Resistance to gentamicin and amikacin of gram-negative organisms isolated from horses

Resistance of gram-negative bacteria to gentamicin has become an increasingly common problem among clinical isolates from human beings. Susceptibility of isolates from horses to gentamicin and amikacin was evaluated for the period from July, 1983 to June, 1985. All isolates of Escherichia coli, and species of Enterobacter, Klebsiella, Proteus, and Pseudomonas examined were susceptible to amikacin, except 2 of the 46 Pseudomonas isolates. In contrast, 13 to 50% of isolates were resistant to gentamicin. Escherichia coli, and Klebsiella, Proteus, and Enterobacter species isolates were highly significantly more susceptible to amikacin (P less than 0.01) than to gentamicin. Pseudomonas spp (P = 0.13) were not significantly different in susceptibility to the 2 drugs. There was significant variation among genera in their susceptibility to gentamicin (P = 0.002), primarily because of the frequency of resistance in isolates of Klebsiella spp and Proteus spp, compared with the other 3 organisms (E coli, Enterobacter spp, and Pseudomonas spp). There was no significant difference of susceptibility to amikacin among the genera studied (P = 0.06).     

Current data on antibiotic resistance of the most important bovine mastitis pathogens in Switzerland

100 strains of Staphylococcus aureus (S. aureus), 100 strains of coagulase-negative staphylococci (CNS), 100 strains of Streptococcus spp. and 100 strains of Escherichia coli (E. coli), isolated from bovine mastitis milk samples between November 2002 and April 2003, were tested for their sensitivity to various antibiotics by means of the agar diffusion method. The antibiotics were chosen on the basis of their licenses for intramammary application in Switzerland (www.vetpharm.unizh.ch). 91% of the S. aureus strains were sensitive to all the antibiotics tested. Only 9% of the strains were resistant to Penicillin G and 7% to Ampicillin. 53% of the CNS strains were sensitive to all the antibiotics tested. 31% exhibited resistance to Penicillin G, 26% to Ampicillin, 16% to Cloxacillin and 14% to Lincomycin. 30% of the Streptococcus spp. strains were sensitive to all the antibiotics tested. 4% were resistant to Penicillin G, 4% to Amoxicillin/clavulanic acid, 1% to Cefoperazone, 2% to Cefquinome, 35% to Neomycin, 22% to Gentamicin, 61% to Kanamy-cin and 11% to Lincomycin. 43% of the strains showed multiple resistance. 79% of the E. coli strains were sensitive to all the antibiotics tested. 20% exhibited resistance to Ampicillin, 9% to Neomycin and 10% to Kanamycin. A comparison of the own results with data of other authors in Switzerland shows no important changes in the resistance situation during the last 20 years. With the exception of two strains (Streptococcus spp.), all tested isolates were sensible against Cefquinome.     

Evaluation of pyelonephritis in culled indoor and outdoor high parity sows

The present trial was conducted in Hungary in neighboring large indoor and outdoor pig production units, belonging to the same breeding company. Rejected kidneys from 201 (out of 241; 83.4%) outdoor, and 191 (out of 512, 37.3%) indoor high parity sows, with previous history of recidiving postparturient fever and excessive postparturient vulvovaginal discharge were gross pathologically bacteriologically, and histologically evaluated. All rejected kidneys revealed chronic pyelonephritis. In outdoor sows Escherichia (E.) coli and Actinobaculum (A.) suis were cultured from all kidneys. Besides E. coli and A. suis, Clostridium spp., Arcanobacterium pyogenes, gram-positive streptococci (enterococci, Streptococcus faecalis), staphylococci (Staphylococcus (S.) albus, S. epidermis, S. aureus), Erysipelothrix rhusiopathiae, and Klebsiella spp. were concurrently found in 131 (64.7%) kidneys; Pseudomonas aeruginosa, Citrobacter, Pasteurella multocida, Proteus spp. were concurrently found beside E. coli and A. suis in 71 (35.3%) kidneys. In indoor sows E. coli and A. suis were cultured from all kidneys as well. Pseudomonas aeruginosa, Citrobacter, Pasteurella multocida, Proteus spp. were found beside E. coli and A. suis in 21 (11%) kidneys. However only 6 sows (3.1%) revealed the concurrent presence of Clostridium spp., Arcanobacterium pyogenes, gram-positive streptococci (enterococci, Streptococcus faecalis), staphylococci (S. albus, S. epidermis, S. aureus), Erysipelothrix rhusiopathiae, and Klebsiella spp. Implications: in Eastern European climate, more high parity outdoor sows with recidiving postparturient fever and vulvovaginal discharge have pyelonephritis and higher diversity of pathogenic bacteria in the renal pelvis compared with indoor sows.     

