Per- and polyfluoroalkyl substances: using comparative medicine to understand exposure and adverse health outcomes in people and their pets.

One of the important human health benefits of keeping pets may be to serve as an early warning system for indoor childhood exposure to toxic chemicals such as per- and polyfluoroalkyl substances (PFAS). The stain-resistant properties and environmental stability of PFAS make them a preferred choice for protective coatings and lubricants, and they have been used for years in various manufacturing and industrial processes around the world. Although the use of PFAS has arguably improved many commercial products, they have been linked to adverse health outcomes such as developmental delays, liver damage, immune suppression, disruption of endocrine and reproductive systems, and some cancers. The current body of literature suggests that serum PFAS levels in dogs and cats are analogous to their human counterparts and that household pets experience similar changes in blood chemistry markers. The proximity of small children and household pets to PFAS-treated carpets and floors, in addition to their tendency to put things into their mouths, potentially allows pets to serve as sentinels for household PFAS exposure. To assess the suitability of pets as indicators for exposure, researchers need to understand the most likely sources of PFAS exposure for household pets and identify the biomarkers of biological effects in those animals. Understanding these parameters may alert veterinary clinicians to potential sources of contamination in the home and ultimately protect the lives of the children and animals who live there.

Current per- and polyfluoroalkyl substance (PFAS) research points to a growing threat in animals.

Per- and polyfluoroalkyl substances (PFAS) are man-made chemicals that are colorless, odorless, and oil and water repellent. Their widespread use in manufacturing and industrial processes has resulted in environmental contamination found across the world. Exposure to PFAS can lead to a variety of adverse human health outcomes such as increased cholesterol, liver damage, immune suppression, and disruption of endocrine and reproductive systems. Exposure to this family of chemicals is considered a significant public health threat. Though nearly every human and animal around the world has been exposed, most of what is known regarding health effects and toxicological processes of PFAS in animals stems from human epidemiological and laboratory animal studies. Discoveries of PFAS contamination on dairy farms and concerns for companion animals have increased interest in PFAS research related to our veterinary patients. In the limited studies published to date, PFAS has been demonstrated in serum, liver, kidneys, and milk of production animals and has been linked to changes in liver enzymes, cholesterol levels, and thyroid hormones in dogs and cats. This is further addressed in the companion Currents in One Health by Brake et al, AJVR, April 2023. There is a gap in understanding the routes of exposure, absorption of PFAS, and adverse health effects among our veterinary patients. The purpose of this review is to summarize the current literature on PFAS in animals and discuss the implications for our veterinary patients.

Seromonitoring of brucellosis in goats and sheep slaughtered at an abattoir in Kampala, Uganda.

We conducted a cross-sectional serologic study at Kampala City abattoir in Uganda on 287 small ruminants (221 goats and 66 sheep) to determine the seroprevalence of brucellosis. The samples were tested using a modified rose bengal test (mRBT) and an indirect ELISA (iELISA). Small ruminant Brucella spp. seropositivity was 18 of 287 (6.3%) by mRBT and 19 of 287 (6.6%) by iELISA. The prevalence of brucellosis by mRBT was non-significantly higher in goats (17 of 221; 7.7%) than in sheep (1 of 66, 1.5%; p = 0.069), and also non-significantly higher by the iELISA in goats (18 of 221; 8.1%) than in sheep (1 of 66, 1.5%; p = 0.057). Brucellosis in slaughtered goats and sheep is a public health hazard to abattoir workers and consumers that calls for control and eradication measures at the farm level, given that testing is not carried out routinely at slaughter points.

Study on supplemental test to improve the detection of bovine tuberculosis in individual animals and herds.

