Bovine Mastitis Research Articles

Succinate exacerbates mastitis in mice via extracellular vesicles derived from the gut microbiota: a potential new mechanism for mastitis.

A high grain diet causes an ecological imbalance in the gut microbiota and serves as an important endogenous trigger of mastitis in dairy cows, but the underlying mechanisms are unclear. Our previous study revealed that subacute rumen acidosis (SARA)-associated mastitis has distinct metabolic profiles in the rumen, especially a significant increase in succinate, but the role of succinate in the pathogenesis of mastitis remains unclear. Succinate treatment exacerbates low-grade endotoxemia-induced mastitis in mice. Specifically, succinate increased the production of gut microbiota-extracellular vehicles (mEVs) containing lipopolysaccharides, which can diffuse across the damaged intestinal barrier into the mammary glands. Administration of mEVs promotes mammary inflammation via activation of the TLR4/NF-κB pathway. Our findings suggest that succinate promotes mastitis through the proliferation of enteric pathogens and mEVs production, suggesting a potential strategy for mastitis intervention on the basis of intestinal metabolic regulation and pathogen inhibition. The role of mEVs in interspecific communication has also been elucidated.

MastitisApp: a software for preventive diagnosis of mastitis in dairy cows

Dairy farming has great economic value in Brazil, however, during production, diseases such as mastitis can occur in animals, which can reduce productivity and, consequently, economic profitability. When mastitis is present in animals, it can cause physical and chemical changes in the milk, affecting its quality, market value and also compromising the health of the animal. MastiteApp is a tool to help producers prevent mastitis in their herds by checking the temperature taken from the four teats of the animal. To perform the analysis, the temperature of all the animals' teats must be measured and, if there is a change in temperature, the system will display a message informing the producer of the possible presence of subclinical mastitis in their animal. The application has proven to be efficient in alerting producers to the possible presence of subclinical mastitis in the first few days of manifestation, thus initiating treatment and preventing the disease from worsening.

Mycoplasma bovis Invades Non-Phagocytic Cells by Clathrin-Dependent Endocytic Pathways and Escapes from Phagocytic Vesicles

Mycoplasma bovis (M. bovis) is capable of causing pneumonia, arthritis, mastitis, and various other ailments in cattle of all age groups, posing a significant threat to the healthy progression of the worldwide cattle industry. The invasion of non-phagocytic host cells serves as a pivotal mechanism enabling M. bovis to evade the immune system and penetrate mucosal barriers, thereby promoting its spread. To investigate the differences in M. bovis invasion into four types of non-phagocytic cells (Madin–Darby bovine kidney (MDBK) cells, embryonic bovine lung (EBL) cells, bovine embryo tracheal (EBTr) cells and bovine turbinate (BT) cells) and further elucidate its invasion mechanism, this study first optimized the experimental methods for M. bovis invasion into cells. Utilizing laser scanning confocal microscopy, transmission electron microscopy, and high-content live-cell imaging systems, the invasion process of M. bovis into four types of non-phagocytic cells was observed. The invasion rates of three different strains of M. bovis (PG45, 07801, 08M) were quantified through the plate counting method. In order to clarify the specific pathway of M. bovis invasion into cells, chlorpromazine (CPZ), amiloride (AMI), and methyl-β-cyclodextrin (M-β-CD) were used to inhibit CLR-mediated clathrin-dependent endocytosis (CDE) pathway, macropinocytosis, and lipid raft pathway, respectively. Subsequently, the invasion rates of PG45 into these four types of cells were measured. Using siRNA technology, the expression of clathrin (CLR) in EBL cells was knocked down to further verify the role of CLR in the invasion process of M. bovis. The results showed that the optimal conditions for M. bovis to invade non-phagocytic cells were a multiplicity of infection (MOI) of 1000 and an optimal invasion time of 4 h. All three strains of M. bovis have the ability to invade the four types of non-phagocytic cells, yet their invasion abilities vary significantly. Observations from transmission electron microscopy further confirmed that at 120 min post-infection, PG45 had successfully invaded EBL cells and was present within endocytic vesicles. It is noteworthy that almost all PG45 successfully escaped from the endocytic vesicles after 240 min of infection had passed. Through chemical inhibition experiments and CLR protein knockdown experiments, it was found that when the CDE and lipid raft pathways were blocked or CLR protein expression was reduced, the invasion rates of PG45, 07801, and 08M in MDBK, EBL, EBTr, and BT cells were significantly decreased (p < 0.05). The above results indicate that M. bovis can invade all types of non-phagocytic cells through endocytic pathways involving CDE (clathrin-dependent endocytosis) or lipid raft-mediated endocytosis, and possesses the ability to escape from phagosomes.

