to feedlot entry and evaluated the effects of type of graze and age on AMR occurrence. Two grazing experiments (16 cow-calf pairs each) were conducted on tall fescue or wheat. Fecal samples were cultured for the detection of tetracycline-resistant (TETr), third-generation cephalosporin-resistant (3GCr), and extended-spectrum ß-lactamase (ESBL)-producing Escherichia coli. Isolates were characterized for resistance to other antibiotics and resistance mechanisms. Concentrations (P < 0.001) and prevalence (P = 0.007) of TETrE. coli isolates were significantly higher in the calves (5.1 log CFU/g and 93%, respectively) than in the cows (4.4 log CFU/g and 80%, respectively). Wheat grazing did not affect TETr isolates phenotypically; however, it significantly expanded (P = 0.005) the resistant population carrying tet(A) over that carrying tet(B). Fecal prevalence of 3GCr and ESBL-producing isolates was 31.3 and 3.4%, respectively, with no significant effects of age (P = 0.340) or wheat grazing (P = 0.597). All 3GCr and ESBL-producing isolates were multidrug resistant (resistant to at least three antimicrobial classes). 3GCr isolates were positive for blaCMY-2 (73%) or blaCTX-M (27%), and blaCTX-M-15 was the most prevalent gene (94%, n = 17) among the CTX-M-positive isolates. Wheat grazing significantly expanded (P < 0.001) the 3GCr population carrying blaCTX-M and reduced the population carrying blaCMY-2. Five of the seven ESBL-producing isolates were positive for blaCTX-M. Our study revealed age-dependent occurrence of TETrE. coli and that wheat grazing expanded the resistant population carrying certain resistance genes. Cow-calf production is a significant reservoir for antibiotic-resistant bacteria of significant public health importance such as 3GCr and CTX-M ESBL-producing E. coli.

Regulation Mechanisms of Virulence Genes in Enterohemorrhagic Escherichia coli.

Enterohemorrhagic Escherichia coli (EHEC) is one of the most common E. coli pathotypes reported to cause several outbreaks of foodborne illnesses. EHEC is a zoonotic pathogen, and ruminants, especially cattle, are considered important reservoirs for the most common EHEC serotype, E. coli O157:H7. Humans are infected indirectly through the consumption of food (milk, meat, leafy vegetables, and fruits) and water contaminated by animal feces or direct contact with carrier animals or humans. E. coli O157:H7 is one of the most frequently reported causes of foodborne illnesses in developed countries. It employs two essential virulence mechanisms to trigger damage to the host. These are the development of attaching and effacing (AE) phenotypes on the intestinal mucosa of the host and the production of Shiga toxin (Stx) that causes hemorrhagic colitis and hemolytic uremic syndrome. The AE phenotype is controlled by the pathogenicity island, the locus of enterocyte effacement (LEE). The induction of both AE and Stx is under strict and highly complex regulatory mechanisms. Thus, a good understanding of these mechanisms, major proteins expressed, and environmental cues involved in the regulation of the expression of the virulence genes is vital to finding a method to control the colonization of reservoir hosts, especially cattle, and disease development in humans. This review is a concise account of the current state of knowledge of virulence gene regulation in the LEE-positive EHEC.

Tetracycline- and Macrolide-Resistant Enterococcus Species Isolated from a Mink Farm in the United States.

Enterococcus species are a normal flora of animals and humans. However, life-threatening opportunistic infections can be caused by antimicrobial resistant strains. Fecal (n = 42) and feed (n = 8) samples were obtained from a mink farm and cultured for the enumeration and detection of erythromycin-resistant (a macrolide; ERYr)- and tetracycline-resistant (TETr) enterococci. ERYr and TETr enterococci were detected from all fecal (mean concentrations = 6 and 7 logs, respectively) and feed (mean concentrations = 5 and 4 logs, respectively) samples. While Enterococcus faecalis and Enterococcus faecium were detected at equal proportions among the fecal TETr isolates, E. faecium predominated among ERYr fecal isolates. All ERYr and 90% of the TETr isolates (n = 50) were multidrug resistant (resistant to three or more antimicrobial classes). Among ERYr isolates, while 83% of E. faecalis (n = 12) were positive for erm(B), 58% of E. faecium (n = 38) isolates were positive for msr(C). Among ERYr isolates, tet(M) was detected from 92% of E. faecalis (n = 12) and 97% of E. faecium (n = 38) isolates. Conversely, however, erm(B) was detected in 18% of E. faecalis (n = 22) and 33% of E. faecium (n = 27) TETr isolates. Our study provides a baseline for future efforts to reduce antimicrobial resistance and improve antimicrobial stewardship in commercial mink production facilities.

