Antimicrobial susceptibility and coagulase gene typing of Staphylococcus aureus isolated from bovine clinical mastitis cases in Turkey

The objectives of this study were to determine antimicrobial resistance patterns of Staphylococcus aureus strains isolated from bovine clinical mastitis cases and to subtype the strains by polymerase chain reaction (PCR) technique based on coagulase gene polymorphism. Two hundred sixty-five S. aureus isolates collected from individual animals in different herds (n = 235) from 1995 to 2004 were tested for susceptibility to penicillin, ampicillin, amoxicillin-clavulanate, oxacillin, oxytetracycline, enrofloxacin, kanamycin-cephalexin, and trimethoprim-sulfamethoxazole using the agar disc diffusion test. Strains were also tested for β-lactamase production. A total of 29.8% of the strains were susceptible to all antimicrobial agents tested. The highest resistance was observed in 63.3% of the strains against β-lactam antibiotics, penicillin and ampicillin. Oxytetracycline resistance was observed in 27.9% of the strains, either alone or in combination with β-lactams. No resistance was detected for amoxicillin-clavulanate, oxacillin, enrofloxacin and kanamycin-cephalexin. β-Lactamase production and resistance to β-lactam antibiotics were usually correlated. Resistance against β-lactams increased from 43.5% in 1995 to 58 to 77% from 1999 to 2004.One hundred twenty-five strains were examined for coagulase gene polymorphism. The isolates were subtyped into 4 types by coagulase gene-based PCR. A predominant 1000-bp PCR product was observed in 60.8% of the isolates typed. The results indicate that a few coagulase gene types of S. aureus are responsible for the majority of bovine clinical mastitis cases in one province of Central Anatolia region, Turkey.     

Short communication: antimicrobial drug susceptibility of Staphylococcus aureus from subclinical bovine mastitis in Italy

The antimicrobial susceptibility of 68 Staphylococcus aureus isolates collected during 2004 from milk of cows affected by subclinical mastitis was examined. The antimicrobial agents tested were the β-lactams, penicillin G, amoxicillin, ampicillin, cloxacillin, amoxicillin + clavulanate, cephalonium, and cefoperazone; and other drugs including lincomycin, oxytetracycline, doxycycline, and kanamycin. Minimum inhibitory concentrations recorded show that only certain β-lactamase-resistant penicillins (specifically cloxacillin) or penicillin combinations (amoxicillin + clavulanate) were consistently effective against Staph. aureus, whereas the other β-lactam derivatives and drugs from other pharmacological groups were either moderately effective or ineffective. Thus, β-lactamase-resistant penicillins are to be considered the antimicrobial agents of choice for treatment of bovine mastitis resulting from infection by Staph. aureus.     

Characterization of Staphylococcus aureus isolates recovered from bovine mastitis in Rio de Janeiro, Brazil

Phenotypic characteristics, antimicrobial susceptibility, and genetic relationships were analyzed in 107 Staphylococcus aureus isolates recovered from cows with subclinical mastitis in Southeastern Brazil. Thirteen different biochemical patterns were detected among isolates. A predominant pattern represented by about 54% of the isolates was distributed among several herds. Isolates of distinct phenotypic profiles were also detected within a herd. Susceptibility to ampicillin, cefotaxime, cephalotin, chloramphenicol, erythromycin, gentamycin, kanamycin, nitrofurantoin, norfloxacin, ofloxacin, oxacillin, penicillin, rifampin, sulfamethoxazole-trimethoprim, tetracycline, trimethoprim, and vancomycin, determined by the disk diffusion method, was observed in 44.9% of isolates. On the other hand, 55.1, 7.4, and 2.8% of the strains were resistant to ampicillin/penicillin, tetracycline, and erythromycin, respectively. Genetic diversity was analyzed by pulsed-field gel electrophoresis using SmaI as the restriction enzyme. All isolates could be typed by pulsed-field gel electrophoresis, which identified 16 types and 24 subtypes. Type A and its subtypes comprised 54.2% of all isolates and were recovered from 6 of the 9 herds analyzed. Other types and subtypes were also found in multiple herds. Although multiple types and subtypes were found within a specific herd, a predominant type was frequently observed.     