BACKGROUND: Bovine tuberculosis (bTB), is a worldwide disease caused by Mycobacterium bovis (M. bovis). The success of bTB eradication and control programs is based on early detection and the removal of reactors from a herd thus routine testing and cull strategy have been applied globally. Since the late nineteenth century, the Tuberculin Skin Test (TST) has been the primary antemortem test available to support bTB eradication campaigns. Due to the TST limitations in terms of Se and Sp, the credibility of the diagnosis is frequently questioned given the occurrence of false-positive and false-negative reactions, therefore, it is necessary to confirm reactive animals using other methods, ensuring the reliability of the diagnosis. The aim of this study was to evaluate the sensitivity and specificity of a multiplex enzyme-linked immunosorbent assay (ELISA) relative to the tuberculin test used for the diagnosis of tuberculosis in cattle in Brazil. RESULTS: Lack of agreement between comparative cervical tuberculin test and ELISA IDEXX TM was observed. The 2 animals positive on the comparative cervical tuberculin test did not react at the ELISA IDEXX TM and 22 negative reactors by comparative cervical tuberculin test were positive by the ELISA IDEXX TM. The ELISA IDEXX TM showed sensitivity that is significantly lower than the official screening test the single cervical tuberculin. ELISA IDEXX TM also detected infected animals and herds undetected by the comparative cervical tuberculin test. The parallel use of comparative cervical tuberculin test and ELISA IDEXX TM increased sensitivity and the feasibility bTB screening. CONCLUSIONS: The results obtained here suggest that the ELISA IDEXX TM may be a supplemental test for the detection of Mycobacterium bovis infection in regions without routine testing and slaughter, where the disease generally progresses to more advanced stages and antibody responses are likely to be more prevalent. Evidence to support the validation of the ELISA IDEXX™ as a supplemental test for bTB eradication programs was provided.

Genetic Diversity and Potential Paths of Transmission of Mycobacterium bovis in the Amazon: The Discovery of M. bovis Lineage Lb1 Circulating in South America.

Bovine tuberculosis (bTB) has yet to be eradicated in Brazil. Herds of cattle and buffalo are important sources of revenue to people living in the banks of the Amazon River basin. A better understanding of Mycobacterium bovis (M. bovis) populational structure and transmission dynamics affecting these animals can significantly contribute in efforts to improve their sanitary status. Herein, we sequenced the whole genome of 22 M. bovis isolates (15 from buffalo and 7 from cattle) from 10 municipalities in the region of the Lower Amazon River Basin in Brazil and performed phylogenomic analysis and Single Nucleotide Polymorphism (SNP)-based transmission inference to evaluate population structure and transmission networks. Additionally, we compared these genomes to others obtained in unrelated studies in the Marajó Island (n = 15) and worldwide (n = 128) to understand strain diversity in the Amazon and to infer M. bovis lineages. Our results show a higher genomic diversity of M. bovis genomes obtained in the Lower Amazon River region when compared to the Marajó Island, while no significant difference was observed between M. bovis genomes obtained from cattle and buffalo (p ≥ 0.05). This high genetic diversity is reflected by the weak phylogenetic clustering of M. bovis from the Lower Amazon River region based on geographic proximity and in the detection of only two putative transmission clusters in the region. One of these clusters is the first description of inter-species transmission between cattle and buffalo in the Amazon, bringing implications to the bTB control program. Surprisingly, two M. bovis lineages were detected in our dataset, namely Lb1 and Lb3, constituting the first description of Lb1 in South America. Most of the strains of this study (13/22) and all 15 strains of the Marajó Island carried no clonal complex marker, suggesting that the recent lineage classification better describe the diversity of M. bovis in the Amazon.

Prevalence, distribution, and risk factors for canine blastomycosis in Michigan, USA.

Blastomycosis is a systemic fungal infection that most commonly affects dogs and humans. The disease is thought to be endemic in southern regions of Michigan, USA, but epidemiologic investigations have not been reported in detail for this state. The primary aims of this study were to investigate the prevalence and distribution of canine blastomycosis cases in Michigan and to identify risk factors for infection. Over 200 primary care veterinary clinics throughout the state were surveyed regarding blastomycosis prevalence, and demographic information was obtained from medical records of affected dogs that were evaluated at these clinics. A retrospective case control study was conducted for an additional 49 dogs with blastomycosis that were evaluated at specialty referral centers located in the southern mid-Michigan region. Prevalence rates were calculated for each county, and cases were mapped using geocoding software. Univariable and multivariable analyses were used to identify risk factors for infection. Prevalence rates were ≥100 cases per 100,000 dogs in five counties. Most blastomycosis cases originated from the Upper Peninsula or from a high-density area in the northern Lower Peninsula. Multivariable regression analysis identified travel or residence north of the 45th parallel as a strong risk factor for infection (P < .001). Blastomycosis was uncommon in southern counties. These results refute previous speculations and should be of value to both human and animal health. Given that many heightened risk areas are popular tourist destinations, practitioners across the USA should be mindful of the spatial distribution of blastomycosis in Michigan.