Prevalence of Antimicrobial Resistant and Extended-Spectrum Beta-Lactamase-producing Escherichia coli in Dairy Cattle Farms in East Tennessee.

Antimicrobials have been widely used in dairy farms to prevent and control dairy cattle diseases since 1960s. This led to the emergence of antimicrobial resistant bacteria (ARB) that, along with their antimicrobial resistance genes (ARGs), can spread from dairy farms to humans. Therefore, regular antimicrobial resistance (AMR) monitoring is important to implement proper mitigation measures. The objective of this study was to determine the prevalence of AMR and extended-spectrum beta-lactamases (ESBLs)-producing Escherichia coli in dairy cattle. A cross-sectional study was conducted in four dairy cattle farms (A-D) in East Tennessee. A total of 80 samples consisting of 20 samples each of bulk tank milk, feces, dairy cattle manure-amended soil, and prairie soil adjacent to the farms were collected and cultured for the isolation of E. coli. Tetracycline (TETr)-, third-generation cephalosporin (TGCr)- and nalidixic acid (NALr)-resistant E. coli (n = 88) were isolated and identified on agar media supplemented with TET, cefotaxime, and NAL, respectively. TGCr E. coli were tested for ESBLs and other coselected ARGs. TETr (74%, n = 88) was the most common, followed by TGCr (20%) and NALr (8%). Farms had significant (p < 0.001) differences: the highest prevalence of TGCr (55%) and TETr (100%) were observed in farm D, while all NALr isolates were from farm C. Over 83% of TGCr isolates (n = 18) harbored ESBL gene blaCTX-M. Majority (78%) of the E. coli isolates were multidrug-resistant (MDR), being positive for beta-lactams (blaCTX-M), TETs tet(A), tet(B), tet(M)), sulfonamides (sul2), aminoglycosides (strA), and phenicols (floR). This study indicated the widespread occurrence of MDR ESBLs-E. coli in dairy cattle farms. AMR surveillance of more dairy farms and identification of farm-level risk factors are important to mitigate the occurrence and spread of ARB of significant public health importance, such as ESBLs-E. coli.

Lagoon, Anaerobic Digestion, and Composting of Animal Manure Treatments Impact on Tetracycline Resistance Genes.

Increased demand for animal protein is met by increased food animal production resulting in large quantities of manure. Animal producers, therefore, need sustainable agricultural practices to protect environmental health. Large quantities of antimicrobials are used in commercial food animal production. Consequently, antimicrobial-resistant bacteria and the resistance genes emerge and are excreted through feces. Manure management is essential for the safe disposal of animal waste. Lagoons, with or without covers, and anaerobic digesters, with the primary purpose of methane production, and composting, with the primary purpose of producing organic fertilizer, are widely used methods of manure treatment. We reviewed manure management practices and their impact on tetracycline resistance genes. Lagoons are maintained at ambient temperatures; especially uncovered lagoons are the least effective in removing tetracycline resistance genes. However, some modifications can improve the performance of lagoons: sequential use of uncovered lagoons and the use of covered lagoons resulted in a one-log reduction, while post-treatments such as biofiltration following covered lagoon treatment resulted in 3.4 log reduction. Mesophilic digestion of animal manure did not have any significant effect; only a 0.7 log reduction in tet(A) was observed in one study. While thermophilic anaerobic digesters are effective, if properly operated, they are expensive for animal producers. Aerobic thermophilic composting is a promising technology if optimized with its economic benefits. Composting of raw animal manure can result in up to a 2.5 log reduction, and postdigestion composting can reduce tetracycline resistance gene concentration by >80%. In general, manure management was not designed to mitigate antimicrobial resistance; future research is needed to optimize the economic benefits of biogas or organic fertilizer on the one hand and for the mitigation of foodborne pathogens and antimicrobial resistance on the other.

Prevalence, Serotypes, and Antimicrobial Resistance of Salmonella from Mink Feces and Feed in the United States.

