Search

In this study, we investigated the variation in the microbial community present in bulk tank milk samples and the potential effect of different farm management factors. Bulk tank milk samples were collected repeatedly over one year from 42 farms located in northern Sweden. Total and thermoresistant bacteria counts and 16S rRNA gene-based amplicon sequencing were used to characterize microbial community composition. The microbial community was in general heterogeneous both within and between different farms and the community composition in the bulk tank milk was commonly dominated by Pseudomonas, Acinetobacter, Streptococcus, unclassified Peptostreptococcaceae, and Staphylococcus. Principal component analysis including farm factor variables and microbial taxa data revealed that the microbial community in milk was affected by type of milking system. Milk from farms using an automatic (robot) milking system (AMS) and loose housing showed different microbial community composition compared with milk from tiestall farms. A discriminant analysis model revealed that this difference was dependent on several microbial taxa. Among farms using an automatic milking system, there were further differences in the microbial community composition depending on the brand of the milking robot used. On tiestall farms, routines for teat preparation and cleaning of the milking equipment affected the microbial community composition in milk. Total bacteria count (TBC) in milk differed between the farm types, and TBC were higher on AMS than tiestall farms (log 4.05 vs. log 3.79 TBC/mL for AMS and tiestalls, respectively). Among tiestall farms, milk from farms using a chemical agent in connection to teat preparation and a more frequent use of acid to clean the milking equipment had lower TBC in milk, than milk from farms using water for teat preparation and a less frequent use of acid to clean the milking equipment (log 3.68 vs. 4.02 TBC/mL). There were no significant differences in the number of thermoresistant bacteria between farm types. The evaluated factors explained only a small proportion of total variation in the microbiota data, however, despite this, the study highlights the effect of routines associated with teat preparation and cleaning of the milking equipment on raw milk microbiota, irrespective of type of milking system used.

Bulk tank milk (BTM) is an easy, inexpensive, and representative sample for detection of Coxiella burnetii infections (Q fever) in dairy herds using real-time PCR. Bulk tank milk PCR can be performed either for initial herd screening or for monitoring the effectiveness of preventive measures. However, one major limitation under field conditions is the need to deliver BTM samples in adequate condition (quickly, safely, and under refrigeration) to a qualified laboratory. In addition, sending non-inactivated biological samples via normal mail may be prohibited. We developed an innovative, easy, and accurate diagnostic tool (QTest) for Q fever to support veterinarians and farmers in overcoming these constraints. The farmer or veterinarian simply places some drops of BTM on a Whatman FTA Elute Micro Card (FTA card) and lets the card dry before mailing it to the laboratory. In a 2-step study, we tested the hypotheses that (1) BTM samples stored on FTA cards are stable over time and at different temperatures, and (2) PCR results obtained via FTA cards are consistent with those obtained from raw BTM samples. The stability of C. burnetii DNA in milk preserved on an FTA card was maintained for at least 29 d at room temperature or 37°C to mimic field conditions. In our field study, of the original 70 positive BTM samples (when tested on raw BTM just after sampling), 58 samples were positive (on either raw BTM or FTA card) by the time of the direct comparison study (10 to 14 d later). Of these 58 samples, 45 raw BTM samples still tested positive after aging, and 53 FTA card BTM samples tested positive, indicating that detection was higher using FTA cards (91.4%) than raw milk (77.6%). Therefore, with inactivation and shipping advantages, this technique facilitates an easier and more practical approach to diagnosis of Q fever at the herd level and would support Q fever control strategies, especially in countries lacking adequate and close laboratory facilities.

Bulk tank milk (BTM) is regularly used for surveillance on dairy farms for disease conditions such as mastitis and Johne's disease. In this study, we used 16S rRNA sequencing and bait-capture enrichment to characterize the microbiota and resistome of BTM, and investigate potential differences between the cream or pellet fractions. A total of 12 BTM samples were taken from 12 Prince Edward Island dairy farms, in Atlantic Canada, in duplicates. The DNA was analyzed by high-throughput sequencing of the 16S rRNA gene and a suite of antimicrobial resistance (AMR) genes. Target-capture enrichment of AMR genes was conducted before shotgun sequencing. The bioinformatics pipelines QIIME 2 and AMR++ were used for microbiota and resistome analysis, respectively. Differences between microbiotae were evaluated qualitatively with nonmetric multidimensional scaling and quantitatively with permutational ANOVA of UniFrac distances. A total of 47 phyla were present across the BTM samples. Firmicutes, Bacteroidetes, Proteobacteria, and Actinobacteria were the most abundant phyla. At the genus level, Corynebacterium, Acinetobacter, Lactobacillus, and Turicibacter were the most abundant. There was no significant difference in the Faith's phylogenetic diversity between the cream and pellet fraction. Faith's phylogenetic diversity differed marginally by stall type. There were 10,217 hits across 80 unique AMR genes, with tetracycline resistance being the most common class.

To reduce antimicrobial use, infusion of antimicrobials into only infected cows at the end of lactation (selective dry cow therapy) is preferable to infusion of every cow with antimicrobials. Use of selective dry cow antimicrobial therapy requires differentiation of probably infected from uninfected cows to enable treatment allocation. Milk somatic cell count (SCC) has been used to distinguish between cows with and without intramammary infection (IMI). However, SCC may be influenced by milk yield, stage of lactation, breed, and herd-level variables such as prevalence of infection. Cut points for SCC, to distinguish between cows with and without an IMI, may need to differ between cow age groups and breeds, or among herds. This study evaluated associations between SCC and major pathogen IMI in one or more quarters of 2,606 cows from 36 herds in 4 regions of New Zealand. In the last week of lactation, cows selected at random had milk samples collected from each quarter, and the teat-end condition and hygiene of the udder were scored. Herd- and cow-level data including age, breed, milk volume, and SCC at each production were recorded, and bulk tank milk SCC and volume of milk shipped were collated. At cow level, the association between average, maximum, and last cow-composite SCC, and presence of a major pathogen IMI in one or more quarters of cows, was examined using receiver operator curves. Predictive logistic regression models were then developed that included potential effect modifiers such as age, milk yield, and bulk tank milk SCC. The population average prevalence of major pathogen IMI was 7.2% of cows (95% confidence interval = 5.9-8.6), and this varied significantly between herds. The average, maximum, and last cow-composite SCC of lactation were all predictive of presence of a major pathogen IMI and did not differ in their ability to discriminate infected from uninfected cows. However, the optimal cut points for the last SCC, the maximum SCC, and average SCC were 108, 152, and 105 × 1,000 cells/mL, respectively. Inclusion of age, bulk tank SCC, and history of clinical mastitis improved overall model fit. However, inclusion of these variables did not improve the discriminatory power of maximum cow-composite SCC used alone. We conclude that cow-composite SCC on its own resulted in sensitivities and specificities of between 0.76 and 0.86, and 0.71 to 0.80, respectively, for determination of presence of major pattern IMI, and the predictive value was not improved by addition of other predictor variables.