Molecular characterization of Mycobacterium bovis infection in cattle and buffalo in Amazon Region, Brazil.

The aim of this study was to characterize Mycobacterium bovis from cattle and buffalo tissue samples, from two Brazilian states, and to analyse their genetic diversity by spoligotyping. Tissue samples from tuberculosis suspect animals, 57 in Amazonas State (12 cattle and 45 buffaloes) and six from Pará State (5 cattle and one buffalo) from slaughterhouses under State Veterinary Inspection, were isolated in culture medium Stonebrink. The positive cultures were confirmed by PCR and analysed by the spoligotyping technique and the patterns (spoligotypes) were identified and compared at the Mycobacterium bovis Spoligotype Database (http://www.mbovis.org/). There was bacterial growth in 44 (69.8%) of the tissues of the 63 animals, of which PCR for region of differentiation 4 identified 35/44 (79.5%) as Mycobacterium bovis. Six different spoligotypes were identified among the 35 Mycobacterium bovis isolates, of which SB0295, SB1869, SB0121 and SB1800 had already been described in Brazil, and SB0822 and SB1608 had not been described. The most frequent spoligotype in this study (SB0822) had already been described in buffaloes in Colombia, a neighbouring country of Amazonas state. The other identified spoligotypes were also described in other South American countries, such as Argentina and Venezuela, and described in the Brazilian states of Rio Grande do Sul, Santa Catarina, São Paulo, Minas Gerais, Mato Grosso do Sul, Mato Grosso and Goiás, indicating an active movement of Mycobacterium bovis strains within Brazil.

Retrospective analysis of diagnoses and outcomes of 45 cats with micturition disorders presenting as urinary incontinence.

BACKGROUND: In contrast to dogs, the causes and outcomes of urinary incontinence (UI) in cats are largely unknown. OBJECTIVES: To determine the causes, identify comorbid conditions, and assess outcomes of cats with micturition disorders presenting as UI. ANIMALS: Forty-five cats with UI. METHODS: Retrospective study. Medical records of cats presented from January 2006 to December 2017 were searched using 45 keywords related to UI. History, presenting complaint, and physical examination findings were used to confirm a diagnosis. Cases were categorized based on functional and anatomic localizations. RESULTS: Forty-five cats met inclusion criteria. Spinal cord disease was the most common cause of UI (n = 18), followed by urethral (n = 17), bladder (n = 9), and ureteral (n = 1) disorders. Proportions of voiding and storage phase disorders were similar (53% and 47%, respectively). However, voiding-phase disorders were observed more frequently in males and younger-aged cats (P < .03). Urinary tract infection was detected in 11 of 28 (39%) cats. Outcomes were available in 38/45 cases; 16 cats (42%) regained continence, 3 (8%) improved with treatment, and 19 (50%) remained incontinent or were euthanized. Multiple variable logistic regressions indicated that spinal cord disease was significantly more likely to be associated with poor outcomes compared to bladder or urethral disorders (P < .04). CONCLUSIONS AND CLINICAL IMPORTANCE: Urinary incontinence in cats was associated with a variety of congenital and acquired disorders that affected both phases of micturition with similar frequency. Incontinent cats with spinal cord disorders were common and warrant a more guarded prognosis than do cats with bladder or urethral disorders.

Matrix Assisted Laser Desorption Ionization-Time-of-Flight mass spectrometry identification of Mycobacterium bovis in Bovinae.

In this study, Matrix Assisted Laser Desorption Ionization-Time-of-Flight (MALDI-TOF) mass spectrometry was used to identify Mycobacterium bovis from cattle and buffalo tissue isolates from the North and South regions of Brazil, grown in solid medium and previously identified by Polymerase Chain Reaction (PCR) based on Region of Difference 4 (RD4), sequencing and spoligotyping. For this purpose, the protein extraction protocol and the mass spectra reference database were optimized for the identification of 80 clinical isolates of mycobacteria. As a result of this optimization, it was possible to identify and differentiate M. bovis from other members of the Mycobacterium tuberculosis complex with 100% specificity, 90.91% sensitivity and 91.25% reliability. MALDI-TOF MS methodology described herein provides successful identification of M. bovis within bovine/bubaline clinical samples, demonstrating its usefulness for bovine tuberculosis diagnosis in the future.