Antimicrobial resistance of bacteria isolated from dairy cow milk samples submitted for bacterial culture: 8,905 samples (1994-2001)

OBJECTIVE: To determine whether antimicrobial resistance patterns of major mastitis pathogens isolated from milk samples from dairy cows have changed over time. DESIGN: Retrospective study. SAMPLE POPULATION: 8905 bacterial isolates obtained from milk samples submitted to the Wisconsin Veterinary Diagnostic Laboratory between January 1994 and June 2001. PROCEDURE: Antimicrobial susceptibility was determined by means of the Kirby-Bauer disk diffusion method. Logistic regression was used to determine whether percentages of isolates resistant to various antimicrobials changed over time. RESULTS: For the gram-positive mastitis pathogens, percentages of isolates resistant to various flactam antimicrobials did not increase over the course of the study. Percentage of Staphylococcus aureus isolates resistant to penicillin decreased from 49 to 30%; percentage of Streptococcus isolates resistant to penicillin decreased from 6 to 1%. Percentage of isolates resistant to erythromycin increased for S aureus, Escherichia coli, Enterobacter spp, Enterococcus spp, and Pasteurella spp. Percentage of isolates resistant to lincomycin increased for S aureus and Staphylococcus spp. Percentage of coagulase-negative Staphylococcus isolates resistant to pirlimycin increased from 6 to 19%. For several pathogens, percentages of isolates resistant to sulfisoxazole and to trimethoprim-sulfamethoxazole decreased. No pathogens had a significant increase in the percentage of isolates resistant to novobiocin-penicillin. CONCLUSIONS AND CLINICAL RELEVANCE: Results did not indicate a trend toward increased antimicrobial resistance among mastitis pathogens isolated from milk samples from dairy cows between 1994 and 2001. Reduced resistance to flactam antimicrobials was identified for several gram-positive mastitis pathogens.     

Rapid identification of bovine mastitis pathogens by high-resolution melt analysis of 16S rDNA sequences

Accurate identification of mastitis pathogens is often compromised when using conventional culture-based methods. Here, we report a novel, rapid assay tested for speciation of bacterial mastitis pathogens using high-resolution melt analysis (HRMA) of 16S rDNA sequences. Real-time PCR amplification of 16S rRNA gene fragment, spanning the variable region V5 and V6 was performed with a resulting amplicon of 290bp. First, a library was generated of melt curves of 9 common pathogens that are implicated in bovine mastitis. Six of the isolates, Escherichia coli, Streptococcus agalactiae, Klebsiella pneumoniae, Streptococcus uberis, Staphylococcus aureus and Mycoplasma bovis, were type strains while the other 3, Arcanobacterium pyogenes, Corynebacterium bovis and Streptococcus dysgalactiae, were bovine mastitis field isolates. Four of the type strains, E. coli, S. agalactiae, K. pneumoniae and S. aureus, were found to be of human origin, while the other 3 type strains were isolated from bovine infections. Secondly, the melt curves and corresponding amplicon sequences of A. pyogenes, E. coli, S. agalactiae, S. dysgalactiae, K. pneumoniae, S. uberis and S. aureus were compared with 10 bovine mastitis field isolates of each pathogen. Based on the distinct differences in melt curves and sequences between human and bovine isolates of E. coli and K. pneumoniae, it was deemed necessary to select a set of bovine strains for these pathogens to be used as reference strains in the HRMA. Next, the HRMA was validated by three interpreters analyzing the differential clustering pattern of melt curves of 60 bacterial cultures obtained from mastitis milk samples. The three test interpreters were blinded to the culture and sequencing results of the isolates. Overall accuracy of the validation assay was 95% as there was difficulty in identifying the streptococci due to heterogeneity observed in the PCR amplicons of S. uberis. The present study revealed that broad-range real-time PCR with HRMA can be used as a powerful, fast and low-cost tool for the differentiation of clinically important bacterial mastitis pathogens.     