Nontyphoidal Salmonella is a leading cause of foodborne illnesses, and concentrated animal production such as commercial mink farming can be a reservoir. The objective of this study was to determine the prevalence, serotypes, antimicrobial resistance, virulence genes and mobile genetic elements, and phylogenetic characterization of Salmonella from mink fecal (n = 42) and feed (n = 8) samples at a commercial mink farm in the United States. Salmonella was detected from 11 (26.2%) fecal and 1 (12.5%) feed samples. Twelve fecal isolates obtained from the positive samples were identified into four serotypes: Salmonella Uganda (eight isolates; two isolates were obtained from one sample), Salmonella Reading (two isolates), Salmonella Heidelberg (one isolate), and Salmonella Orion (one isolate). Two isolates from the positive feed sample were identified as Salmonella Cerro. Twelve isolates, one isolate per positive sample, were whole genome sequenced for further characterization. The two Salmonella Reading isolates were multidrug resistant (MDR) with the classical ASSuT (ampicillin, streptomycin, sulfisoxazole, and tetracycline) phenotype. Genotypically, the isolates were correspondingly positive, both by polymerase chain reaction and by whole-genome sequencing (WGS), for blaTEM-1, aph(3″)-Ib and aph(6)-Id, sul2, and tet(A). WGS additionally identified trimethoprim resistance gene dfrA8 in Salmonella Reading, and fosfomycin resistance gene fosA7 in Salmonella Heidelberg. All isolates carried Salmonella-specific multidrug resistance (mdsA/B), gold resistance, and intimin-like virulence genes. In addition, Salmonella Orion carried copper and silver resistance gene clusters. The two Salmonella Reading isolates also carried a cytolethal distension toxin (cdtB) gene. Salmonella Reading isolates belonged to ST412, a predominant sequence type among turkey and human isolates obtained in relation to recent salmonellosis outbreaks in North America. Phylogenetically, Salmonella Uganda, Salmonella Heidelberg, and Salmonella Reading were mostly associated with historic human/clinical, and poultry and swine source isolates. This study indicates that mink production can be a reservoir for foodborne pathogens such as Salmonella with MDR serovars commonly associated with human illnesses.

Evaluation of Streptococcus uberis Surface Proteins as Vaccine Antigens to Control S. uberis Mastitis in Dairy Cows.

There is no effective vaccine against Streptococcus uberis mastitis in dairy cows. Objectives of this study were (1) to extract S. uberis surface proteins (SUSP) and determine immunoreactivity in vitro and (2) immunogenicity and efficacy in vivo. SUSP was extracted from S. uberis, and their immunoreactivity was tested by western blot. In total, 26 Jersey dairy cows were randomly divided into four groups. Groups 1, 2, and 3 were vaccinated subcutaneously with 4 mg, 1 mg, and 100 µg of SUSP, respectively, with Freund's incomplete adjuvant. Group 4 (control) was injected with placebo. S. uberis UT888 was infused into two contralateral quarters of each cow during early lactation. Somatic cell count (SCC), bacteria count in milk, and mastitis were monitored. Our results show that SUSP contains multiple protein bands, that ranged from 10 to 100 kDa. All vaccinates showed an increased anti-SUSP IgG antibody. The SCC of all experimentally infected quarters increased after challenge but slightly decreased after day 3 with no significant difference among groups. Milk bacterial count was significantly (p < 0.05) reduced in high and medium doses vaccinated groups than low and control groups. In conclusion, SUSP vaccine is immunogenic and showed a promising efficacy to control bovine S. uberis mastitis.

Prevalence of Mastitis and Antibiotic Resistance of Bacterial Isolates from CMT Positive Milk Samples Obtained from Dairy Cows, Camels, and Goats in Two Pastoral Districts in Southern Ethiopia.

A study was carried out from August 2017 to February 2018 on lactating dairy cows, one-humped dromedary camels, and goats to determine mastitis in the Bule Hora and Dugda Dawa districts of in Southern Ethiopia. Milk samples from 564 udder quarters and udder halves from 171 animals consisting of 60 dairy cows, 51 camels, and 60 goats were tested for mastitis. Sixty-four positive udder milk samples were cultured, and bacterial mastitis pathogens were isolated and identified. The antibiotic resistance of bacterial isolates from milk with mastitis was tested against nine antimicrobials commonly used in the study area. Cow- and quarter-level prevalence of mastitis in dairy cows, camels, and goats was 33.3%, 26.3%, and 25% and 17.6%, 14.5%, and 20%, respectively. In cattle, the prevalence was significantly higher in Dugda Dawa than in Bule Hora. Major bacterial isolates were coagulase-negative Staphylococcus species (39.1%), S. aureus (17.2%), S. hyicus (14.1%), and S. intermedius and Escherichia coli (9.4% each). In camels, udder abnormality and mastitis were significantly higher in late lactation than in early lactation. Mastitis tends to increase with parity in camels. E. coli isolates were highly resistant to spectinomycin, vancomycin, and doxycycline, whereas most S. aureus isolates were multidrug-resistant. Most of the rural and periurban communities in this area consume raw milk, which indicates a high risk of infection with multidrug-resistant bacteria. We recommend a community-focused training program to improve community awareness of the need to boil milk and the risk of raw milk consumption.

Detection of Extended-Spectrum Beta-Lactamase-Producing and Carbapenem-Resistant Bacteria from Mink Feces and Feed in the United States.