Seroprevalence of Q fever in cattle, sheep and goats in the Volta region of Ghana.

Q fever is a zoonotic disease caused by Coxiella burnetii, a causative agent of abortion in livestock and febrile illness in humans. Outbreaks of human cases of Q fever have been reported in Australia and the Netherlands, which was linked to abortions in goat and sheep farms. In Ghana, information on Q fever in both livestock and humans is scanty. This study sought to determine the seroprevalence of Q fever in livestock in the Tongu area of the Volta region of Ghana. It was a cross sectional study with blood sampled from 204 cattle, 158 sheep and 100 goats. An indirect ELISA test was performed to detect Q fever antibodies in the serum of livestock. A total of 20 farms were sampled across the municipalities and an overall prevalence of Q fever was 21.6%. Specie-specific prevalence was 28.4% (45/158) for sheep, 21.7% (45/204) for cattle and 10% (10/100) for goats. Abortions were reported on all the farms sampled and most farmers lived in close proximity to the farms sampled. Q fever is prevalent in the North Tongu area and requires the attention of the veterinary and health authorities, using the One- Health approach in order to control its occurrence and save lives.

Epidemiological Study of Mycobacterium bovis Infection in Buffalo and Cattle in Amazonas, Brazil.

Bovine Tuberculosis (BTB) is an endemic disease in about one hundred countries, affecting the economy causing a decrease in productivity, condemnation of meat, and damaging the credibility on international trade. Additionally, Mycobacterium bovis the major causative agent for BTB can also infect humans causing a variety of clinical presentations. The aim of this study was to determine BTB prevalence and the main risk factors for the Mycobacterium bovis prevalence in cattle and buffalos in Amazonas State, Brazil. Tissue samples from 151 animals (45 buffalo and 106 cattle from five herds with buffalo only, 22 herds with cattle only, and 12 herds with buffalo and cattle) were obtained from slaughterhouses under State Veterinary Inspection. M. bovis were isolated on Stonebrink medium. The positive cultures were confirmed by polymerase chain reaction (PCR) testing. The apparent herd and animal prevalence rates were 56.4 and 5.40%, respectively. Regarding animal species, the apparent prevalence rates were 3% in cattle and 11.8% in buffalo. Generalized Linear Mixed Models (GLMM) with random effect were used to assess the association with risk factors on the prevalence. Species (buffalo), herds size (>100 animals) and the presence of both species (buffalo and cattle) in the herd were the major risk factors for the infection by Mycobacterium bovis in the region. The findings reveal an urgent need for evidence-based effective intervention to reduce BTB prevalence in cattle and buffalo and prevent its spread to the human population. Studies are needed to understand why buffalo are more likely to be infected by M. bovis than cattle in Amazon. Recommendations for zoning, use of data from the inspection services to generate information regarding BTB focus, adoption of epidemiological tools, and discouragement of practices that promote the mixing of cattle and buffalo, were made.

Viral diversity and abundance in polluted waters in Kampala, Uganda.

Waterborne viruses are a significant cause of human disease, especially in developing countries such as Uganda. A total of 15 virus-selective samples were collected at five sites (Bugolobi Wastewater Treatment Plant (WWTP) influent and effluent, Nakivubo Channel upstream and downstream of the WWTP, and Nakivubo Swamp) in July and August 2016. Quantitative PCR and quantitative RT-PCR was performed to determine the concentrations of four human viruses (adenovirus, enterovirus, rotavirus, and hepatitis A virus) in the samples. Adenovirus (1.53*105-1.98*107 copies/L) and enterovirus (3.17*105-8.13*107 copies/L) were found to have the highest concentrations in the samples compared to rotavirus (5.79*101-3.77*103 copies/L) and hepatitis A virus (9.93*102-1.11*104 copies/L). In addition, next-generation sequencing and metagenomic analyses were performed to assess viral diversity, and several human and vertebrate viruses were detected, including Herpesvirales, Iridoviridae, Poxviridae, Circoviridae, Parvoviridae, Bunyaviridae and others. Effluent from the wastewater treatment plant appears to impact surface water, as samples taken from surface water downstream of the treatment plant had higher viral concentrations than samples taken upstream. Temporal fluctuations in viral abundance and diversity were also observed. Continuous monitoring of wastewater may contribute to assessing viral disease patterns at a population level and provide early warning of potential outbreaks using wastewater-based epidemiology methods.