Background Colonization of dairy cows by Staphylococcus aureus (S. aureus), especially those which are multi-drug resistant and toxin producing, is a concern for animal health and well-being as well as public health. The objective of this study was to investigate the prevalence, antibiotic resistance, gene content and virulence determinants of S. aureus in bulk tank milk samples (BTM) from U.S. dairy herds. Results BTM samples were collected, once in winter and once in summer, from 189 U.S. dairy herds. Of 365 BTM samples cultured, the sample and herd prevalence of S. aureus in BTM was 46.6% (170 of 365 samples) and 62.4% (118 of 189 herds), respectively. Among a subset of 138 S. aureus isolates that were stored for further analysis, 124 were genome sequenced after being confirmed as S. aureus using phenotypic tests. The most commonly identified antimicrobial resistance-associated gene was norA (99.2%) and mecA gene responsible for methicillin resistance (MRSA) was identified in one isolate (0.8%). The most frequently detected putative virulence genes were aur (100%), hlgB (100%), hlgA, hlgC, hlb (99.2%), lukE (95.9%) and lukD (94.3%). In the 53 staphylococcal enterotoxin positive isolates, sen (37.9%), sem (35.5%), sei (35.5%) and seg (33.1%) were the most frequently detected enterotoxin genes. Among the 14 sequence types (ST) and 18 spa types identified, the most common was ST2187 (20.9%) and t529 (28.2%), respectively. The most predominant clone was CC97 (47.6%) followed by CC unknown (36.3%). The single MRSA isolate belonged to ST72-CC8, spa type t126 and was negative for the tst gene but harbored all the other virulence genes investigated. Conclusion Our findings indicated a high prevalence of S. aureus in BTM of U.S. dairy herds, with isolates showing little evidence of resistance to antibiotics commonly used to treat mastitis. However, isolates often carried genes for the various enterotoxins. This study identified predominant genetic clones. Despite lower prevalence, the presence of MRSA and multi-drug resistant strains in BTM poses a significant risk to animal and public health if their number were to increase in dairy environment. Therefore, it is necessary to continuously monitor the use of antibiotics in dairy cows.

This study aimed to determine the occurrence of methicillin-resistant (MR) non-aureus staphylococci (NAS) on 20 preselected German dairy farms. Farms were selected based on the detection of methicillin-resistant Staphylococcus aureus (MRSA) during previous diagnostic investigations. Bacterial culture of presumptive MR-NAS was based on a 2-step enrichment method that has been recommended for MRSA detection. Quarter milk samples (QMS), bulk tank milk, swab samples from young stock, and environmental samples were collected for bacterial culture. Methicillin-resistant NAS were detected on all study farms. The MR-NAS positive test rate was 3.3% (77/2,347) in QMS, 42.1% (8/19) in bulk tank milk, 29.1% (59/203) in nasal swabs from milk-fed calves, 18.3% (35/191) in postweaning calves, and 7.3% (14/191) in nasal swabs from prefresh heifers. In the environment, MR-NAS were detected in dust samples on 25% (5/20) of the dairy farms as well as in teat liners and suckers from automatic calf feeders. The geometric mean somatic cell count in QMS affected by MR-NAS (183,000 cells/mL) was slightly higher compared with all QMS (114,000 cells/mL). Nine MR-NAS species were identified; Staph. sciuri, Staph. lentus, Staph. fleurettii, Staph. epidermidis, and Staph. haemolyticus were the most common species. In addition, 170 NAS isolates were identified that showed reduced cefoxitin susceptibility (4 mg/L) but did not harbor the mecA or mecC genes. On some farms, similar mobile genetic elements were detected in MR-NAS and MRSA. It was suggested that resistance genes may be transferred between NAS and Staph. aureus on the respective farms.

Staphylococcus aureus is one of the most frequent pathogens causing intramammary infections in dairy herds. Consequently, virulence factors, pathobiology, and epidemiology of Staphylococcus aureus strains have been widely assessed through the years. Nevertheless, not much has been described about the epidemiology of Staph. aureus strains from bulk tank milk (BTM) and adherences on milking equipment (AMES), even when these strains may play a role in the quality of milk that is intended for human consumption. The objective of this study was to assess the strain diversity of 166 Staph. aureus isolates collected from 3 consecutive BTM samples, and from AMES in contact with milk from 23 Chilean dairy farms. Isolates were analyzed and typed using pulsed-field gel electrophoresis. Diversity of strains, both within and among farms, was assessed using Simpson's index of diversity (SID). On farms where Staph. aureus was isolated from both AMES and BTM (n = 8), pulsotypes were further analyzed to evaluate the role of AMES as a potential source of Staph. aureus strains in BTM. Among all Staph. aureus analyzed by pulsed-field gel electrophoresis, a total of 42 pulsotypes (19 main pulsotypes and 23 subtypes) were identified. Among dairy farms, strain diversity was highly heterogeneous (SID = 0.99). Within dairy farms, Staph. aureus strain diversity was variable (SID = 0 to 1), and 18 dairy operations (81.8%) had one pulsotype that was shared between at least 2 successive BTM samples. In those farms where Staph. aureus was isolated in both AMES and BTM (n = 8), 7 (87.5%) showed a clonal distribution of Staph. aureus strains between these 2 types of samples. The overlapping of certain Staph. aureus strains among dairy farms may point out common sources of Staph. aureus among otherwise epidemiologically unrelated farms. Indistinguishable Staph. aureus strains between AMES and BTM across dairy farms suggest that Staph. aureus-containing AMES may represent a source for BTM contamination, thus affecting milk quality. Our study highlights the role of viable Staph. aureus in AMES as a source for BTM contamination on dairy farms, and also describes the overlapping and presence of specific BTM and AMES pulsotypes among farms.

This study was carried out on 200 random milk samples сolleсted from different areas at Dakahlia Governorate for detection of prevalence and antimicrobial resistance of some zoonotic bacteria in milk as Staphylococcus aureus and Streptococci by using biochemical tests, antibiotic sensitivity test, PCR for confirmation and detection of some resistance and virulence genes. Staphyloсoссus aureus (S. aureus) was detected in 63%, 76%, and 42%, while Streptococcus agalaсtiae (St. agalaсtiae) was found in 8%, 10% and 4% of examined individual milk samples, retail milk samples, and milk of Bulk tank, respectively. S. aureus isolates revealed high levels of resistance to ampicillin (100%), nalidxiс acid (90%), oxaсillin (85%), peniсillin (85%), cefoxitin (75%), rifampin (25%), tetraсyсline (20%) and erythromyсin and sulphamethoxazole/ trimethoprim (10%). Whereas, St. agalaсtiae show high resistance to tetracycline (90%), ampiсillin (80%), rifampin (60%), sulphamethoxazole/ trimethoprim (50%), gentamyсin (45%), erythromyсin and nalidxiс aсid (40%), cholorampheniсol (30%), and streptomyсin (25%). PСR results revealed that 4 out of 5 (80%) methicillin-resistant (MRSA) isolates had meсAand 2 (40%) had meсС,while 1 (20%) had Seaenterotoxin. Three isolates (100%) of St. agalaсtiae had sul1 gene and one out of three (33.3%) had tetK genes, while dfrA could not be detected. The main outcome of the current work is that milk can cause severe public health hazards to people because it had a variety of microorganisms. It is important to ensure using good hygienic practices in farms and prevent the haphazard abuse of antibiotics.