Bactericidal activities and biochemical features of 16 antimicrobial peptides against bovine-mastitis causative pathogens.

Mastitis, often caused by bacterial infection, is an inflammatory condition affecting the mammary glands. The condition is particularly prevalent in dairy cattle. Current treatment of bovine mastitis heavily relies on the use of antibiotics. To identify alternative solutions to antibiotic use, we evaluated the antimicrobial activity of 14 cathelicidins reported from 10 animal species. In conjunction, we assessed two bacteriocins against the bovine-mastitis causative bacterial panel, consisting of Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus, Bacillus cereus, Enterococcus faecalis, Streptococcus agalactiae, Streptococcus dysgalactiae, and Streptococcus equi. Among the antimicrobial peptides (AMPs), cc-CATH3, ML-CATH, and PD-CATH proved to be highly active (minimum inhibitory concentration of 2-41μg/mL, 0.2-10.3μM) against all bacterial strains in the panel and field isolates from milk, with elevated somatic cell counts (≥ 500,000 cells/mL). Of the AMPs tested in this study, ML-CATH presented the highest level of effectiveness in controlling mastitis-associated bacterial strains while also possessing minimal cytotoxicity and functional stability against pH change and a high salt condition. The results of in silico analyses on the biochemical features of 12 helical cathelicidins revealed that the charge of AMPs appears to be a major determinant in killing Gram-negative bacteria. Furthermore, we observed a unique motif, "N(n≥3)-P(n≥1)-N(n≥3)", from the sequences of PMAP-36, cc-CATH3, ML-CATH, and PD-CATH that exhibits potent antimicrobial activity against a broad spectrum of bacteria compared to others. Our findings support the proposition that AMPs could serve as effective antimicrobial alternatives to conventional antibiotics in treating complex animal diseases caused by microbial infection, such as bovine mastitis.

Blood-feeding stable flies, Stomoxys calcitrans (Diptera: Muscidae), are attracted to, and transmit Staphylococcus aureus (Bacillales: Staphylococcaceae), a causal agent of bovine mastitis: a laboratory pilot study.

Stable flies, Stomoxys calcitrans (Linnaeus) (Diptera: Muscidae), are common blood-feeding ectoparasites of cows and thus potential vectors of the skin-dwelling bacterium Staphylococcus aureus, a causal agent of bovine mastitis which inflicts udder inflammation in cows. Our objectives were to determine whether stable flies (i) are attracted to disease-causing strains of S. aureus, and (ii) transmit S. aureus from infected blood to sterile blood. In 3-chamber olfactometers, five of eight S. aureus strains grown on agar and tested versus sterile agar attracted female stable flies. When flies ingested droplets of blood inoculated with S. aureus at doses of 0 (control), 105 (low), 107 (medium), and > 109 (high) colony-forming units per milliliter and subsequently ingested sterile blood, they transmitted S. aureus to the sterile blood. The dose of S. aureus in blood droplets fed upon by flies during their first feeding bout dose-dependently affected the amount of bacteria that flies transmitted to sterile blood during their second feeding bout, but the time elapsed between feeding bouts (0h, 1h, 8h, and 24h) had no effect on the amount of microbes transmitted to sterile blood. Our data infer the existence of a positive feedback loop. First, stable flies carrying S. aureus and feeding on cows transmit S. aureus, thereby causing mastitis. As S. aureus bacteria of afflicted cows proliferate, they attract even more flies which, in turn, worsen the infection. This type of feedback loop underscores the need for effective stable fly control tactics that curtail the incidence of bovine mastitis in cows.