奶牛临床型乳房炎的细菌分离鉴定与耐药性分析

2011年山西省多个奶牛场发生了较严重的乳房炎,对76份采集的奶样进行细菌分离鉴定并采用药敏纸片法检测主要分离菌的抗生素耐药情况。所分离细菌以革兰氏阳性菌为主,革兰氏阳性球菌和其他革兰氏阳性菌分别占60.67%和23.59%。分离出多种病原菌和机会致病菌,主要的病原菌有链球菌、金黄色葡萄球菌、大肠杆菌等,检出率为2.24%~11.24%,其中无乳链球菌的检出率最高;机会致病菌有粪链球菌、微球菌、克雷伯菌、凝固酶阴性葡萄球菌等,检出率为1.12%~11.24%,其中粪链球菌和微球菌的检出率较大,分别为11.24%和6.74%。药敏试验检测结果显示,在所选的15种药物中,主要分离菌均对丁胺卡那霉素、氟哌酸和恩诺沙星3种药物敏感;大肠杆菌、克雷伯菌、凝固酶阴性葡萄球菌对青霉素类和β-内酰胺/β-内酰胺酶抑制剂类药物产生了极强的耐药性,耐药率均为100%;乳房链球菌对该类药物也产生不同程度的耐药,耐药率为20%~100%;对链霉素产生100%耐药的细菌有大肠杆菌、克雷伯菌、无乳链球菌、乳房链球菌、停乳链球菌等细菌;部分分离菌对卡那霉素、庆大霉素、链霉素、四环素、红霉素、先锋霉素Ⅴ、复方新诺明等药物产生不同程度耐药。被检奶牛场混合感染较为严重,应进一步加强环境卫生管理,临床治疗应合理有效用药。     

A retrospective study of the aetiology and temporal distribution of bovine clinical mastitis in smallholder dairy herds in the Dar es Salaam region of Tanzania

A 31-year record-based retrospective study was carried out to determine the aetiology and temporal distribution of bovine clinical mastitis in smallholder dairy herds in the Dar es Salaam region of Tanzania over the period November 1971-December 2002. Laboratory information on 1964 quarter samples from 1365 cows in 281 smallholder dairy herds were retrieved, compiled and studied. Eighty-eight percent of the quarter samples were culture-positive and the predominant mastitis pathogens isolated were Staphylococcus aureus (25.7%), Streptococcus agalactiae (15.4%), Klebsiella pneumoniae (14.3%) and Escherichia coli (14.1%). Other isolates included Pseudomonas aeruginosa (7.5%), Streptococcus dysgalactiae (5.2%) and Streptococcus uberis (4.2%). Contagious mastitis pathogens were isolated from 45.6% of the culture-positive samples, whereas environmental and miscellaneous pathogens were isolated from 48.2% and 5.7%, respectively. Thirty percent of the miscellaneous mastitis pathogens were Candida species. The results demonstrate a steady increase in clinical Candida albicans mastitis. The prevalence of Candida albicans has increased from 1% in 1971 to 17.0% in November 2002. Conversely, despite some fluctuations, the prevalence of Staphylococcus aureus, Streptococcus agalactiae, E. coli and K. pneumoniae remain above 10%. The possible risk factors for these observations are discussed.     

Identification of Streptococcus spp. causing bovine mastitis by PCR-RFLP of 16S-23S ribosomal DNA

Traditional microbiological methods for identification of Streptococcus spp. causing bovine mastitis have been demonstrated to be less than highly reliable. PCR-RFLP analysis of 16S-23S ribosomal DNA was used to characterise seven reference strains of streptococcal mastitis pathogens as well as four reference strains of other gram-positive, catalase-negative cocci of bovine origin to allow comparative identification of field isolates. RFLP analysis of PCR products, using a combination of two restriction endonucleases in single reactions (HaeIII and AluI, HaeIII and RsaI or AluI and RsaI) generated unique patterns for species of Streptococcus, Enterococcus and Lactococcus. One hundred field isolates of Streptococcus spp. collected from cows with clinical or subclinical mastitis were tested. Fifty-seven isolates, classified by conventional tests as S. uberis, were identified as 47 S. uberis and six S. parauberis by their unique RFLP patterns. The remaining four isolates had RFLP patterns distinct from the reference strains and two of these were identified as closely related to S. iniae and two to Aerococcus viridans by 16S rRNA sequencing. Conventional identification of 17 S. agalactiae and 26 S. dysgalactiae subsp. dysgalactiae was confirmed by RFLP. Use of a combination of restriction enzymes in a single tube enabled the rapid, accurate, cost effective and easily performed identification of all major streptococcal mastitis pathogens.     