Nontuberculous mycobacteria (NTM) compose a group of mycobacteria that do not belong to the Mycobacterium tuberculosis complex group. They are frequently isolated from environmental samples such as water, soil, and, to a lesser extent, food samples. Isolates of NTM represent a major health threat to humans worldwide, especially those who have asthma or are immunocompromised. Human disease is acquired from environmental exposures and through consumption of NTM-contaminated food. The most common clinical manifestation of NTM disease in human is lung disease, but lymphatic, skin and soft tissue, and disseminated disease are also important. The main objective of the current study was to profile the farm-level contamination of cow milk with NTM by examining milk filters and bulk tank milk samples. Five different NTM species were isolated in one dairy herd in Wisconsin, with confirmed 16S rRNA genotypes including Mycobacterium fortuitum, Mycobacterium avium ssp. hominissuis, Mycobacterium abscessus, Mycobacterium simiae, and Mycobacterium avium ssp. paratuberculosis (Mycobacterium paratuberculosis). In tank milk samples, M. fortuitum was the predominant species in 48% of the samples, whereas M. chelonae/abscessus and M. fortuitum were the only 2 species obtained from 77 and 23% of the examined filters, respectively. Surprisingly, M. avium ssp. hominissuis, M. paratuberculosis, and M. simiae were isolated from 16.7, 10.4, and 4% of the examined milk samples, respectively, but not from milk filters. Interestingly, NTM isolates from human clinical cases in Wisconsin clustered very closely with those from milk samples. These findings suggest that the problem of NTM contamination is underestimated in dairy herds and could contribute to human infections with NTM. Overall, the study validates the use of bulk tank samples rather than milk filters to assess contamination of milk with NTM. Nontuberculous mycobacteria represent one type of pathogens that extensively contaminate raw milk at the farm level. The significance of our research is in evaluating the existence of NTM at the farm level and identifying a simple approach to examine the potential milk contamination with NTM members using tank milk or milk filters from dairy operations. In addition, we attempted to examine the potential link between NTM isolates found in the farm to those circulating in humans in Wisconsin.

Mycoplasmabovine mastitis is a highly contagious disease, usually associated with clinical cases refractory to antibiotic treatment. The aim of this study was the isolation ofMycoplasmaspecies in cattle milk samples from dairy herds in Chile. Bulk tank milk samples selected by convenience from 91 Holstein Friesian dairy herds located in Los Rios (66) and Los Lagos (25), the two most important dairy Regions in the country, were collected. Additionally, 100 individual milk samples from cows with a high incidence of clinical mastitis, refractory to antibiotic therapy, and negative bacteriological results for traditional mastitis pathogens, all from the Biobío Region and received in our diagnostic laboratory, were included. All samples were cultured for 10 days on PPLO medium. The differentiation of suspect colonies between genusMycoplasmaandAcholeplasmawas performed by the digitonin test and a specific PCR. The species identification was performed by aM. bovisspecific PCR and 16S rRNA sequencing.Mycoplasmawas isolated from 3 (3.3%) bulk tank milk samples and 2 (2%) individual cow milk samples. All colonies were identified asMycoplasmaby the digitonin test and by a specific PCR. At species level, one strain isolated from a bulk tank milk sample was identified asM. bovis.The remaining two strains isolated from bulk tank milk samples were identified asM. bovigenitalium,while the two strains isolated from milk of individual cows were identified asM. alkalescens.These results show that not onlyM. bovisis present in Chilean dairy herds, but also other pathogenic species not previously described in Chile such asM. bovigenitaliumandM. alkalescens,which pose a potential risk for dairy herds in southern Chile.

Cleanliness of milking equipment is known to be important for the safety of dairy products and to prevent the spread of diseases among cows. We investigated the cleaning procedures of milking equipment and suckling equipment on Japanese dairy farms, and the cleanliness of bucket milkers, suckling buckets, milk receivers, and bulk tanks, using adenosine triphosphate (ATP) bioluminescence test. Bulk tanks (except one bulk tank) and milk receivers were washed by automated cleaning, but all bucket milkers and suckling buckets were washed by manual cleaning. Detergents were often not used to clean bucket milkers and suckling buckets. The log10 transformed relative luminescence units (LRLU) of equipment washed by manual cleaning was higher than equipment washed by automated cleaning. Clean surfaces (≤2.2 LRLU) were only observed on the bulk tank and the milk receiver. More than 50% of the LRLU of the mouthpiece, the rubber packing of claw, and the nipple of the suckling bucket were determined dirty. These results suggest that the cleanliness of the bucket milkers and the suckling buckets washed by manual cleaning was lower than that of the equipment washed by automated cleaning, and may be due to insufficient cleaning procedures.

Escherichia coli (E. coli) can easily contaminate the bovine mammary gland; hence, monitoring of bulk tank milk (BTM) samples helps detect mastitis. In this study, we investigated the characteristics of E. coli from BTM of a dairy company. A total of 58 BTM batches were collected from 29 dairy farms of a dairy company, and 40 E. coli were tested in this study. E. coli showed the highest resistance to cephalothin (52.4%), followed by colistin (28.6%), ampicillin (19.0%) and tetracycline (19.0%). However, E. coli showed low resistance from 0% to 9.5% against other 14 antimicrobials. In the distribution of antimicrobial resistant gene, β-lactamases genes blaOXA and tetracyclines-resistant gene tetB were detected in 15 (37.5%) and 2 (5.0) isolates, respectively, and blaTEM, aac (6')-Ib, aac (3)-II, tetA, sul1 and sul2 genes were only detected once in each (2.5%) isolate. The O127 (22.5%) serogroup was the most predominant, but all E. coli isolates were divided into 20 serogroups. The occurrence of antimicrobial-resistant bacterial infection can be problematic as it not only deteriorates the milk quality but also limits the therapeutic choices of antimicrobials in humans. Therefore, continuous monitoring is recommended to track antimicrobial-resistant E. coli in BTM and raw milk.

These findings showed that raw milk from dairy cattle farms and collector’s bulk tank in Sebeta town was contaminated with public health important bacterial species like E. coli and Salmonella species and the observed resistant to certain antimicrobial drugs also needs attention. To ensure the quality of raw milk, stakeholders engaged in milk and dairy production chain should be trained on hygienic practices.

Objective: This study was conducted to screen the antibacterial activity of Acacia nilotica plant extract and four disinfectants(TH4, Tek-trol, Virkon S and peracetic acid)against pathogens causing mastitis (E.coli, Staphylococcus aureus and Streptococcus agalactiae). Design: Descriptive study Animals: Three hundred eighty-two dairy cows Procedures: Three thousands and seventy seven samples were collected from three dairy farms at Dakahlia province. Samples include animal samples (1528 of both quarter milk and teat skin swabs) and 221 environmental samples include 60 bedding, 60 Milk linear, 36 feed, 36 water, 11 bulk tank milk and 18 workers' hand swabs. All samples were examined bacteriologically for isolation and identification of mastitis causing pathogens (S. aureus, E. coli and St. agalactiae). Furtherly, minimum inhibitory concentration (MIC) and minimum bacterial concentration (MBC) tests were used to investigate the antimicrobial activities of selected disinfectants and Acacia nilotica extract toward isolated strains. Results: TH4 and aqueous Acacia nilotica extract were effective against S. aureus and St. agalactiae, with MIC values as low as 3.13 μg/ml of the original concentrations. Meanwhile, the antimicrobial actions of Tek-trol and Acacia nilotica extract toward E. coli isolates only reached 12.5 μg/ml. The in-vitro bactericidal effect showed that, MBC values of Acacia nilotica plant extract achieved the highest inhibitory concentration up to 25 μg/ml among the tested disinfectants. Conclusion and clinical relevance: In conclusion, Acacia nilotica plant extract have antibacterial activity comparable to commercialdisinfectants for the control of mastitogenic pathogens in dairy farms, providing a promising tool for mastitis control in dairy farms.