Milk metabolite profiling of dairy cows as influenced by mastitis

Mastitis is a disease with frequent incidence in dairy cows, causing huge financial losses to the dairy industry globally. The identification of certain biomarkers is crucial for the early diagnosis and management of mastitis. Metabolomics technology is a useful tool to accurately and efficiently analyze the changes of metabolites in biofluids in response to internal and external stimulations. Milk is the secreted by udder, and milk metabolites can directly reflect whether the udder are in the healthy or diseased state. The milk metabolomics analysis of mastitis can reveal the physiological and pathological changes of mammary gland and screen the related biomarkers, so as to offer useful reference for the prediction, diagnosis, and management of mastitis. Therefore, the aim of the present study was to comprehensively summarize milk metabolic change caused by naturally occurring or experimentally induced mastitis in dairy cows. In addition, comparative analysis and enrichment analysis were conducted to further discover potential biomarkers of mastitis and to identify the relevant pathways differentiating the healthy and mastitic cows. Multiple milk metabolites were identified to be altered during mastitis based on different metabolomics platforms. It was noteworthy that there were 28 metabolites not only identified by at least two different studies, but also showed consistent change tendency among the different studies. By comparison with literature, we further identified 12 milk metabolites, including acetate, arginine, β-hydroxybutyrate, carnitine, citrate, isoleucine, lactate, leucine, phenylalanine, proline, riboflavin, and valine that were linked with the occurrence of mastitis, which suggested that these 12 milk metabolites could be potential biomarkers of mastitis in dairy cows. Several pathways were revealed to explain the mechanisms of the variation of milk metabolites caused by mastitis, such as phenylalanine, tyrosine and tryptophan biosynthesis, arginine and proline metabolism, riboflavin metabolism, and tricarboxylic acid (TCA) cycle. These results offer a further understanding for the alteration of milk metabolites caused by mastitis, which have a potential significance in the development of more reliable biomarkers for mastitic diagnosis in dairy cows.

Mycoplasma bovis 5'-nucleotidase is a virulence factor conferring mammary fitness in bovine mastitis.

Nucleases and 5' nucleotidase (5'-NT) play essential roles in cell biology and are often associated with bacterial virulence. In Mycoplasma spp., which have limited metabolic capacities and rely on nutrient availability, these enzymes are of significant importance for nucleotide salvage. This study explores the potential role of 2 membrane-associated lipoproteins, the major nuclease MnuA and 5'-NT, in Mycoplasma bovis mastitis. Mutants deficient in MnuA (mnuA::Tn) and in 5'-NT (0690::Tn) were identified through genome-wide transposon mutagenesis of M. bovis PG45 type strain and their fitness and virulence were assessed both in vitro, in axenic medium, and in vivo, using murine and cow mastitis models. The mnuA::Tn mutant demonstrated reduced nuclease activity, while 0690::Tn exhibited slow log-phase growth and impaired hydrolase activity towards nucleotides as well as deoxynucleotides (dAMP and dGMP). In comparison to the parent strain, the 0690::Tn mutant displayed markedly reduced fitness, as evidenced by a significant decrease or even absence in post-challenge mycoplasma counts in murine and cow mammary tissues, respectively. Moreover, the 0690::Tn mutant failed to induce mastitis in both experimental models. Conversely, the mnuA::Tn mutant induced inflammation in murine mammary glands, characterized by neutrophil infiltration and increased expression of major inflammatory genes. In cows, the mnuA::Tn was able to cause an increase in somatic cell counts in a manner comparable to the wild type, recruit neutrophils, and induce mastitis. Collectively, these findings provide complementary insights, revealing that disruption of 5'-NT significantly attenuated M. bovis pathogenicity, whereas a MnuA-deficient mutant retained the ability to cause mastitis.