The occurrence of Staphylococcus aureus on a farm with small-scale production of raw milk cheese

In recent years, the small-scale production of raw milk products has increased in Norway, and there is some concern that such foods may pose a risk of staphylococcal food poisoning to consumers. The aim of the study was to evaluate potential sources of contamination of raw milk cheese with Staphylococcus aureus on a bovine dairy farm with small-scale production. Samples for bacteriological analyses (n = 144) were collected from the animals, the environment, processing equipments, from humans, and from cheeses at various stages of production. Staphylococcus aureus was isolated from 10 of 11 cows, the farmer, equipment, the environment, and the cheese. Seventy-five Staph. aureus isolates were genotyped by pulsed-field gel electrophoresis, tested for enterotoxin (SE) production by reversed passive latex agglutination, for SE genes by multiplex polymerase chain reaction, and for penicillin resistance by the cloverleaf method. Five different pulsotypes were identified and SE gene fragments were identified in 11 isolates, but no isolates produced SE or were penicillin resistant. Staphylococcus aureus was found throughout the farm, and appeared to be spread with the milk to the environment, equipment, and to products. One pulsotype dominated and was identified from most sample sites on the farm. Raw milk products are vulnerable to contamination with Staph. aureus. Strategies to reduce the occurrence of Staph. aureus in bulk milk are of particular importance on farms where milk is used for raw milk products.     

Phenotypic and genetic antibiogram of methicillin-resistant staphylococci isolated from bovine mastitis in Korea

Staphylococcus aureus belongs to the group of major contagious mastitis pathogens, whereas the coagulase-negative staphylococci (CNS) are also capable of causing opportunistic bovine mastitis. Many of these strains are resistant to penicillin or ampicillin because of the long-term use of β-lactam antibiotics in agricultural and healthcare settings. Based on the simple and highly specific coagulase genotyping by PCR-RFLP used for discriminating among Staph. aureus strains, the relationship between phenotypic antibiogram and the polymorphism of coagulase gene was determined in this study. The staphylococci strains (835 Staph. aureus and 763 CNS) were isolated from 3,047 bovine mastitic milk samples from 153 dairy farms in 8 provinces from 1997 to 2004 in the Republic of Korea. Twenty-one (2.5%) Staph. aureus and 19 (2.4%) CNS strains were resistant to methicillin [oxacillin minimum inhibitory concentration (MIC) ≥4 μg/mL]. The mecA gene was also found in 13 methicillin-resistant Staph. aureus (MRSA) and 12 methicillin-resistant CNS (MRCNS) isolates with a significantly higher detection rate of the mecA gene in MRSA with high MIC (≥16 μg/mL) compared with those with MIC ≤ 8 μg/mL. Methicillin-resistant Staph. aureus and MRCNS were also more resistant to other antibiotics (ampicillin, cephalothin, kanamycin, and gentamicin) than methicillin-susceptible staphylococci. Among 10 different coa PCR-RFLP patterns (A to J) in 706 Staph. aureus strains, the main types were A (26.9%), B (17.0%), G (10.5%), and H (15.4%), with the frequent observation of the A and H types (6 and 10 isolates) in MRSA. This study indicates that major epidemic Staph. aureus clones may be spread between different dairy farms, and the profile of coa genotype can be applied for epidemiological investigations and control of bovine mastitis, particularly one caused by MRSA with specific prevalent coa types.     

Bacteriological characteristics of Staphylococcus aureus isolates from humans and bulk milk

The aim of this study was to clarify the epidemiological association and bacteriological characteristics of human and animal Staphylococcus aureus isolates. Pulsed-field gel electrophoresis showed that pulsotypes (PT) of isolates from bulk milk differed from PT from human isolates, suggesting that there is no epidemiological association between isolates from these 2 sources. The absence of a common PT could result from the lack of contact between the sources. Methicillin-resistant S. aureus from human secretions and S. aureus from bulk milk in Japan consisted of 1 and 2 dominant clusters, respectively, whereas methicillin-susceptible S. aureus from humans consisted of assorted clusters. Isolates belonging to the dominant clusters showed the coagulase serotype, the capsule serotype, detection of exotoxin genes, and antimicrobial susceptibility. Isolates from bulk milk did not show the penicillin-binding protein 2a gene, and 252 of 275 isolates belonging to the 2 dominant clusters of bulk milk were susceptible to ampicillin, cefazolin, erythromycin, chloramphenicol, oxacillin, and vancomycin. Moreover, the LukM/LukF′-PV leukotoxin gene was detected in 233 of 275 isolates belonging to the dominant clusters in bulk milk isolates. These results support the hypothesis that a number of factors play a role in the adaptation of S. aureus isolates to specific hosts.     