In relation to milk hygiene and safety, the milking phase is considered a critical moment since it represents a probable pathway for the introduction of unwanted microorganisms in the dairy chain. In particular, Listeria monocytogenes (L. monocytogenes) and Escherichia coli ( E. coli ) are known as possible microbial contaminants of raw sheep milk although extensive knowledge regarding their contamination dynamics in sheep farms is still lacking. This study aimed to examine the occurrence and concentration of these microorganisms in milk samples collected from farm bulk tanks in the region of Lazio (Central Italy) and investigate the related risk factors. Over a period of one year, we collected 372 milk samples from 87 sheep farms and administered a questionnaire in order to acquire information regarding relevant farm management variables. The results showed that L. monocytogenes was not present in any of the samples, suggesting that the contamination of sheep bulk tank milk with this pathogen has a low probability. Meanwhile, E. coli was found in almost two thirds of milk samples (61%) but was quantified below 10 2 CFU/ml in most of them (around 75%). Statistical analysis indicated that the presence of E. coli tends to be more probable and counts are significantly higher during the warmest seasons. Unexpectedly, milk collected by hand milking resulted in lower level of contamination. We believe that, although further studies are necessary to clarify some aspects, the reported data contributed to deepen the knowledge about the occurrence of L. monocytogenes and E. coli in raw sheep milk and provided useful data for future risk assessments.

Simple Summary Staphylococci are present in the microbiota of both humans and animal species, being recognized as the most important opportunistic pathogens. Antimicrobial resistance (AMR) has become a global public health issue presenting a significant risk because it severely limits treatment options. Methicillin resistance in staphylococci (MRS) poses a specific problem as it may cause serious human and animal infections, eventually resulting in death. The increasing observation of MRS in different animal species has raised the concern of their impact on animal health and the potential of zoonotic transmission. The availability of comprehensive data on the ecology and distribution of MRS in animals and food products worldwide is necessary to understand their relevance in the "One Health" domain. However, there is a gap in information in terms of MRS and the Arab countries. Therefore, our study aimed to provide an overview of the situation of MRS in these countries by reviewing the available data on livestock and animal products and making recommendations for the future. The prevalence of methicillin resistance in staphylococci has been increasing globally and is currently one of the major public health concerns. In particular, treating infections caused by staphylococci with acquired antimicrobial resistance is problematic, as their treatment is more difficult. The resistance is found both in human and animal staphylococcal strains. Methicillin-resistant staphylococci (MRS) have also been increasingly reported in wildlife. In Arab countries, MRS has been detected in food producing animals and food products; however, the risk this poses is somewhat unclear, and still a significant lack of information on the trend and distribution of these pathogens in these countries, which have a specific ecosystem (desert) and traditions (Muslim culture). In this manuscript, we aim to provide an overview of the prevalence and the major MRS clonal lineages circulating in these specific countries and compare to them other situations with different ecosystems and cultures.

Q fever is one of the commonest infectious diseases worldwide. A Coxiella burnetii prevalence of 97.6% has been found by ELISA and PCR tests of the bulk tank milk in dairy cattle farms of Hungary. The herd- and individual-level seroprevalence rates of C. burnetii in the examined dairy cows and farms have dramatically increased over the past ten years. Three high-producing industrial dairy farms were studied which had previously been found ELISA and PCR positive for C. burnetii by bulk tank milk testing. Coxiella burnetii was detected in 52% of the 321 cows tested by ELISA. Pregnancy loss was detected in 18% of the cows between days 29–35 and days 60–70 of gestation. The study found a higher seropositivity rate (80.5%) in the cows that had lost their pregnancy and a seropositivity of 94.4% in the first-bred cows that had lost their pregnancy at an early stage. The ELISA-positive pregnant and aborted cows were further investigated by the complement fixation test (CFT). In dairy herds an average of 66.6% individual seropositivity was detected by the CFT (Phase II) in previously ELISA-positive animals that had lost their pregnancy and 64.5% in the pregnant animals. A higher (Phase I) seropositivity rate (50.0%) was found in the cows with pregnancy loss than in the pregnant animals (38.5%). The high prevalence of C. burnetii in dairy farms is a major risk factor related to pregnancy loss.

Objectives To investigate the prevalence and characteristics of methicillin-resistant staphylococci on dairy farms in England and Wales including zoonotic MRSA. Methods Bulk tank milk was sampled from 363 dairy farms in 2015-2016 and methicillin-resistant staphylococci were isolated by salt broth enrichment and plating on MRSA Brilliance selective agar. Isolates were characterised through antimicrobial susceptibility testing and whole-genome sequencing. Results Methicillin-resistant staphylococci were isolated from ∼5% of dairy farms and belonged to six different species, Staphylococcus aureus, Staphylococcus epidermidis, Staphylococcus lentus, Staphylococcus saprophyticus, Staphylococcus fleurettii and Staphylococcus sciuri. Whole-genome sequencing revealed a large variety of antimicrobial resistance genes and SCCmec elements were present, including mecA and mecC alleles. Potentially zoonotic methicillin-resistance S. aureus were found at a low prevalence (0.83% of sampled dairy farms). Whole-genome sequencing also provided evidence for the mobility of a primordial mec gene complex, independently of a SCCmec element, which appears to have been acquired by S. saprophyticus from S. fleurettii. Conclusions These data give new insight into the epidemiology of veterinary methicillin-resistant staphylococci to inform future surveillance and zoonotic risk evaluation. Our data indicate that MRSA has likely decreased in prevalence since earlier survey work in England and Wales during 2011-12 and highlights the diversity of methicillin resistance and other resistance determinants among bovine-associated staphylococci with implications for veterinary and human medicine.

This Research Reflection short review presents an overview of the effects of heat stress on dairy cattle udder health and discusses existing heat stress mitigation strategies for a better understanding and identification of appropriate abatement plans for future stress management. Due to high ambient temperatures with high relative humidity in summer, dairy cows respond by changes of physical, biochemical and biological pathways to neutralize heat stress resulting in decreased production performance and poorer immunity resulting in an increased incidence of intramammary infections (IMI) and a higher somatic cell count (SCC). In vitro studies on bovine polymorphonuclear cells (PMN) suggested that heat stress reduces the phagocytosis capacity and oxidative burst of PMN and alters the expression of apoptotic genes and miRNA which, together with having a negative effect on the immune system, may explain the increased susceptibility to IMI. Although there are limited data regarding the incidence rate of clinical mastitis in many countries or regions, knowledge of SCC at the cow or bulk tank level helps encourage farmers to improve herd health and to develop strategies for infection prevention and cure. Therefore, more research into bulk tank SCC and clinical mastitis rates is needed to explain the effect of heat stress on dairy cow udder health and functions that could be influenced by abatement plans.