Effect of nanoemulsion loaded with macela (Achyrocline satureioides) on the ultrastructure of Staphylococcus aureus and the modulating activity of antibiotics

Nanoformulations with herbal actives for treating bovine mastitis present an alternative for controlling bacterial infections in the emerging scenario of antimicrobial resistance. In this study, we investigated macela (Achyrocline satureioides) nanoemulsion (NE-ML), a formulation developed for the treatment of bovine mastitis (registered under Brazilian patent application BR 10 2021 008630 0), in the context of its bactericidal mechanism(s) of action and potential synergism with commercial antimicrobials. The effect of NE-ML on the integrity and cell permeability of Staphylococcus aureus was evaluated by measuring the electrical conductivity of bacterial suspensions exposed to different concentrations of NE-ML and by assessing the release of cellular constituents. Damage to bacterial ultrastructures was analyzed by transmission electron micrographs. The synergism of NE-ML with beta-lactam antibiotics and aminoglycosides was evaluated by the checkerboard test method against S. aureus (n = 6). The relative electrical conductivity of the bacterial solution gradually increased over time, reaching high values after exposure to 1xMIC (52.3%) and 2xMIC (75.34%) of NE-ML. Total proteins were detected in the bacterial suspensions exposed to NE-ML, increasing in concentration over exposure time (p < 0.05). Through bacterial micrographs, we observed that exposure to NE-ML (1xMIC) affected the integrity of the plasma membrane with invaginations in the cytosolic region and alterations in the cell wall. The increase in NE-ML concentration resulted in greater damage to the ultrastructure of S. aureus with changes in bacterial cell division patterns. When NE-ML was combined with the beta-lactam antimicrobials, the interaction was indifferent, indicating no modulation of antimicrobial resistance. In contrast, when combined with the aminoglycoside, a synergistic interaction did occur. These general findings suggest that the bactericidal action of NE-ML begins in the plasma membrane, causing alterations in its permeability and integrity, and extends to the cell wall, cytoplasm, and cell division. Although synergy was restricted to the aminoglycoside by destabilizing the bacterial cell membrane, this suggests that NE-ML can induce the entry of other actives, potentially reducing their therapeutic doses. Understanding the mechanism of action of this new nanoformulation is certain to drive pharmacological advances, broaden the perspective of its in vivo use, and improve the treatment of bovine mastitis.

A New Way to Identify Mastitis in Cows Using Artificial Intelligence

Mastitis is a disease that is considered an obstacle in dairy farming. Some methods of diagnosing mastitis have been used effectively over the years, but with an associated relative cost that reduces the producer’s profit. In this context, this sector needs tools that offer an early, safe, and non-invasive diagnosis and that direct the producer to apply resources to confirm the clinical picture, minimizing the cost of monitoring the herd. The objective of this study was to develop a predictive methodology based on sequential knowledge transfer for the automatic detection of bovine subclinical mastitis using computer vision. The image bank used in this research consisted of 165 images, each with a resolution of 360 × 360 pixels, sourced from a database of 55 animals diagnosed with subclinical mastitis, all of which were not exhibiting clinical symptoms at the time of imaging. The images utilized in the sequential learning transfer were those of MammoTherm, which is used for the detection of breast cancer in women. The optimized model demonstrated the most optimal network performance, achieving 92.1% accuracy, in comparison to the model with manual search (86.1%). The proposed predictive methodologies, based on knowledge transfer, were effective in accurately classifying the images. This significantly enhanced the automatic detection of both healthy animals and those diagnosed with subclinical mastitis using thermal images of the udders of dairy cows.

Diversity of Staphylococcus aureus enterotoxin genes and its potential impact on severity of mastitis in dairy cows.

Staphylococcus aureus enterotoxins (SEs) can lead to food poisoning and appear to play a pivotal role in the development and severity of mastitis. This study aimed to evaluate whether the presence and diversity of S. aureus Enterotoxin genes mastitis-causing are associated with the development and severity of mastitis in dairy cows. Comparative analysis of S. aureus isolates from cows with clinical and subclinical mastitis (103 isolates each, CM and SM respectively) was conducted, examining the occurrence of enterotoxins (sea-see, seg-sez, sel26, sel27, sel01 and se02). Clinical isolates exhibited greater diversity, with 14 genes compared to four in subclinical isolates, and higher gene frequency (100% vs. 86.4%). Among the classical SEs, only sec (1%) and sed (4.9%) were detected in clinical isolates, suggesting a potential insignificance in the development of mastitis. Conversely, the selw and selx genes were among the most prevalent in both clinical and subclinical mastitis isolates. While their role in human food poisoning has not been definitively established, they appear to influence mastitis development. Statistical analysis revealed significant differences in clinical isolates carrying seg, seh, sei, seo, seul1, seul2, selw, and selx genes compared to subclinical isolates, suggesting their potential involvement in the severity of mastitis. Determining the frequency of SE gene and its role in mastitis establishment and in clinical mastitis severity, as well as understanding how enterotoxins contribute to pathogenicity in clinical and subclinical mastitis, is crucial for guiding optimal medical therapy for animals and establishing effective veterinary treatments to mitigate economic losses in the dairy industry.