Investigation of blood lead concentrations in dogs living in Flint, Michigan.

OBJECTIVE To measure blood lead concentrations (BLCs) in dogs living in Flint, Mich, following a declared water crisis and to assess potential associations of BLCs with demographic data, water sources, and clinical signs in these dogs. DESIGN Cross-sectional study. ANIMALS 284 dogs residing in Flint, Mich (test population), and 47 dogs residing in East Lansing, Mich (control population), and immediately adjacent areas. PROCEDURES Blood samples were collected at free screening clinics in Flint (test population) and at the Michigan State University College of Veterinary Medicine Veterinary Medical Center (control population). Owners of test population dogs completed questionnaires providing demographic and clinical information. Hematologic evaluations were performed; BLCs were measured by inductively coupled plasma-mass spectrometry. RESULTS 4 of 284 test population dogs had BLCs > 50 ppb; an additional 20 had BLCs > 20 ppb. Overall, BLCs of test population dogs were higher than those of control dogs. Within the test population, young dogs (≤ 2 years of age) had higher BLCs than old dogs (≥ 6 years of age). Only 7.2% of test population dogs were drinking unfiltered tap water at the time of screening; however, dogs that had been receiving filtered or bottled water for ≤ 3 months before screening had higher BLCs than did those that received such water for > 3 months. CONCLUSIONS AND CLINICAL RELEVANCE Taken together, findings suggested that the impact of the Flint water crisis extended to companion animals. Results highlighted the importance of maintaining awareness of lead exposure and considering both human and animal well-being in cases of environmental toxicant exposures.

Use of whole-genome sequencing and evaluation of the apparent sensitivity and specificity of antemortem tuberculosis tests in the investigation of an unusual outbreak of Mycobacterium bovis infection in a Michigan dairy herd.

OBJECTIVE To describe use of whole-genome sequencing (WGS) and evaluate the apparent sensitivity and specificity of antemortem tuberculosis tests during investigation of an unusual outbreak of Mycobacterium bovis infection in a Michigan dairy herd. DESIGN Bovine tuberculosis (bTB) outbreak investigation. ANIMALS Cattle, cats, dog, and wildlife. PROCEDURES All cattle in the index dairy herd were screened for bTB with the caudal fold test (CFT), and cattle ≥ 6 months old were also screened with a γ-interferon (γIFN) assay. The index herd was depopulated along with all barn cats and a dog that were fed unpasteurized milk from the herd. Select isolates from M bovis-infected animals from the index herd and other bTB-affected herds underwent WGS. Wildlife around all affected premises was examined for bTB. RESULTS No evidence of bTB was found in any wildlife examined. Within the index herd, 53 of 451 (11.8%) cattle and 12 of 21 (57%) cats were confirmed to be infected with M bovis. Prevalence of M bovis-infected cattle was greatest among 4- to 7-month-old calves (16/49 [33%]) followed by adult cows (36/203 [18%]). The apparent sensitivity and specificity were 86.8% and 92.7% for the CFT and 80.4% and 96.5% for the γIFN assay when results for those tests were interpreted separately and 96.1% and 91.7% when results were interpreted in parallel. Results of WGS revealed that M bovis-infected barn cats and cattle from the index herd and 6 beef operations were infected with the same strain of M bovis. Of the 6 bTB-affected beef operations identified during the investigation, 3 were linked to the index herd only by WGS results; there was no record of movement of livestock or waste milk from the index herd to those operations. CONCLUSIONS AND CLINICAL RELEVANCE Whole-genome sequencing enhanced the epidemiological investigation and should be used in all disease investigations. Performing the CFT and γIFN assay in parallel improved the antemortem ability to detect M bovis-infected animals. Contact with M bovis-infected cattle and contaminated milk were major risk factors for transmission of bTB within and between herds of this outbreak.

Milk Hygiene in Rural Southwestern Uganda: Prevalence of Mastitis and Antimicrobial Resistance Profiles of Bacterial Contaminants of Milk and Milk Products.