The aim of our research was to determine the influences of some environmental factors on somatic cell count (SCC) and chemical composition of cow bulk tank milk. A total of 428 bulk tank milk samples of Brown Swiss cows were collected between January and October 2006 from family dairy farms located in Kırşehir province of Turkey. To evaluate the effect of farm scale, daily total bulk tank milk production was classified in three groups (small: 40 kg/d). Also, three groups were designed according to average SCC values as low (500x103 cells/ml). SCC was transformed according to the base of log10, because it did not show normal distribution.The results showed that sampling season was significantly affected fat content (P500x103 cell/ml). As a result, It was concluded that season was primarily responsible among the most important environmental factor to increase bulk tank milk quality., The aim of our research was to determine the influences of some environmental factors on somatic cell count (SCC) and chemical composition of cow bulk tank milk. A total of 428 bulk tank milk samples of Brown Swiss cows were collected between January and October 2016 from family dairy farms located in Kırşehir province of Turkey. To evaluate the effect of farm scale, daily total bulk tank milk production was classified in three groups (small: 40 lt/d). Also, three groups were designed according to average SCC values as low (500x103 cells/ml). SCC was transformed according to the base of log10, because it did not show normal distribution. The results showed that sampling season was significantly affected fat content (P500x103 cell/ml). As a result, it was concluded that season was primarily responsible among the most important environmental factor to increase bulk tank milk quality.

Objective: A cross sectional study was conducted between June 2017 and August 2018 in around Dakahlia governorate to identify Methicillin resistant Staphylococcus aureus (MRSA) and to determine risk factors associated with the occurrence of mastitis in dairy cows. Design: Cross sectional study Animals and samples: 130 dairy cattle (744 samples) and 181 environmental samples (36 bedding, 37 bulk tank milk BTM, 27 feed troughs, 27 water troughs, 27 milk linear and 27 worker’s nasal swabs), with a total of 925 samples were studied. Procedures: Three hundred and seventy-two quarters were examined to detect clinical and subclinical mastitis by physical examinations and California mastitis test. In addition, bacteriological isolation and identification of Staphylococcus aureus and MRSA were carried on the above milk samples of above mentioned quarters, teat swabs and environmental samples. Results: The overall prevalence of Staphylococcus aureus and MRSA was 59.4 and 22.4, respectively. The MRSA were more prevalent in mastitic quarter (X2 = 31.146, P-value = 0.000), Holstein Friesian breed (P-value = 0.021), Old aged dairy cattle (>8 years) (P-value = 0.000), Multiparous cows (P-value = 0.000), large herd size (>=2000) (P-value = 0.000). Conclusion and clinical relevance: The present result indicates that MRSA is a major cause of mastitis in dairy farms at northern Egypt, where one of the key elements for controlling its spreading depends mainly on determining its potential risk factors responsible for its existence.

This study analyzed differences between dairies that have closed compared with dairies still operating in the southeastern United States. Significant changes have occurred in the US dairy industry in the last decade, involving the number of dairy farms, herd size, milk quality, and management practices, yet the dairy industry remains the fourth leading agricultural sector in the United States, with $38 billion of milk sales in 2017. Although the number of dairy cows in the United States has remained relatively constant over the past decade, at approximately 9 million head, the number of dairy operations has decreased by 30%, resulting in larger dairies. This trend is even more prevalent in the southeastern United States, where the number of dairies has decreased by 39% from 5,315 in 2008 to only 3,235 in 2017. Additionally, downward pressure on bulk tank somatic cell count, which is used as a milk quality metric and has implications regarding animal health, intensified with US processors' introduction of incentive and penalty systems for quality milk production, necessitating better management of mastitis in dairy herds. In this context, this study examines factors that affect southeastern US dairy farms' persistence in the industry by using primary survey data collected in 2013 through a mail survey of Grade A dairies in Georgia, Mississippi, Kentucky, North Carolina, South Carolina, Tennessee, and Virginia. Dairies that were no longer operational had exited the industry from 2007 through 2014. A probit regression was used to determine which farm and operator characteristics were associated with the dairy's operational status. Dairy farms with more cows and higher average milk production per cow were more likely to be operational. For an additional 10 kg/d of milk production per cow, the dairy was 1.5% more likely to be operational. For each 100 additional cows a dairy had, it was 4% more likely to be operational. The analysis also identifies nonpecuniary determinants of operational status for southeastern US dairies, such as mastitis management practices. Findings suggest that operations capable of leveraging scale effects are more likely to remain operational, with results affirming the consolidation of the US dairy industry and demonstrating that more productive farms are more likely to stay in operation. Results also suggest that factors other than farm size affect a dairy's operational status.

This research communication explores the value of routinely collected bulk tank milk quality data for estimating dairy cattle welfare at herd level. Selected bulk tank milk quality parameters (somatic cell count, total bacterial count, urea, protein and fat contents) recorded during the years 2014–2016 in 287 Italian dairy farms were compared with the animal welfare data of each farm. The welfare assessment data were extracted from the database of the Italian Reference Centre for Animal Welfare (CReNBA), which includes the outputs of the application of the CReNBA welfare assessment protocol for dairy cows, used at national level for on-farm controls. The statistical analysis was carried out using the correlation coefficient for Kendall's Tau ranks, in order to investigate the presence of a categoric relationship between the selected bulk tank milk quality parameters and the overall animal welfare score or the scores of the single areas A (farm management and staff training), B (housing) and C (animal-based measures). Somatic cell count, total bacterial count, urea and proteins demonstrated only a few statistically significant and very weak correlations with farm animal welfare data, while no significant correlations were obtained for milk fat content. Given the weak correlations found, the selected bulk tank milk parameters seems to be able to provide only limited information about the welfare level of the herd, thus it could be difficult to use them for drawing up a pre-screening model for identifying herds at risk of poor welfare.

Staphylococcus aureus is an important intramammary pathogen for dairy cows that also is remarkably important for public health. Multiple virulence factors can be involved simultaneously during the pathogenesis of a staphylococcal disease, including adhesion proteins, extracellular enzymes, and toxins. The main objective of this study was to assess virulence factors that are associated with cow intramammary infection (IMI) and of human health concern among Staph. aureus isolates obtained from bulk tank milk (BTM) and adherences on milking equipment surfaces. A total of 166 Staph. aureus isolates from 23 dairy farms were characterized according to their virulence profiles. For virulence factors of importance in IMI, the presence of the virulence markers thermonuclease (nuc) and coagulase (coa) and virulence genes such as fibronectin (fnbA) and intercellular adhesion (icaA, icaD) were assessed. For virulence factors of public health concern, presence of antimicrobial resistance (mecA and mecC) and enterotoxin (sea and seb) genes were analyzed. Among all Staph. aureus isolates, 5 virulence profiles were found; the profile nuc(+)coa(+)fnbA(+)icaA(+)icaD(+)mecA(-)mecC(-)sea(-)seb(-) was the most frequently observed (21 out of 23 dairy farms). No differences were found between the virulence profile frequencies of Staph. aureus from BTM and adherences on milking equipment surfaces. The virulence profiles most frequently observed included genes involved in the adherence and biofilm-forming ability of Staph. aureus, which could represent a potential advantage for the bacterium during the early stages of IMI colonization and for persistence on surfaces. Our results indicate a greater frequency of virulence factors of importance for IMI pathogenesis than virulence factors of public health concern, consistent with the dairy origin of isolates. The mecA, mecC, and seb genes were not observed among Staph. aureus isolates analyzed in this study. However, the sea gene was detected in 3 Staph. aureus isolated from BTM, thus posing a potential public health threat. Our results emphasize the importance of understanding the epidemiology and dynamics of Staph. aureus on dairy farms as a tool for the improvement of udder health and milk safety.