Are SNPs Linked to Somatic Cell Score Suitable Markers for the Susceptibility to Specific Mastitis Pathogens in Holstein Cows?

Mastitis in cattle is often caused by microorganism infections in the udder. The three most common pathogens are esculin-positive streptococci (SC+), coagulase-negative staphylococci (CNS), and Escherichia coli (E. coli). In a previous study, 10 SNPs were associated with somatic cell score and mastitis in diverse Holstein populations. We tested these SNPs for their effects on individual pathogen presence. Milk and pathogen samples of 3076 Holstein cows were collected from four farms. Samples were excluded if multiple pathogens were present at the same time. Records of the same pathogen within 14 days of each other were counted as one infection. This resulted in 1129 pathogen-positive samples. Cases and controls were in ratios of 20:80 for SC+, 8:92 for CNS, and 11:89 for E. coli. The lasso, backward, and forward methods were used to narrow down SNPs associated with pathogen presence. The suitability of the SNPs to separate the samples into cases or controls for each pathogen was indicated using ROC curves. The Cochran-Armitage (CAT) and the Jonckheere-Terpstra (JTT) tests evaluated the influence of the SNPs on pathogen presence. Finally, a generalised linear mixed model (GLMM) including fixed environmental effects and a random sire effect was fitted to the binary trait of pathogen presence to test for association. In total, six out of the 10 investigated SNPs showed associations with pathogen presence based on the forward method: Two SNPs each for SC+ (rs41588957, rs41257403) and CNS (rs109934030, rs109441194), and three for E. coli (rs109934030, rs41634110, rs41636878). The CAT and GTT tests linked four SNPs (rs41588957, rs41634110, rs109441194, rs41636878) to pathogen presence, two of which were confirmed with the GLMM (rs41634110, rs109441194), with effects on CNS and E. coli. The SNPs linked to CNS and those linked to E. coli explained 13.2% and 13.8% of the variance, compared to 19% and 18.4%, respectively, of the full model with all 10 SNPs. Half of the SNP genotypes previously linked to lower SCS also decreased the probability for pathogen presence and might therefore be targets not just for lower SCS but for a better pathogen resistance. Trial Registration: Not applicable, no new data were collected for this study.

Selective Dry Cow Therapy: Clinical Field Trial on Prevention and Cure of Intramammary Infections