Characterization of Staphylococcus aureus isolated from chronically infected dairy goats

A herd of 88 Alpine goats in Northern Italy was monitored for a complete lactation. Milk samples were taken from each udder half during 8 monthly visits. Goats (n = 28) with ≥2 consecutive positive tests for Staphylococcus aureus in the same udder half were identified as chronically infected, and all of those had ≥4 positive tests of the 8 samples. Goats with no infections in either udder half during any visit were considered healthy (n = 26). Linear mixed models were used to examine the relationship between chronic infection by S. aureus and SCC and production traits. The bacteria isolated from one sample from each infected goat were genotyped on the basis of polymorphism in several genes and evaluated for the presence of genes encoding for enterotoxins. The bacteria isolated from each animal were also subject to a test for β-lactamase production and to minimum inhibitory concentration tests for 11 antimicrobial agents. As expected, SCC (log₂) was significantly higher in infected goats than in healthy goats (7.55 vs. 5.50). Also, mean log SCC from infected udder halves (8.02) was greater than that in uninfected udder halves from the same goats (6.44). No significant differences were observed in milk yield or for fat and protein percentages between infected and healthy goats. No genetic variability was observed among the bacteria isolated, suggesting that all were from the same strain, although isolates did vary in susceptibility to various antimicrobial agents. All S. aureus isolates were negative for the β-lactamase production test. The most effective drugs when tested in vitro were benzylpenicillin, amoxicillin plus clavulanic acid, cloxacillin, and cephalosporins.     

Long-term persistence of specific genetic types of mastitis-causing Staphylococcus aureus on three dairies

Pulsed-field gel electrophoresis (PFGE) after SmaI digestion was used to investigate the persistence of specific genotypes of bovine mammary gland isolates of Staphylococcus aureus on 3 dairy herds. A total of 341 isolates of Staph. aureus were available from cows in 3 herds, collected over a period of 15 yr. Pulsed-field gel electrophoresis band patterns of Staph. aureus isolates were analyzed visually and with gel analysis and comparison software. Based on this analysis, isolates were classified by PFGE type. Persistence was determined as the time period from the first to the last isolation of a particular PFGE type of Staph. aureus within a herd. Specific types of mastitis-causing Staph. aureus persisted long-term on these dairies. For example, PFGE type 3 isolates persisted on farms A, B, and C for 15, 15, and 13 yr, respectively. Type 6 was found to persist for 13 yr on farm C. Despite the application of standard mastitis control practices, mastitis-causing Staph. aureus types appeared to persist long-term, as detected by PFGE, and were isolated coincident with herd problems of increased milk somatic cell counts and decreased milk production.     

Host adaptation of bovine Staphylococcus aureus seems associated with bacteriological cure after lactational antimicrobial treatment

Staphylococcus aureus causes a wide range of diseases in multiple species. Some sequence types (ST) are observed in a variety of hosts, whereas other strains are mainly associated with bovine mastitis, suggesting host adaptation. We propose that host adaptation of Staph. aureus may influence bacteriological cure of bovine subclinical mastitis after antimicrobial treatment. To test this hypothesis, multilocus sequence typing was performed on Staph. aureus isolates from 60 treated and 79 untreated control quarters that were obtained from well-defined cohorts of dairy cows from a recently conducted randomized field trial on early treatment of subclinical mastitis. Bovine-associated ST were distinguished from non-bovine-associated ST based on the literature and public databases. The association between host adaptation and bacteriological cure was investigated using population-averaged logistic regression models. Thirteen ST were identified, with approximately 80% of isolates belonging to bovine-associated ST. The odds for cure were around 2.5 times as high for non-bovine-associated ST as for bovine ST in treated quarters, whereas no difference in spontaneous cure was observed in untreated control quarters. In addition, host adaptation was related to known predictors of cure, such as penicillin susceptibility and somatic cell count. All isolates belonging to non-bovine-associated ST were resistant to penicillin, whereas the majority of isolates belonging to bovine-associated ST were penicillin susceptible. Penicillin-resistant bovine-associated strains were associated with high somatic cell counts compared with other strains. The correlation between penicillin resistance, cell counts, and host adaptation may affect the association between host adaptation and cure. For diagnostic purposes, a simple and fast alternative to multilocus sequence typing of Staph. aureus to determine host adaptation may be valuable.     