Intramammary infections in a dairy herd with a low incidence of Streptococcus agalactiae and Staphylococcus aureus infections.

In a dairy herd with a low incidence of intrammary infections due to Streptococcus agalactiae and Staphylococcus aureus, clinical mastitis remained a serious problem despite good control of nonclinical mastitis through postmilking teat disinfection and antibiotic therapy of known infected quarters at the end of lactation. During the 2-year study, the incidence of clinical mastitis was 0.88 cases/cow-year; 32.2% were caused by streptococcal species other than Str agalactiae and 33.5% by gram-negative organisms. Among all new infections detected, 54.1% were caused by streptococcal species other than Str agalactiae and 25.7% by gram-negative bacteria. Among new infections, 41.6% occurred during the nonlactating period or within a few days of calving. Incidence of clinical mastitis was highest in the 1st month of lactation. Among 84 gram-negative infections, 42.8% were caused by Klebsiella pneumoniae, 20.2% by Escherichia coli, and 23.8% by Enterobacter spp. Among the many serotypes of K pneumoniae and E coli, none was predominant.     

Bacterial isolates from blood and their susceptibility patterns in critically ill foals: 543 cases (1991-1998)

OBJECTIVE: To assess microorganisms isolated from blood specimens obtained from critically ill neonatal foals and to evaluate their antimicrobial susceptibility patterns. DESIGN: Retrospective study. ANIMALS: 543 neonatal foals. PROCEDURE: Medical records of foals that were < 1 month old and were admitted to a referral neonatal intensive care unit were reviewed for results of bacteriologic culture of blood and antimicrobial susceptibility patterns. RESULTS: At least 1 microorganism was isolated from 155 of 543 (28.5%) foals. Escherichia coli was the most commonly isolated bacterium. A single gram-positive organism was detected in 49 foals. Although 90% of the E coli isolates were susceptible to amikacin, some gram-negative and gram-positive organisms had resistance against multiple antimicrobials. CONCLUSIONS AND CLINICAL RELEVANCE: Gram-negative bacteria remain the most common isolates from blood of neonatal foals; however, gram-positive organisms were also found, and with greater prevalence than reported elsewhere. Susceptibility patterns may vary, and resistance to multiple antimicrobials may develop. This is especially true for organisms such as Enterobacter spp and Enterococcus spp. Prudent empirical treatment for neonatal sepsis should include broad-spectrum antimicrobials.     

Staphylococcus aureus lipotechoic acid induces differential expression of bovine serum amyloid A3 (SAA3) by mammary epithelial cells: Implications for early diagnosis of mastitis

Mastitis is one of the most costly diseases of agriculturally important animals and is a common problem for lactating cows. Current methods used to detect clinical and especially subclinical mastitis are either inadequate or problematic. Pathogens such as the gram-positive bacterium Staphylococcus aureus or the gram-negative bacterium Escherichia coli typically cause mastitis. E. coli induces clinical mastitis, whereas, S. aureus causes a subclinical, chronic infection of the mammary gland. In this study we report the differential expression and secretion of mammary-derived serum amyloid A3 (SAA3) by bovine mammary epithelial cells following stimulation with the S. aureus cell wall component, lipotechoic acid (LTA). Two-dimensional immunoblot analyses confirmed that bovine SAA3 is the predominant SAA isoform produced by LTA stimulated mammary epithelial cells. Our previous study showed that bovine SAA3 is also differentially expressed in response to the gram-negative bacterial endotoxin lipopolysaccharide. Collectively, these data indicate that the local production of SAA3 by mammary epithelial cells in response to either gram-positive or gram-negative bacterial components may provide a sensitive indicator for early detection and treatment of mastitis in vivo, minimizing chronic cases of infection, the spread of mastitis to other animals, and economic losses.     

Bacterial flora of the conjunctiva and nasal cavity in normal and diseased captive bustards.