Intramammary antibiotic dry cow therapy (DCT) at the end of lactation is one of the key measures in the management and control of bovine mastitis. Currently, livestock production is under pressure to reduce antibiotic consumption, emphasizing the need to avoid medicating all cows at dry-off, and instead to treat only infected cows. The study objective was to evaluate IMI cure risk, new IMI risk, and post-calving IMI risk between DCT-treated and untreated quarters over the dry period. The sample totaled 269 cows from 12 Finnish automatic milking system herds entering the dry period between 2019 and 2021. Cows eligible for the study had a final DHI test-day SCC ≥ 100,000 cells/mL and had no clinical signs of mastitis. Based on odd or even identification numbers, cows with an SCC between 100,000 and 250,000 cells/mL were sequentially assigned to either receive DCT at dry-off (DCT100-250) or remain untreated (NoDCT100-250). All cows with an SCC > 250,000 cells/mL received DCT (DCT > 250). No quarters received teat sealants. Farmers collected aseptic quarter-milk samples for microbiological quantitative real-time PCR (qPCR) analysis 0 to 4 d before dry-off and 0 to 4 d after calving. The outcome of the statistical model was the odds of a quarter having an IMI at freshening 0 to 4 d after calving. The statistical tool was a generalized linear mixed-effects model with logit link function and 2-level random intercepts, cows nested within herds. Quarter-level IMI prevalence at dry-off was NoDCT100-250 17.6% (45/256), DCT100-250 22.2% (83/374), and DCT > 250 32.0% (132/413). Quarter-level IMI prevalence at freshening was NoDCT100-250, 36.2% (84/232), DCT100-250, 13.6% (50/369), and DCT > 250, 11.6% (46/397). The untreated quarters in the NoDCT100-250 group had a 1.1 times higher risk of failing to cure over the dry period than did the treated quarters in the DCT100-250 group (unadjusted RR 1.10, 95% CI 0.94-1.28) and a 1.2 times higher risk than the treated quarters in the DCT > 250 group (unadjusted RR 1.15, 95% CI 1.00-1.33). The untreated quarters in the NoDCT100-250 group had a 1.5 times higher risk of having a new IMI over the dry period than did the treated quarters in the DCT100-250 group (unadjusted RR 1.48, 95% CI 1.35-1.63) and a 1.4 times higher risk than the treated quarters in the DCT > 250 group (unadjusted RR 1.35, 95% CI 1.23-1.49). The principal cause of new IMI was NAS, followed by Streptococcus dysgalactiae. Quarters in the NoDCT100-250 group had 6.6 times higher odds for having an IMI at freshening than did quarters in the DCT100-250 group (95% CI 3.52-12.21). According to our findings within the framework of this experiment, selectively DCT-treated cows with an SCC between 100,000 and 250,000 cells/mL at dry-off demonstrated that, although the group-specific cure risks over the dry period were high, untreated quarters had a higher risk of acquiring a new IMI over the dry period than did treated quarters, highlighting the effectiveness of DCT in reducing IMI risks. The prophylactic use of antibiotics, however, is not recommended, and in some countries is even prohibited.

Correlation Analysis of Antimicrobial Resistance Coexistence among Bacterial Pathogens Associated with Bovine Mastitis

Correlation Analysis of Antimicrobial Resistance Coexistence among Bacterial Pathogens Associated with Bovine Mastitis

Prevalence, Risk Factors, and Antimicrobial Resistance of Staphylococcus and Streptococcus Species Isolated from Subclinical Bovine Mastitis.

Subclinical mastitis (SCM) is a prevalent serious disease among dairy cows worldwide. It poses a significant challenge to the dairy industry, animal welfare, and a threat to public health. The present study aimed to investigate the molecular detection, prevalence, and antimicrobial resistance of Staphylococcus spp. and Streptococcus spp. isolated from raw composite milk samples obtained from SCM dairy cattle in Bangladesh. A total of 612 quarters milk samples obtained from 153 cows were analyzed for SCM using the California Mastitis Test. Bacterial isolation and identification were carried out and bacterial species were confirmed using molecular polymerase chain reaction methods. Antimicrobial susceptibility testing was performed using disc diffusion method. The findings revealed that the prevalence of SCM was 70.3% (26/37), 35.95% (55/153), and 23.04% (141/612) in the herd, cow, and quarter levels, respectively. Among the positive samples, 92.7% (51/55) were Staphylococcus spp. (S. aureus, S. chromogenes, and S. simulans) and the remaining isolates were 7.3% (4/55) Streptococcus spp. (Streptococcus agalactiae and Streptococcus dysgalactiae). The most prevalent species was S. chromogenes, accounting for 67.3% (37/55). Antimicrobial susceptibility testing showed that 65.5% of isolates were susceptible to cefoxitin, whereas, 89.1% were resistant to penicillin. Overall, 12 isolates (21.8%) out of 55 were resistant to more than three classes of antimicrobials and were defined as multidrug-resistant isolates. Methicillin-resistance gene was detected in 61.1% of the cefoxitin-resistant isolates. A multivariate logistic regression analysis identified five potential risk factors including the lack of post-milking teat disinfection (OR: 3.06), absence of immediate feeding after milking (OR: 9.81), poor udder hygiene (OR: 7.83), tick infestation (OR: 13.76), and absence of dry cow therapy (OR: 3.31). The findings of the current study underscore the urgent requirement for targeted interventions, considering the identified factors to effectively manage and control SCM in dairy cows.