Characterization for enterotoxin production,virulence factors,and antibiotic susceptibility of Staphylococcus aureus isolates from various foods in Portugal

Staphylococcus aureus represents a public health challenge worldwide. The aim of this study was the characterization of different food isolates of S. aureus on the basis of their production of enterotoxins, hemolysins and resistance to antibiotics. A total of 148 coagulase-positive staphylococcal strains isolated from different food origins were identified to the species level. By multiplex PCR, 69% of the isolates were shown to be enterotoxigenic (SEs); the most common were sea seg, sea seg sei and seg sei. According to CLSI [CLSI, Clinical and Laboratory Standards Institute, 2007. Performance Standards for Antimicrobial Susceptibility Testing; Fifteenth Informational Supplement. CLSI document M100-S15. Clinical and Laboratory Standards Institute, Wayne, PA], 38% of the isolates were resistant to oxacillin (≥6 μg/mL; MRSA positives) but only 0.68% showed the presence of mecA gene. 70 and 73% of the S. aureus strains were resistant to β-lactams, ampicillin and penicillin, respectively. The virulence pattern was demonstrated to be origin and strain dependent. These findings emphasise the need to prevent the presence of S. aureus strains and SEs production in foods.     

Short communication: Staphylococcus aureus isolated from colostrum of dairy heifers represent a closely related group exhibiting highly homogeneous genomic and antimicrobial resistance features

In heifers, intramammary infections caused by Staphylococcus aureus affect milk production and udder health in the first and subsequent lactations, and can lead to premature culling. Not much is known about Staph. aureus isolated from heifers and it is also unclear whether or not these strains are readily transmitted between heifers and lactating herd mates. In this study, we compared phenotypic characteristics, spa types, and DNA microarray virulence and resistance gene profiles of Staph. aureus isolates obtained from colostrum samples of dairy heifers with isolates obtained from lactating cows. Our objective was to (1) characterize Staph. aureus strains associated with mastitis in heifers and (2) determine relatedness of Staph. aureus strains from heifers and lactating cows to provide data on transmission. We analyzed colostrum samples of 501 heifers and milk samples of 68 lactating cows within the same herd, isolating 48 and 9 Staph. aureus isolates, respectively. Staphylococcus aureus strains from heifers, lactating herd mates, and an unrelated collection of 78 strains from bovine mastitis milk of mature cows were compared. With 1 exception each, characterization of all strains from heifers and lactating cows in the same herd yielded highly similar phenotypic and genotypic results. The strains were Staphaurex latex agglutination test negative (Oxoid AG, Basel, Switzerland) and belonged to agr type II, CC705, and spa types tbl 2645 and t12926. They were susceptible to all antimicrobial agents tested. In contrast, the strains from mature cows in other herds were spread across different clonal complexes, spa types, and SplitsTree clusters (http://www.splitstree.org/), thus displaying a far higher degree of heterogeneity. We conclude that strains isolated from colostrum of heifers and mastitis milk of lactating cows in the same herd feature highly similar phenotypic and genomic characteristics, suggesting persistence of the organism during the first and potentially subsequent lactations or transmission between heifers and mature herd mates.     

Antibiogram and coagulase diversity in staphylococcal enterotoxin-producing Staphylococcus aureus from bovine mastitis