Antibiotic-resistant infections caused by extended-spectrum ß-lactamases (ESBLs) and carbapenemases are increasing worldwide. Bacteria resistant to extended-spectrum cephalosporins and last resort carbapenems have been reported from food animals and their environments. Other concentrated nonfood-producing animals such as mink farming can be a reservoir of bacteria resistant to these critically important antibiotics. The objective of this study was to determine the prevalence of ESBL-producing bacteria and carbapenem-resistant (CR) bacteria from mink fecal (n = 42) and feed (n = 8) samples obtained from a commercial mink farm in the United States. The most prevalent ESBL-producing bacteria identified from the fecal samples were Escherichia coli (93%), Klebsiella pneumoniae (76%), and Proteus species (88%). E. coli (100%) and K. pneumoniae (75%) were also the most prevalent ESBL-producing bacteria identified from feed samples. All ESBL E. coli isolates were resistant to penicillin and most cephem beta-lactam antibiotics. Among the ESBL E. coli isolates, co-resistance was observed to ciprofloxacin (33%) and gentamicin (28%) indicating multidrug resistance. ESBL E. coli isolates predominantly carried blaCTX-M-14 and blaCTX-M-15 genes. Although all feed K. pneumoniae isolates carried blaCTX-M-9, all fecal K. pneumoniae isolates carried blaSHV. CR Pseudomonas species (7%), Hafnia alvei (24%), and Myroides odoratimimus (9.5%) were detected from fecal samples. H. alvei (37.5%) was the only CR bacteria detected from the feed samples. All CR isolates were polymerase chain reaction negative for the tested carbapenemases that are commonly reported, which may indicate intrinsic rather than acquired resistance. This study indicates that mink production can be a reservoir for bacteria resistant to the highest priority critically important antibiotics for human health.

Assessment of Hygienic Practices in Beef Cattle Slaughterhouses and Retail Shops in Bishoftu, Ethiopia: Implications for Public Health.

Understanding the potential drivers of microbial meat contamination along the entire meat supply chain is needed to identify targets for interventions to reduce the number of meatborne bacterial outbreaks. We assessed the hygienic practices in cattle slaughterhouses (28 employees) and retail shops (127 employees) through face-to-face interviews and direct personal observations. At the slaughterhouses, stunning, de-hiding and evisceration in vertical position, carcass washing and separate storage of offal were the identified good practices. Lack of hot water baths, absence of a chilling room, infrequent hand washing, insufficiently trained staff and irregular medical check-up were practices that lead to unhygienic handling of carcasses. At the retail shops, cleaning equipment using soap and hot water (81%), storing unsold meat in refrigerators (92%), concrete floors and white painted walls and ceilings were good practices. Adjacently displaying offal and meat (39%), lack of a cold chain, wrapping meat with plastic bags and newspapers, using a plastic or wooden cutting board (57%), infrequent washing of equipment and floors, and inadequately trained employees were practices that could result in unhygienic handling of beef. Our study identified unhygienic practices both at the slaughterhouses and retail shops that can predispose the public to meatborne infections, which could be improved through training and implementation of quality control systems.

Efficacy of diminazene diaceturate and isometamidium chloride hydrochloride for the treatment of Trypanosoma evansi in mice model.

Diminazene diaceturate (DIM) and isometamidium chloride hydrochloride (ISMM) have been widely used for the treatment of animal trypanosomosis. We evaluated the efficacy of standard doses of DIM and ISMM followed by their double doses for the treatment of Trypanosoma evansi in experimentally infected mice. A T. evansi strain obtained from a naturally infected camel in Afar was used. 25 swiss white mice randomly divided in to five groups were inoculated with 0.2 mL of blood containing 103 trypanosomes. At the peak of parasitemia (≈ 2 weeks post infection), groups A and B were treated with the standard dose (3.5 mg/kg body weight [BWT]) of DIM; groups C and D were treated with the standard dose (0.5 mg/kg BWT) of ISMM; and group E served as infected control. In the DIM standard dose groups, relapses and peak parasitemia were observed 20- and 25-days post treatment respectively. Similarly, relapses and peak parasitemia were observed 21- and 27-days post treatment in the ISMM standard dose groups. All mice in the control group died within two weeks post infection. Following relapses, mice were treated with the double doses of DIM (7 mg/kg BWT) or ISMM (1 mg/kg BWT). Parasitemia was not detected for 3 months following the double dose treatments. Following dexamethasone administration for 7 days, all but one mouse in the DIM group remained negative for another month. In general, although the T. evansi strain was resistant to the standard doses of DIM and ISMM their double doses completely cleared the infection.