Mastitis and antimicrobial resistance are a big challenge to the dairy industry in sub-Saharan Africa. A study was conducted in Kashongi and Keshunga subcounties of Kiruhura District (in Uganda) where the government and private sector have deliberate programs to improve production efficiency, quality, and safety of milk and its products. The study aimed to determine the prevalence of mastitis, its common causative agents, antimicrobial sensitivity of mastitis causing organisms, and contaminants of processed milk products: yoghurt and ghee. Seventy-one milk, fourteen yoghurt, and three ghee samples were collected from nine farms. Of the 71 cows tested, 54 (76.1%) had mastitis. The mastitis cases from Keshunga were 32 (59.3%) and Kashongi contributed 22 (40.7%) of the cases. The common mastitis causative agents were Staphylococcus spp. (30.8%), Streptococcus spp. (12.3%), Corynebacterium spp.(15.4%), and E. coli (7.7%). Some of the isolates were resistant to tetracycline and penicillin. Prevalent contaminants of yoghurt were Staphylococcus spp. (8.3%), Streptococcus spp. (8.3%), Corynebacterium spp. (8.3%), and E. coli (8.3%), whereas all ghee contained Streptococcus spp. (100%). Prevalence of mastitis, antimicrobial resistance, and contamination of milk products are high in the study area. Targeted programs to prevent and control mastitis as well as antibiotic resistance are recommended.

Survivability of Mycobacterium bovis on salt and salt-mineral blocks fed to cattle.

OBJECTIVE To determine the survivability of Mycobacterium bovis on salt and salt-mineral blocks in typical weather conditions in Michigan over two 12-day periods at the height of summer and winter. SAMPLE 4 salt (NaCl) and 4 salt-mineral blocks inoculated with pure cultures of a strain of M bovis currently circulating in Michigan livestock and wildlife. PROCEDURES In the summer and again in the winter, inoculated blocks were placed in secured outdoor facilities where equal numbers of each block type (2/type/season) were exposed to shade or sunlight. Samples were collected from randomly selected areas on the surface of each block beginning within 1 hour after placement (day 0) twice a day for the first 4 days and once a day from days 7 through 11. Bacterial culture of samples was performed to detect viable M bovis. RESULTS Depending on the exposure conditions, salt blocks yielded viable M bovis for up to 2 days after inoculation and salt-mineral blocks yielded viable M bovis for > 3 days. Survival time was greatest on salt-mineral blocks kept outdoors in the shade during the winter. The odds of recovering viable M bovis from salt-mineral block samples were 4.9 times as great during the winter (vs the summer) and 3.0 times as great with exposure to shade (vs sunlight). CONCLUSIONS AND CLINICAL RELEVANCE Results from this study indicated that salt and salt-mineral blocks should be considered potential sources of bovine tuberculosis when designing risk mitigation programs for cattle herds in areas with wildlife reservoirs of M bovis.

Herd outbreak of bovine tuberculosis illustrates that route of infection correlates with anatomic distribution of lesions in cattle and cats.

An outbreak of bovine tuberculosis (TB) in a Michigan dairy herd resulted in quarantine, depopulation, pathology, and epidemiologic investigations. This herd, compared to other TB-infected herds in Michigan, was unusual in the long-term feeding of waste milk to its replacement calves. The herd had 80 cattle with positive results on caudal fold test or gamma interferon testing, which were reclassified as suspects because the herd had never been known to be tuberculous previously. Autopsy revealed striking variation in the anatomic distribution of gross anatomic lesions, microscopic lesions, and culture-positive lymph nodes between the adult cattle, the calves, and the domestic cats present on the farm. Adult cattle had lesions and culture-positive lymph nodes predominantly within the thoracic lymph nodes, whereas cats had 50% of their lesions and culture-positive lymph nodes in their abdomens, and 50% of positive calves had culture-positive lymph nodes in their abdomens. This difference in anatomic distribution correlated with the likely routes of infection, which are believed to be by direct airborne transmission in adult cattle and indirect ingestion of contaminated milk in both calves and cats. Although TB literature over the past 100-plus years states that the route of infection may manifest itself in differences in lesion anatomic distribution, our team has been working with TB for over 20 years, and we have never encountered such striking variation between different groups of animals on the same farm.

Thirty-minute screening of antibiotic resistance genes in bacterial isolates with minimal sample preparation in static self-dispensing 64 and 384 assay cards.