We investigated antibiogram and coagulase gene diversity in staphylococcal enterotoxin (StE)-producing Staphylococcus aureus isolated from raw milk samples of cows infected with mastitis from 140 dairy farms in Korea between 1997 and 2004. Of the 696 Staph. aureus isolates collected in this study, 164 isolates (23.6%) produced one or more staphylococcal enterotoxins (A to D), and 19 isolates (2.7%) were methicillin-resistant. The percentage of StE-producing Staph. aureus (SES) isolates resistant to methicillin, kanamycin, neomycin, amikacin, and tetracycline was greater than that of non-SES. Ten coagulase genotype patterns were observed, including 4 main types comprising I (25.4%), II (13.9%), VII (13.2%), and VIII (17.8%). More than 4 Staph. aureus types were isolated from each of 82 dairy farms in different geographic locations, and only 1 coagulase genotype pattern was observed in 39 of the herds (47.6%). There was no significant correlation between coagulase genotypes harbored by Staph. aureus and their specific StE type. The percentage of isolates producing major StE types (A, B, AC, and ABCD) and being resistant to cephalothin and methicillin was greater among the Staph. aureus isolates with the 4 predominant coagulase genotypes (I, II, VII, and VIII) than among the isolates harboring the 6 rare coagulase types (III, IV, V, VI, IX, and X). Based on coagulase gene polymorphisms, our data indicate that a broad distribution of identical or closely related enterotoxin-producing Staph. aureus strains seem to contribute to bovine mastitis in the Republic of Korea.     

Interference of Origanum vulgare L. essential oil on the growth and some physiological characteristics of Staphylococcus aureus strains isolated from foods

The aim of this study was to investigate the chemical composition of Origanum vulgare L. essential oil, the inhibitory effect of the oil on the cell viability of Staphylococcus aureus strains isolated from foods, and the influence of sub-inhibitory concentrations of the oil on some physiological attributes of these strains. GC-MS analysis showed that carvacrol (57.71%) was the most prevalent compound in the oil, followed by p-cymene (10.91%), γ-terpinene (7.18%), terpinen-4-ol (6.68%) and thymol (3.83%). The results showed that O. vulgare essential oil at 0.03, 0.6 and 0.12 μL mL⁻¹ inhibited the cell viability of Staph. aureus. At 0.12 μL mL⁻¹ the oil caused cidal effect with decrease ≥3 log cycles of the initial inoculum after 15 min of exposure. Sub-inhibitory concentrations (0.03 and 0.015 μL mL⁻¹) of the oil suppressed some physiological attributes of the Staph. aureus strains such as coagulase, lipase and salt tolerance. The oil interfered on the microbial metabolic activity in a dose-dependent manner. O. vulgare essential oil could be a novel antimicrobial with capability to suppress some physiological characteristics, in addition to inhibit the growth and survival of pathogen bacteria in foods, particularly Staph. aureus.     

Invited Review: The role of cow, pathogen, and treatment regimen in the therapeutic success of bovine Staphylococcus aureus mastitis

Staphylococcus aureus is an important cause of udder infections in dairy herds. Both lactational and dry cow therapy are part of Staph. aureus control programs. Reported cure rates for Staph. aureus mastitis vary considerably. The probability of cure depends on cow, pathogen, and treatment factors. Cure rates decrease with increasing age of the cow, increasing somatic cell count, increasing duration of infection, increasing bacterial colony counts in milk before treatment, and increasing number of quarters infected. Staphylococcus aureus mastitis in hind quarters has a low cure rate compared with front quarters. Antimicrobial treatment of intramammary infections with penicillin-resistant Staph. aureus strains results in a lower cure rate for treatment with either β-lactam or non-β-lactam antibiotics. Other strain-specific factors may affect the probability of cure but routine diagnostic methods for use in bacteriology laboratories or veterinary practices are not yet available. The most important treatment factor affecting cure is treatment duration. Increased duration of treatment is associated with increased chance of cure. Economically, extended treatment is not always justified, even when indirect effects of treatment such as prevention of contagious transmission are taken into consideration. Usefulness of treatment trials could be improved by standardization of case definitions, consideration of host and strain factors, and sufficient statistical power. Treatment of young animals with penicillin-sensitive Staph. aureus infections is often justified based on bacteriological cure and economic outcome, whereas treatment of older animals, chronic infections, or penicillin-resistant isolates should be discouraged.     

Analysis of antibiotic resistance patterns and detection of mecA gene in Staphylococcus aureus isolated from packaged hamburger

Presence of Staphylococcus aureus, antibiotic resistance pattern and PCR detection of mecA gene in isolated strains were investigated in total of 256 packaged hamburgers in Iran-Tehran. For this purpose we used standard disk-diffusion method and sensitive and specific PCR technique, respectively. Results showed that 25% of samples were positive for S. aureus. Resistance to meticillin, erythromycin, penicillin G, cefazolin, ciprofloxasin, vacomycin and amoxiclave was determined 89%, 20.3%, 18.7%, 15.6%, 14%, 26.6% and 12.5%, respectively. According to the obtained results from PCR analysis of methicillin-resistant S. aureus (MRSA), mecA gene was present in 100% of the resistant isolates, 0% of intermediate-resistance isolates and 25% of susceptible isolates. The results obtained from PCR detection of mecA gene showed high correlation with standard disk diffusion test.     