A survey was carried out to describe the normal aerobic bacterial flora of the conjunctiva and nasal cavity of captive houbara bustards (Chlamydotis undulata), kori bustards (Ardeotis kori), and white-bellied bustards (Eupodotis senegalensis) maintained at the National Avian Research Center, Abu Dhabi, United Arab Emirates. A total of 58 samples were examined from the nasal cavity and 55 samples from the conjunctiva of healthy bustards. There was no bacterial growth in 45% of conjunctival samples. Bacteria isolated from the conjunctiva of healthy birds included Micrococcus spp., Staphylococcus auricularis, Staphylococcus xylosus, Staphylococcus capitis, Staphylococcus warneri, Bacillus spp., and Enterobacter amigenus. Bacteria isolated from the nasal cavity of healthy birds included Bacillus spp., Micrococcus spp., S. auricularis, S. xylosus, Staphylococcus simulans, Staphylococcus saprophyticus, Staphylococcus hyicus, Staphylococcus cohnii, Staphylococcus sciuri, Aerococcus spp., and Providencia rettgeri. These findings were compared with bacterial isolates from bustards with clinical signs of ocular or upper respiratory tract diseases. Mycoplasma spp., Pseudomonas aeruginosa, Pseudomonas stutzeri, Proteus mirabilis, Escherichia coli, Klebsiella spp., Aeromonas hydrophila, and Staphylococcus aureus were the pathogenic bacteria isolated from the conjunctiva of 34.3% bustards with ocular discharges. Mycoplasma spp., P. aeruginosa, Pseudomonas spp., P. mirabilis, E. coli, Klebsiella pneumoniae, and S. aureus were the pathogenic bacteria isolated from the nasal cavity of 74% bustards with upper respiratory tract diseases.     

Microbial pathogens from goat mastitis and phage-typing of Staphylococcus aureus isolates

Examination of milk from goats yielded 41 strains from 40 clinically affected halves; 15 were Staphylococcus aureus, 6 Staph. epidermis, 1 Streptococcus agalactiae, 2 Strept. dysgalactiae, 5 Strept. uberis, 2 Corynebacterium pyogenes, 3 Escherichia coli, 3 Pasteurella spp. and 4 Mycoplasma spp. One half had dual infection of Staph. aureus and Strept. dysgalactiae. Twenty two of the 297 milk samples from apparently normal halves also harboured pathogens comprising of 9 Staph. aureus, 1 Strept. agalactiae, 2 E. coli, 2 Pasteurella spp., 2 Candida albicans and 6 Mycoplasma spp. Most of the bacterial isolates were sensitive to many broad spectrum antibiotics. Twenty of the 24 Staph. aureus isolates were phase typable by a set of 23 human Staphylococcal International Phages suggesting the utility of these phages for the typing of goat strains. The isolates were grouped into 15 phage-types, many of which have been reported from human infections in Iraq. This indicates the possibility of association of human strains of Staph. aureus in caprine mastitis. No definite correlation could be noted between antibiogram and phage types of Staph. aureus strains.     

Etiology and antimicrobial susceptibility of udder pathogens from cases of subclinical mastitis in dairy cows in Sweden

Background

A nationwide survey on the microbial etiology of cases of subclinical mastitis in dairy cows was carried out on dairy farms in Sweden. The aim was to investigate the microbial panorama and the occurrence of antimicrobial resistance. Moreover, differences between newly infected cows and chronically infected cows were investigated.

Methods

In total, 583 quarter milk samples were collected from 583 dairy cows at 226 dairy farms from February 2008 to February 2009. The quarter milk samples were bacteriological investigated and scored using the California Mastitis Test. Staphylococci were tested for betalactamase production and presence of resistance was evaluated in all specific udder pathogens. Differences between newly infected cows and chronically infected cows were statistically investigated using logistic regression analysis.