Polymerase chain reaction (PCR) is typically used for the early detection of mycoplasma in bovine milk; it requires 3 days to obtain results because of the necessary enrichment process. A more rapid, simple, and accurate detection method is required to directly detect the Mycoplasma bovis (M. bovis) gene in milk. In this study, we assess the utility of combining the following two methods to achieve this goal: the loop-mediated isothermal amplification (LAMP), which is more sensitive than PCR, and the procedure for ultra rapid extraction (PURE), which adsorbs and filters components that inhibit DNA amplification/detection. LAMP was examined using DNA extracts obtained by four methods. This showed that PURE had the highest sensitivity and specificity and that the combination of PURE and LAMP was able to detect M. bovis in milk. We then showed that the detection limit of M. bovis was 102 colony-forming units per milliliter of milk using the PURE-LAMP. Finally, the respective sensitivities of the PURE-LAMP and PCR were 57% and 86% for bulk tank milk, 89% and 74% for mature milk, 85% and 92% for colostrum/transitional milk, and 97% and 95% for mastitis milk. The specificity was 100% for all milk samples in both LAMP and PCR. We conclude that PCR was suitable for detecting mycoplasma in bulk tank milk and that the PURE-LAMP could detect mycoplasma within 2 hr and was also effective for mature and mastitis milk.

A qualitative 3 min one-step assay for detecting beta-lactam, sulfonamide, and tetracycline antibiotics was validated following milk screening test guidelines developed by FDA-CVM, AOAC-RI, and IDF. The validated 90% detection levels with 95% confidence were: penicillin G 2 part per billion (ppb); amoxicillin 4 ppb; ampicillin 9 ppb; ceftiofur plus metabolites 50 ppb; cloxacillin 9 ppb; cephapirin 15 ppb; sulfadimethoxine 8 ppb; sulfamethazine 9 ppb; chlortetracycline 34 ppb; oxytetracycline 53 ppb; and tetracycline 42 ppb. Detection levels were lower than U.S. and Canadian allowable limits for milk and were consistent with most European Maximum Residue Limits. Tests of raw commingled cows’ milk indicated a low positive error rate of

Staphylococcus aureus is a Gram-positive bacterium that causes intramammary infections and bulk tank milk (BTM) contamination in dairy operations around the world in spite of on-farm application of preventive measures. The study was conducted on a 30-cow dairy farm in the Nuble Region of Chile. For BTM culture and somatic cell count (SCC) analysis, three consecutive BTM samples were collected. Samples for bacterial culture (n = 16) were collected from macroscopic adherence on previously washed, sanitized, and dry milking equipment surfaces in direct contact with milk during milking or cooling. A total of 48 S. aureus isolates from BTM, milking equipment, and cows' quarters with intramammary infections were analyzed by pulsed-field gel electrophoresis (PFGE). Selected milking equipment pieces were removed for biofilm visualization using scanning electron microscopy (SEM). S. aureus was isolated from all three BTM samples; the average SCC for the three BTM samples was 1,429,333 cells/mL. Fourteen of the 16 samples of milking equipment (87.5%) were culture positive for S. aureus. Biofilms were visualized by SEM in all four removed milking equipment pieces. Microorganisms observed by SEM in those biofilms were mainly coccus-shaped bacteria, and microbiological culture of these biofilms yielded viable S. aureus isolates in all samples. All pulsotypes observed among S. aureus isolates from BTM were indistinguishable from those in milking equipment surfaces. All PFGE pulsotypes observed among S. aureus isolates from biofilms on rubber liners were indistinguishable from isolates from intramammary infections in cows. Our findings suggest that milking equipment films may act as source of S. aureus contamination for BTM and cows during milking, thus compromising the microbiological quality of milk used for manufacturing dairy products.

Background and Aim: Consistency in producing raw milk with less variation in bulk tank milk somatic cell count (BMSCC) is important for dairy farmers as their profit is highly affected by it in the long run. Statistical process control (SPC) is widely used for monitoring and detecting variations in an industrial process. Published reports on the application of the SPC method to smallholder farm data are very limited. Thus, the purpose of this study was to assess the capability of the SPC method for monitoring the variation of BMSCC levels in milk samples collected from smallholder dairy farms. Materials and Methods: Bulk tank milk samples (n=1302) from 31 farms were collected 3 times/month for 14 consecutive months. The samples were analyzed to determine the BMSCC levels. The SPC charts, including the individual chart (I-chart) and the moving range chart (MR-chart), were created to determine the BMSCC variations, out of control points, and process signals for each farm every month. The interpretation of the SPC charts was reported to dairy cooperative authorities and veterinarians. Results: Based on a set of BMSCC values as well as their variance from SPC charts, a series of BMSCC data could be classified into different scenarios, including farms with high BMSCC values but with low variations or farms with low BMSCC values and variations. Out of control points and signals or alarms corresponding to the SPC rules, such as trend and shift signals, were observed in some of the selected farms. The information from SPC charts was used by authorities and veterinarians to communicate with dairy farmers to monitor and control BMSCC for each farm. Conclusion: This study showed that the SPC method can be used to monitor the variation of BMSCC in milk sampled from smallholder farms. Moreover, information obtained from the SPC charts can serve as a guideline for dairy farmers, dairy cooperative boards, and veterinarians to manage somatic cell counts in bulk tanks from smallholder dairy farms.

Comparison of bacterial counts (BCs) among common bedding types used for dairy cows, including straw, is needed. There is concern that the microbial content of organic bedding is elevated and presents risks for dairy cow udder health and milk quality. The objectives of this study were to investigate: (1) % DM and BCs (Streptococcus spp., all gram-negatives and specifically Klebsiella spp.) in different types of bedding sampled, and to investigate housing and farm management factors associated with % DM and BCs; (2) if bedding type was associated with hygiene of cow body parts (lower-legs, udder, upper-legs and flank) and housing and management factors associated with hygiene and (3) bedding types associated with higher BCs in cow milk at the farm level and bulk tank milk and management factors that were associated with highest BCs. Seventy farms (44 free-stall and 26 tie-stall) in Ontario, Canada were visited 3 times, 7 days apart from October 2014 to February 2015. At each visit, composite samples of unused and used bedding were collected for % DM determination and bacterial culture. Used bedding samples were collected from the back third of selected stalls. Data were analyzed using multivariable linear mixed models. Bedding classification for each farm were: new sand (n = 12), straw and other dry forage (n = 33), wood products (shavings, sawdust; n = 17) and recycled manure solids (RMSs)-compost, digestate (n = 8). In used bedding, across all bedding samples, sand was driest, compared to straw and wood, and RMS; higher % DM was associated with lower Streptococcus spp. count. Streptococcus spp. and all Gram-negative bacteria counts increased with increasing days since additional bedding was added. Gram-negative bacteria counts in used bedding varied with type: RMS = 16.3 ln colony-forming units (cfu)/mL, straw = 13.8 ln cfu/mL, new sand = 13.5 ln cfu/mL, and wood = 10.3 ln cfu/mL. Klebsiella spp. counts in used bedding were lower for wood products (5.9 ln cfu/mL) compared to all other bedding types. Mean cow SCC tended to be higher on farms with narrower stalls. Farms with mattress-based stalls had a higher prevalence of cows with dirty udders compared to those using a deep bedding system (often inorganic sand). Wider stalls were associated with lower bulk milk bacteria count. Lower % DM of used bedding was associated with higher bulk milk bacteria count. In conclusion, bedding management may have a profound impact on milk quality, bacterial concentrations in the bedding substrates, and cow hygiene.