Microbial dysbiosis in the gut–mammary axis as a mechanism for mastitis in dairy cows

Mastitis is a significant and costly disease in dairy cows, reducing milk production and affecting herd health. Recent research highlights the role of gastrointestinal microbial dysbiosis in the development of mastitis. This review focuses on how microbial imbalances in the rumen and intestines can compromise the integrity of the gastrointestinal barriers, allowing harmful bacteria and endotoxins, such as lipopolysaccharide, to enter the bloodstream and reach the mammary gland, triggering inflammation. This process links gastrointestinal health to mammary gland inflammation through the gut–mammary axis. Furthermore, disruptions in glucose metabolism and immune responses are implicated in the progression of mastitis. This review underscores the potential for non‐antibiotic interventions aimed at restoring microbial balance to reduce mastitis incidence, providing new insights into improving dairy cow health and farm productivity. Our findings emphasise the critical need to explore preventive measures targeting the rumen and intestinal microbiota for effective mastitis control.

Genetic framework sequencing analysis of Candida tropicalis in dairy cow mastitis and study of pathogenicity and drug resistance

Candida tropicalis (C. tropicalis) is a zoonotic pathogen that is widespread in the environment and in recent years an increasing number of dairy cows have been infected with the fungus causing mastitis in cows.In this study, 37 milk samples from the udders of cows with clinical mastitis were collected from a dairy farm in Guangxi Province, China, from which C. tropicalis was isolated and identified, and then the isolated fungi were subjected to genome frame map sequencing, genome functional analysis as well as comparative genome analysis of the sequencing results, and combined with the virulence test of the fungi and drug sensitivity test of the fungi determined in infected mice, the resistance genes and pathogenicity of the fungi were Analysis of resistance genes and pathogenicity.Our study results revealed the isolation and characterisation of C. tropicalis from diseased cows, with a genome length of approximately 14.27 Mb. Functional annotation of the genome identified 4068 genes associated with C. tropicalis. The strain exhibited a chemoresistance mutation in the gene cyp51,a virulence-enhancing mutation in the gene VTC4, and mutations in genes linked to drug resistance. Pathogenicity tests demonstrated that C. tropicalis could induce damage to the internal organs of mice, leading to different levels of cyanosis in the abdominal cavity, white necrotic foci on the surface of internal organs, lung hemorrhage, and enlargement of the spleen and thymus.Histological sections also revealed varying degrees of hemorrhage and degenerative changes in the cells of different organs in the mice. Drug sensitivity tests showed that the fungus was highly sensitive to nystatin and ketoconazole, moderately sensitive to amphotericin B, and insensitive to antibiotics such as itraconazole, gentamicin, and penicillin. In conclusion, C. tropicalis isolated from dairy cows in the Guangxi region in this study was pathogenic and resistant to azoles such as itraconazole and fluconazole, and this study provides a theoretical basis for the further screening of novel resistance genes in C. tropicalis, as well as providing a certain reference for the drugs used for the treatment of fungal cow mastitis in this region.

Whole genome sequence analysis of multi-drug resistant and biofilm-forming Staphylococcus haemolyticus isolated from bovine milk

BackgroundMilk is an excellent growth medium for microorganisms due to its nutritive composition. Microorganisms have been implicated in bovine mastitis (BM) in dairy cows as well as causing infections in animals and humans. Despite extensive endeavours to manage BM, this condition continues to persist as the most prevalent and economically burdensome problem affecting dairy cattle on a global scale. Non-aureus staphylococci (NAS) species such as Staphylococcus haemolyticus, S. epidermidis, and S. xylosus are currently the predominant microbiological agents identified as the main cause of subclinical udder infections and are also considered opportunistic pathogens in cases of clinical mastitis in dairy cows. Therefore, it is crucial to elucidate the genetic profile of these species. The primary objective of this study was to characterise three phenotypically determined multidrug-resistant NAS environmental strains (NWU MKU1, NWU MKU2, and NWU MKS3) obtained from dairy cows milk via whole-genome sequencing.ResultsThe results confirmed that the three isolates were S. haemolyticus with genome sizes of 2.44, 2.56, and 2.56 Mb and a G + C content of 32.8%. The genomes contained an array of antibiotic resistance genes that may potentially confer resistance to a range of antibiotic classes, such as macrolides, fluoroquinolones, aminoglycosides, cephalosporins, tetracyclines, peptides, and phenicol. Furthermore, all the genomes carried virulence genes, which are responsible for several functions, such as adhesion, enzyme and toxin production. The genomes of these organisms contained signatures encoding mobile genetic elements such as prophages and insertion sequences.ConclusionThese findings indicate there is a need for diligent monitoring with improved management practices and quality control strategies on farms to safeguard milk production systems and human health.