In a clinical setting, molecular assays such as polymerase chain reaction offer a rapid means to infer or confirm identity and therapeutic decisions. Accordingly, a number of molecular assays targeting identity and antibiotic resistance (AR) genes have been developed; however, these methods can be technically complex and relatively expensive. Herein, we describe a diagnostic concept utilizing isothermal amplification technology with non-purified heat-lysed cells and self-dispensing cards for testing multiple primers in parallel. This proof-of-concept study, performed with Staphylococcus aureus isolates and associated AR genes, was compared with culture-based susceptibility and quantitative PCR (qPCR). Results demonstrate reduced sample processing steps resulting in a turnaround time (starting from bacterial culture to ending in the antibiotic resistance gene profile) in less than 30 min. For antibiotics tested in which an associated AR gene was targeted on the Gene-Z card, 69% (18/26) of culture-based resistance events were positive for related AR genes. A comparison of loop-mediated isothermal amplification (LAMP) and qPCR assays targeting the same antibiotic resistance genes showed a 98.2% agreement in terms of presence and absence calls. Identity-based discrepancies between conventional (phenotypic) and molecular (genotypic) results were further resolved, and we were able to demonstrate higher accuracy in identification with the molecular analysis.

Effect of feeding a direct-fed microbial on total and antimicrobial-resistant fecal coliform counts in preweaned dairy calves.

OBJECTIVE: To determine whether feeding a direct-fed microbial (DFM) to dairy calves would reduce total and antimicrobial-resistant coliform counts in feces and affect average daily gain (ADG). ANIMALS: 21 preweaned Holstein heifer calves. PROCEDURES: The study had a randomized complete block design. Within each block, 3 consecutively born calves were randomly assigned to 1 of 3 treatment groups within 24 hours after birth (day 0). Calves were fed the DFM at 1.0 g (DFM1; n = 7) or 0.5 g (DFM2; 7) twice daily or no DFM (control; 7) from days 0 through 29. A fecal sample was collected from each calf daily on days 0 through 3 and then every other day through day 29. Fecal samples were cultured, and mean numbers of total coliforms and coliforms resistant to ampicillin, ceftiofur, and tetracycline were compared among the 3 treatment groups. Calves were weighed on days 0 and 29 to calculate ADG. RESULTS: Mean total fecal coliform counts did not differ significantly among the 3 treatment groups. Mean ceftiofur-resistant and tetracycline-resistant coliform counts for the control group were significantly lower, compared with those for the DFM1 and DFM2 groups. Mean ADG did not differ significantly between the DFM1 and DFM2 groups; however, the mean ADG for all calves fed the DFM was 0.15 kg less than that for control calves. CONCLUSION AND CLINICAL RELEVANCE: Results suggested that the DFM fed to the preweaned calves of this study did not reduce total or antimicrobial-resistant coliform counts in feces.

Within-Farm Changes in Dairy Farm-Associated Salmonella Subtypes and Comparison to Human Clinical Isolates in Michigan, 2000-2001 and 2009.

Temporal changes in the distribution of Salmonella subtypes in livestock populations may have important impacts on human health. The first objective of this research was to determine the within-farm changes in the population of subtypes of Salmonella on Michigan dairy farms that were sampled longitudinally in 2000-2001 and again in 2009. The second objective was to determine the yearly frequency (2001 through 2012) of reported human illnesses in Michigan associated with the same subtypes. Comparable sampling techniques were used to collect fecal and environmental samples from the same 18 Michigan dairy farms in 2000-2001 and 2009. Serotypes, multilocus sequence types (STs), and pulsed-field gel electrophoresis (PFGE) banding patterns were identified for isolates from 6 farms where >1 Salmonella isolate was recovered in both 2000-2001 and 2009. The distribution of STs was significantly different between time frames (P < 0.05); only two of 31 PFGE patterns were identified in both time frames, and each was recovered from the same farm in each time frame. Previously reported within-farm decreases in the frequency of multidrug-resistant (MDR) Salmonella were due to recovery of MDR subtypes of S. enterica serotypes Senftenberg and Typhimurium in 2000-2001 and genetically distinct, pansusceptible subtypes of the same serotypes in 2009. The annual frequency of human illnesses between 2001 and 2012 with a PFGE pattern matching a bovine strain decreased for patterns recovered from dairy farms in 2000-2001 and increased for patterns recovered in 2009. These data suggest important changes in the population of Salmonella on dairy farms and in the frequency of human illnesses associated with cattle-derived subtypes.