Aim: This study aimed to evaluate and quantify the different factors affecting the costs of mastitis in cattle, to quantify the annual and monthly financial losses attributed to mastitis, and to estimate production losses using average linear scores found on The Dairy Herd Improvement Association somatic cell count (SCC) sheets and bulk tank SCC per lactation number. Materials and Methods: All data (bovine mastitis-associated costs and expenditures in Egyptian dairy herds) were analyzed using SPSS/PCT, 2001. A partial budget technique using spreadsheet software and the general linear model procedure was used to analyze the productive and financial measures. Results: Mastitis was present in 57.1% of cows (240/420), while clinical mastitis (CM) was present in 19% of them (80/420). The geometric mean of SCC/ml in bulk tank milk samples of 150 cattle dairy farms was 556.3×103. The annual subclinical mastitis (SCM)-related economic loss was 21,933,258.6 LE, and the two most important cost components were the subsequent decrease in milk production and quality premium losses (93% and 7% of SCM costs, respectively). The quality premium loss was 1,369,602.1 LE. On the other hand, the annual economic loss due to decreased milk production as a result of SCM was 20,563,656.5 LE. The total cost of 80 CM cases, including the failure and preventive costs, was 1,196,871.4 LE, including 1,169,150.4 LE failure costs (106,336.0 LE in direct costs and 1,062,814.4 LE in indirect costs) and 27,721.0 LE preventive costs. The average cost per CM case was 28,760.9 LE, including veterinary time and consultation fees of 250.0 LE (1%), labor 562.5 LE (2%), premature culling 736,000.0 LE (77%), decreased milk production 4085.18 LE (13.7%), discarded milk 185.3 LE (1%), and drugs and treatments 328.9 LE (1%). The total costs of CM (expenditures) extra control and preventive measures, including the diagnosis of CM for 80 cows annually in 20 Egyptian dairy farms were 27,721.0 LE, representing 346.5 LE or 1% of the total cost of CM cases. The cost of monitoring and diagnostic measures was 8635.2 LE, representing 107.9 LE or 1% of the total cost of a case of CM. Conclusion: The method used for cost estimation, in this study, is highly adaptable to individual cattle farms and had a major role in assessing specific control and management measures. The concepts described in this paper help to improve our understanding of the full economic impact of clinical and subclinical mastitis in cattle in Egypt. Assessing the economic losses from mastitis to determine the economic costs and losses occurring in Egyptian dairy farms is critical for encouraging farmers to acknowledge the scale of the problem and implement effective management practices aimed at improving mastitis control and reducing the associated costs.

Food contaminated with Shiga toxin-producing Escherichia coli (STEC) represents a hazardous public health problem worldwide. Therefore, the present study was performed to elucidate the virulent and antimicrobial resistance characteristics of STEC isolated from milk and dairy products marketed in Egypt. A total of 125 samples (raw market milk, bulk tank milk, Kareish cheese, white soft cheese, and small scale-produced ice cream, 25 each) were collected for determination the prevalence and antimicrobial resistance profiling of STEC. Thirty-six STEC isolates were recovered from milk and dairy products. Serological analysis illustrated that three isolates were E. coli O157:H7 and 33 isolates belonged to different serotypes. Molecular examination indicated that all isolates harboured stx1 and/or stx2 genes, 14 isolates expressed eaeA gene and 3 isolates possessed rfbE gene. Antimicrobial resistance profiling of the isolates was both phenotypically and genetically examined. Interestingly, 31 out of 36 (86.11%) isolates were multidrug-resistant and harboured the extended-spectrum β-lactamase encoding genes, namely, blaCTX-M-15, blaSHV-12 and blaCTX-M-14. Moreover, 12 isolates (33.33%) harboured plasmid-mediated quinolone resistant gene, qnrS. The overall conclusion of the current investigation indicated insufficient hygienic measures adopted during milking, handling, and processing leading to development of pathogenic and multidrug-resistant STEC.

Sporeforming bacteria are responsible for the spoilage of several dairy products including fluid milk, cheese, and products manufactured using dried dairy powders as ingredients. Sporeforming bacteria represent a considerable challenge for the dairy industry because they primarily enter the dairy product continuum at the farm, survive processing hurdles, and subsequently grow in finished products. As such, strategies to reduce spoilage due to this group of bacterial contaminants have focused on understanding the effect of farm level factors on the presence of spores in bulk tank raw milk with the goal of reducing spore levels in raw milk, as well as understanding processing contributions to spore levels and outgrowth in finished products. The goal of the current study was to investigate sources of spores in the farm environment and survey farm management practices to identify variables using multimodel inference, a model averaging approach that eliminates the uncertainty of traditional model selection approaches, that affect the presence and levels of spores in bulk tank raw milk. To this end, environmental samples including feed, bedding, manure, soil, water, and so on, and bulk tank raw milk were collected twice from 17 upstate New York dairy farms over a 19-mo period and the presence and levels of various spore types (e.g., psychrotolerant, mesophilic, thermophilic, highly heat resistant thermophilic, specially thermoresistant thermophilic, and anaerobic butyric acid bacteria) were assessed. Manure had the highest level of spores for 4 out of 5 aerobic spore types with mean counts of 5.87, 5.22, 4.35, and 3.68 log cfu/g of mesophilic, thermophilic, highly heat resistant thermophilic, and specially thermoresistant thermophilic spores, respectively. In contrast, bulk tank raw milk had mean spore levels below 1 log cfu/mL across spore types. Multimodel inference was used to determine variables (i.e., management factors, environmental spore levels, and meteorological data from each sampling) that were important for presence or levels of each spore type in bulk tank raw milk. Analyses indicated that variables of importance for more than one spore type included the residual level of spores in milk from individual cows after thorough teat cleaning and forestripping, udder hygiene, clipping or flaming of udders, spore level in feed commodities, spore level in parlor air, how often bedding was topped up or changed, the use of recycled manure bedding, and the use of sawdust bedding. These results improve our understanding of how spores transfer from environmental sources into bulk tank raw milk and provide information that can be used to design intervention trials aimed at reducing spore levels in raw milk.