Short communication: profile of virulence factors of Staphylococcus aureus isolated from subclinical bovine mastitis in the state of Rio de Janeiro, Brazil

Staphylococcus aureus produces exoproteins that contribute to its ability to colonize the mammary gland such as hemolysins, coagulase, slime, and protein A. This study characterized phenotypically and genotypically these virulence factors in 50 Staph. aureus isolates. These isolates were obtained from milk samples from subclinical mastitis cases identified in 15 dairy cattle farms located in the state of Rio de Janeiro, Brazil. All of the confirmed Staph. aureus samples were PCR positive for the coa gene, which displayed 3 different size polymorphisms. The amplification of the spaA X region yielded a single amplicon for each isolate with the prevalent amplicon sized 315 bp. The Staph. aureus isolates were 24 and 16% positive for the hla and hlb genes, respectively, and 22 and 20% positive for the icaA and icaD genes, respectively. Amplification of the agr gene RNAIII was positive in 74% of the strains. Twenty-seven different profiles were identified among the samples, indicating a great diversity of Staph. aureus involved in the etiology of mastitis cases in the analyzed region. These findings are valuable to the comprehension of the distribution of the profiles of Staph. aureus strains isolated from subclinical mastitis cases in the state of Rio de Janeiro.     

Short communication: Fluoroquinolone susceptibility of Staphylococcus aureus strains isolated from caprine clinical mastitis in southeast Spain

The antibiotic susceptibility of 32 strains of Staphylococcus aureus isolated from the mastitic milk of dairy goats was evaluated. The antibiotics tested were 3 fluoroquinolones that have been developed especially for use in veterinary medicine: danofloxacin, marbofloxacin, and orbifloxacin. Minimum inhibitory concentration (MIC) tests were performed according to the microdilution broth method. The MIC₉₀ (concentration at which 90% inhibition is achieved) values obtained were 0.5, 1, and 1 mg/L for danofloxacin, marbofloxacin, and orbifloxacin, respectively. Danofloxacin was the most active fluoroquinolone tested against Staph. aureus strains isolated from milk; however, specific testing is required before using these drugs as therapy for the control of clinical mammary infections in goats.     

Prevalence,antimicrobial susceptibility,and molecular characterization of Staphylococcus aureus isolated from dairy herds in northern China

Staphylococcus aureus is one of the main pathogens involved in dairy cow mastitis. Monitoring of antibiotic use would prove useful to assess the risk of Staph. aureus in raw milk. The objective of this work was to investigate the prevalence of Staph. aureus strais isolated from raw milk in northern China, and to characterize antimicrobial susceptibility of these strains and their key virulence genes. In total, 195 raw milk samples were collected from 195 dairy farms located in 4 cities of northern China from May to September 2015. Out of 195 samples, 54 (27.7%) were positive for Staph. aureus. Among these 54 samples, 54 strains of Staph. aureus were isolated, and 16 strains were identified as methicillin-resistant Staph. aureus. The strains exhibited high percentages of resistance to penicillin G (85.2%), ampicillin (79.6%), and erythromycin (46.3%). Moreover, 72% of the strains showed resistance to more than one antimicrobial agent. Overall, 63% of penicillin-resistant strains possessed the blaZ gene, and 60% of the erythromycin-resistant strains possessed erm(A), erm(B), erm(C), msr(A), or msr(B) genes with 8 different gene patterns. All isolates resistant to gentamicin, kanamycin, and oxacillin carried the aac6'-aph2”, ant(4')-Ia, and mecA genes, respectively. Two tet(M)-positive isolates carried specific genes of the Tn916-Tn1545 transposon. The most predominant virulence genes were sec, sea, and pvl, which encode staphylococcal enterotoxins (sec and sea) and Panton-Valentine leukocidin, respectively. Thirty-two isolates (59.2%) harbored one or more virulence genes. The majority of Staph. aureus strains were multidrug resistant and carried multiple virulence genes, which may pose a risk to public health. Our data indicated that antimicrobial resistance of Staph. aureus was prevalent in dairy herds in northern China, and that antibiotics, especially penicillin G and ampicillin, to treat mastitis caused by Staph. aureus should be used with caution in northern China.     