Characteristics of methicillin-resistant Staphylococcus aureus isolates from bovine mastitis milk in South Korea: molecular characteristics, biofilm, virulence, and antimicrobial resistance.

This study reports on the presence and characteristics of methicillin-resistant Staphylococcus aureus strains in milk from mastitis-infected cows. To our knowledge, this is the first report of a Panton-Valentine leukocidin- and toxic shock syndrome toxin-1-positive ST22-SCCmec IV strain in South Korea.

In Situ Expression of Yak IL-22 in Mammary Glands as a Treatment for Bovine Staphylococcus aureus-Induced Mastitis in Mice.

Since the development of dairy farming, bovine mastitis has been a problem plaguing the whole industry, which has led to a decrease in milk production, a reduction in dairy product quality, and an increase in costs. The use of antibiotics to treat mastitis can cause a series of problems, which can bring a series of harm to the animal itself, such as the development of bacterial resistance and dramatic changes in the gut flora. However, the in vivo and in vitro antibacterial activity of yak Interleukin-22 (IL-22) and its application in mastitis caused by Staphylococcus aureus have not been reported. In this study, the mammary gland-specific expression plasmid pLF-IL22 of the yak IL-22 gene was constructed and expressed in MAC-T cells and mammary tissue of postpartum female mice. The coding region of the IL-22 gene in yaks is 573 bp, which can encode 190 amino acids, and the homology difference in the IL-22 gene in yaks is less than 30%, which indicates certain conservation. IL-22 is a hydrophilic protein with a total positive charge of four, the presence of a signal peptide, and the absence of a transmembrane domain. Sufficient expression of IL-22 effectively inhibited the high expression of inflammatory factors caused by Staphylococcus aureus, reduced the symptoms of mammary gland histopathology, and alleviated mastitis. Under the action of IL-22, the intestinal flora of mastitis mice also changed, the abundance of intestinal Bacilli, Prevotellaceae, and Alloprevotella in mice increased after treatment, and the pathogenic bacteria decreased. These findings provide new insights into the potential application of the yak IL-22 gene in the treatment of bovine mastitis in the future.

Whole-genome sequencing of Streptococcus uberis isolated from cows with mastitis in Thuringia.

Introduction. Streptococcus uberis is a common cause of mastitis in cattle, leading to significant economic losses. The widespread use of antimicrobials has contributed to the emergence of resistance, which poses a severe challenge in controlling S. uberis infection.Aim. The objective of this study was to gain insights into the antimicrobial resistance (AMR) and epidemiological typing of S. uberis isolated from milk collected from bovine mastitis on dairy farms in Thuringia.Methodology. In this study, 84 S. uberis isolates were obtained from cattle with clinical mastitis in Thuringia, their phenotypic and genotypic AMR were analyzed and their phylogenetic relationship was explored using whole-genome sequencing.Results. Genetically heterogeneous strains were found on the farms, but clusters of highly similar strains also circulated within the same farms. All isolates were sensitive to ampicillin, penicillin, ceftiofur, and vancomycin. However, 42.9%, 42.9%, 22.6%, 19.0%, and 13.0% were resistant to tetracycline, doxycycline, clindamycin, pirlimycin, and erythromycin, respectively. Thirty-nine strains were phenotypically resistant to two or more tested antibiotics. We identified a plasmid associated with macrolide and lincosamide resistance in 12% of the strains.Conclusion. The emergence of S. uberis strains resistant to multiple antibiotics highlights the importance of S. uberis surveillance and the prudent use of antimicrobials.