Recommended milking practices (RMP) are protective against mastitis. However, many producers do not adopt, or only partially adopt, these measures. This study aimed to explore the attitudes and perceptions of Ontario dairy farmers toward barriers to implementation of RMP and to investigate what motivates behavior change in relation to milking hygiene. Four focus groups with Ontario dairy producers were conducted, and verbatim transcripts were analyzed thematically. The main barriers to adoption of RMP were identified and categorized into 2 groups: intrinsic barriers and physical barriers. Intrinsic barriers included personal habits and convenience, not perceiving udder health as a priority on their farm, and lack of information. Physical barriers included employee training and compliance, convenience of implementing RMP, and time, money, and labor barriers. Producers used their bulk tank somatic cell count (SCC) as a measure of perceived severity of udder health problems on farm. Those with lower SCC were less likely to prioritize udder health compared with peers experiencing elevations in SCC. Lack of udder health problems translated for some producers into non-adoption of certain RMP, as they felt these practices were not needed unless a problem arose. Others felt motivated to implement more practices and work toward better udder health if such efforts translated into rewards for better-quality milk. Some producers perceived RMP as not meaningful or useful, seemingly due to a lack of education about the reasons behind RMP implementation. Understanding the importance of these practices is one key to implementing them. To overcome some of the intrinsic barriers, increased efforts in knowledge translation are needed, including efforts in retraining current practices, as well as in establishing best practices.

Staphylococcus aureus (S. aureus) is now regarded as a zoonotic agent. Methicillin-susceptible S. aureus (MSSA) ST398 is a livestock-associated bacterium that is most prevalent in China, but there are currently no data available for Shandong. Therefore, the aim of this study was to investigate the epidemiology and characterization of MSSA ST398 from retail pork and bulk tank milk (BTM) in Shandong. A total of 67 S. aureus isolates were collected from retail pork between November 2017 and June 2018. Among the isolates, high antimicrobial resistance rates were observed for penicillin (97.0%), and 92.5% of the isolates were multi-drug resistant (MDR). Eight sequence types (STs) were identified in the retail pork isolates, and the predominant type was ST15 (n=26), which was followed by ST398 (n=14). Staphylococcal protein A gene (spa) typing identified spa types t034 and t1255 in MSSA ST398 from retail pork. Using whole-genome sequencing analysis, we described the phylogeny of 29 MSSA ST398 isolates that were obtained from retail pork (n=14) and BTM (n=15). The phylogenetic tree showed that the MSSA ST398 isolates from different sources had the same lineage. Among the 29 MSSA ST398 isolates, five resistance genes were detected, and all isolates carried DHA-1. Fifteen toxin genes were detected, and all isolates carried eta, hla, and hlb. In conclusion, this study found that a high risk for MSSA ST398 was present in retail pork and BTM. These findings have major implications for how investigations of MSSA ST398 outbreaks should be conducted in the One-Health context.

The bovine udder is colonized by a huge quantity of microorganisms that constitute the intramammary ecosystem, with a specific role in modulating not only udder homeostasis and mastitis susceptibility, but also the quality of the dairy products. However, generating high-quality bacterial DNA can be critical, especially starting from a complex biological matrix like milk, characterized by high fat, protein, and calcium contents. Here, bacterial DNA was recovered from a commercial ultra-high-temperature (UHT) milk sample artificially spiked with a predetermined mock community composition and from three bulk tank milk (raw milk) samples. The DNA was isolated using three different protocols to evaluate the effect of the extraction procedures on the milk microbiota composition. In the mock community experiment, the bacterial profiles generated by the three DNA extraction protocols were profoundly different, with the genera Staphylococcus, Lactobacillus, Listeria, and Salmonella underestimated by all the protocols. Only one protocol revealed values close to the expected abundances for Escherichia/Shigella spp., Bacillus spp., Enterococcus spp., and Pseudomonas spp. On the other hand, the nonspiked UHT milk sample exhibited a similar microbiota composition, revealing the prevalence of Acinetobacter spp., for all the DNA extraction protocols. For the raw milk samples, the three DNA extraction kits performed differently, revealing significant separations in both the microbial richness (alpha diversity) and composition (beta diversity). Our study highlights the presence of significant differences among these procedures, probably due to the different DNA extracting capacities and to the different properties of the milk samples, revealing that the selection of DNA extraction protocol is a critical point. IMPORTANCE The advance of high-throughput technologies has increased our knowledge of the world of microorganisms, especially of microbial populations inhabiting living animals. This study provides evidence that milk, as other complex sources, could be critical for generating high-quality DNA for microbiota analysis. In addition, it demonstrates that the microbial population highlighted by metagenomic studies changes in relation to different DNA extraction procedures, revealing that attention should be paid especially when comparing different studies.

Cleaning of the production environment is vital to ensure the safety and quality of dairy products. Although cleaning with chlorine-based agents is widely adopted, it has been associated with detrimental effects on milk quality and safety, which has garnered increasing interest in chlorine-free cleaning. However, the influence of these methods on the milk microbiota is not well documented. This study investigated the factors that influence the raw milk microbiota, with a focus on the differences when chlorine-based and chlorine-free cleaning of milking equipment are used. Bulk tank raw milk was sampled during three sampling months (April, August, and November), from farms across Ireland selected to capture the use of different cleaning methods, i.e., exclusively chlorine-based (n = 51) and chlorine-free cleaning (n = 92) and farms that used chlorine-free agents for the bulk tank and chlorine-based cleaning agents for the rest of the equipment (n = 28). Shotgun metagenomic analysis revealed the significant influence of seasonal and geographic factors on the bulk tank milk microbiota, indicated by differences in diversity, taxonomic composition, and functional characteristics. Taxonomic and functional profiles of samples collected in November clustered separately from those of samples collected in other months. In contrast, cleaning methods only accounted for 1% of the variation in the bulk tank milk bacterial community, and samples collected from farms using chlorine-based versus chlorine-free cleaning did not differ significantly, suggesting that the chlorine-free approaches used did not negatively impact microbiological quality. This study shows the value of shotgun metagenomics in advancing our knowledge of the raw milk microbiota. IMPORTANCE The microbiota of raw milk is affected by many factors that can control or promote the introduction of undesirable microorganisms. Chlorine-based cleaning agents have been commonly used due to their effectiveness in controlling undesirable microorganisms, but they have been associated with the formation of chlorine residues that are detrimental to product quality and may impact consumer health. Chlorine-free alternatives have been recommended in some countries, but the influence of cleaning agents on the milk microbiota is unknown. Here, we investigated the influence of cleaning methods and other factors on bulk tank raw milk. Results showed that season and location had a greater influence on the milk microbiota than the cleaning agents used. Indeed, the similar microbiota compositions of raw milk from farms that used chlorine-based and those that used chlorine-free cleaning methods supports the further use of chlorine-free cleaning agents in dairy production.

The treatment of clinical mastitis on farm is always under the spotlight for veterinary surgeons due to the financial penalty of withholding milk from the bulk tank, the challenge of achieving a bacteriological cure during lactation compared with during a dry period and the importance of antibiotic use in dairy herds. This article focuses on the management of the FIRST clinical mastitis case in a cow's lactation, as this not only represents the best chance of achieving a bacteriological cure as well as a symptomatic one, but also puts emphasis on the need to measure the outcome of treatment approaches for first cases and the likely origin of first cases so that treatment can be avoided in the future.