High-intensity pulsed electric field variables affecting Staphylococcus aureus inoculated in milk

Staphylococcus aureus is an important milk-related pathogen that is inactivated by high-intensity pulsed electric fields (HIPEF). In this study, inactivation of Staph. aureus suspended in milk by HIPEF was studied using a response surface methodology, in which electric field intensity, pulse number, pulse width, pulse polarity, and the fat content of milk were the controlled variables. It was found that the fat content of milk did not significantly affect the microbial inactivation of Staph. aureus. A maximum value of 4.5 log reductions was obtained by applying 150 bipolar pulses of 8 μs each at 35 kV/cm. Bipolar pulses were more effective than those applied in the monopolar mode. An increase in electric field intensity, pulse number, or pulse width resulted in a drop in the survival fraction of Staph. aureus. Pulse widths close to 6.7 μs lead to greater microbial death with a minimum number of applied pulses. At a constant treatment time, a greater number of shorter pulses achieved better inactivation than those treatments performed at a lower number of longer pulses. The combined action of pulse number and electric field intensity followed a similar pattern, indicating that the same fraction of microbial death can be reached with different combinations of the variables. The behavior and relationship among the electrical variables suggest that the energy input of HIPEF processing might be optimized without decreasing the microbial death.     

Antimicrobial susceptibilities in dairy herds that differ in dry cow therapy usage

Intramammary infusion of antimicrobials at the end of lactation (dry cow therapy; DCT) is a central part of mastitis control programs and is one of the major indications for antimicrobial use in dairy cows. However, with increasing focus on prudent use of antimicrobials and concerns about emergence of antimicrobial resistance, the practice of treating every cow at the end of lactation with DCT is in question. This cross-sectional, observational study determined the minimum inhibitory concentrations (MIC) of 10 antimicrobials for coagulase-negative staphylococci (CNS), Staphylococcus aureus, Streptococcus dysgalactiae, and Streptococcus uberis isolates from milk samples from dairy cows with somatic cell counts >200,000 cells/mL in herds that had been organic for >3 yr (n = 7), or had used either ampicillin-cloxacillin DCT (n = 11) or cephalonium DCT (n = 8) in the preceding 3 yr. The organic herds were certified under the United States Department of Agriculture National Organic Program, meaning that there was no blanket DCT, and minimal use of antimicrobials in general, with a loss of organic status of the animal if treated with antimicrobials. Breakpoints (where available) were used to categorize isolates as resistant, intermediate, or susceptible to antimicrobials. The MIC distributions of isolates from different herd types were compared using binomial or multinomial logistic regression. Of 240 CNS isolates, 12.9, 0.8, 7.1, 32.6, and 1.2%, were intermediate or resistant to ampicillin, cephalothin, erythromycin, penicillin, and tetracycline, respectively. Of 320 Staph. aureus isolates, 29.0, 2.5, 1.2, and 34.9% were intermediately resistant or resistant to ampicillin, penicillin, erythromycin, and oxacillin, respectively. Of 184 Strep. uberis isolates, 1.1, 25.0, 1.6, and 1.6% were intermediately resistant or resistant to erythromycin, penicillin, pirlimycin, and tetracycline, respectively. Generally, the MIC of CNS and streptococcal isolates from organic herds were lower than isolates from herds using DCT. However, the differences in MIC distributions occurred at MIC below clinical breakpoints, so that the bacteriological cure rates may not differ between isolates of differing MIC. Bimodal distributions of MIC for ampicillin and penicillin were found in Staph. aureus isolates from organic herds, suggesting that isolates with a higher MIC are a natural part of the bacterial population of the bovine mammary gland, or that isolates with higher MIC have persisted within these organic herds from a time when antimicrobials had been used. Given these observations, further work is required to determine if exposure to DCT is causally associated with the risk of elevated MIC, and whether reduction or removal of DCT from herds would reduce the risk of elevated MIC of mastitis pathogens.