Results

The most common isolates of 590 bacteriological diagnoses were Staphylococcus (S) aureus (19%) and coagulase-negative staphylococci (CNS; 16%) followed by Streptococcus (Str) dysgalactiae (9%), Str. uberis (8%), Escherichia (E.) coli (2.9%), and Streptococcus spp. (1.9%). Samples with no growth or contamination constituted 22% and 18% of the diagnoses, respectively. The distribution of the most commonly isolated bacteria considering only bacteriological positive samples were: S. aureus - 31%, CNS - 27%, Str. dysgalactiae - 15%, Str. uberis - 14%, E. coli - 4.8%, and Streptococcus spp. - 3.1%. There was an increased risk of finding S. aureus, Str. uberis or Str. dysgalactiae in milk samples from chronically infected cows compared to findings in milk samples from newly infected cows. Four percent of the S. aureus isolates and 35% of the CNS isolates were resistant to penicillin G. Overall, resistance to other antimicrobials than penicillin G was uncommon.

Conclusions

Staphylococcus aureus and CNS were the most frequently isolated pathogens and resistance to antimicrobials was rare.     

张家口地区奶牛临床型乳腺炎病原菌的分离鉴定

采集具有临床型乳腺炎奶牛的奶样,经细菌的分离培养、纯培养和生化试验,以确定奶牛临床型乳腺炎的致病菌种类。结果发现患临床型乳腺炎奶牛的致病菌和占分离菌株的百分率为:腐生葡萄球菌19.7%、金黄色葡萄球菌16.5%、兽疫链球菌17.3%、停乳链球菌9.4%、乳房链球菌3.9%、无乳链球菌1.6%、产气肠杆菌17.3%、聚团肠杆菌4.7%、奇异变形杆菌3.0%、大肠杆菌3.0%、蜡样芽胞杆菌3.0%,样品中单纯感染与混合感染的百分率分别为66.0%和27.8%。     

Development of resistant bacteria isolated from dogs with otitis externa or urinary tract infections after exposure to enrofloxacin in vitro

Minimum inhibitory concentrations for enrofloxacin were determined for 63 bacterial isolates from dogs with otitis externa or urinary tract infections. Development of resistant mutants was determined after exposing the isolates to enrofloxacin in vitro for up to five serial passages. Results indicated that Pseudomonas aeruginosa and Enterococcus spp isolates exposed to enrofloxacin developed resistance rapidly, whereas Klebsiella, Proteus, and Streptococcus spp were less likely to develop resistance. Despite the presence of enrofloxacin pressure, no resistant bacteria developed in the Escherichia coli and staphylococcal isolates. In many isolates, susceptibility patterns changed from susceptible to intermediate.     

Survey of marbofloxacin susceptibility of bacteria isolated from cattle with respiratory disease and mastitis in Europe

A monitoring programme conducted in Europe since 1994 to survey the marbofloxacin susceptibility of bacterial pathogens isolated from cattle has established the susceptibility of bacterial strains isolated before any antibiotic treatment from bovine mastitis and bovine respiratory disease (BRD) cases between 2002 and 2008. Minimum inhibitory concentration (MIC) was determined by a standardised microdilution technique. For respiratory pathogens, Pasteurella multocida and Mannheimia haemolytica isolates (751 and 514 strains, respectively) were highly susceptible to marbofloxacin (MIC≤0.03 μg/ml for 77.39 per cent of the strains) and only 1.75 per cent of M haemolytica strains were resistant (MIC≥4 μg/ml). Histophilus somni isolates (73 strains) were highly susceptible to marbofloxacin (0.008 to 0.06 μg/ml). Mycoplasma bovis MIC (171 strains) ranged from 0.5 to 4 μg/ml. For mastitis pathogens, the majority of Escherichia coli isolates were highly susceptible to marbofloxacin (95.8 per cent of 617 strains). Staphylococcus aureus and coagulase-negative staphylococci (568 and 280 strains) had a homogenous population with MIC centred on 0.25 μg/ml. Streptococcus uberis and Streptococcus dysgalactiae (660 and 217 strains) were moderately susceptible with MIC centred on 1 μg/ml. Marbofloxacin MIC for these various pathogens appeared stable over the seven years of the monitoring programme and was similar to previously published MIC results.     

Udder pathogens and their resistance to antimicrobial agents in dairy cows in Estonia

Background

The goal of this study was to estimate the distribution of udder pathogens and their antibiotic resistance in Estonia during the years 2007-2009.

Methods

The bacteriological findings reported in this study originate from quarter milk samples collected from cows on Estonian dairy farms that had clinical or subclinical mastitis. The samples were submitted by local veterinarians to the Estonian Veterinary and Food Laboratory during 2007-2009. Milk samples were examined by conventional bacteriology. In vitro antimicrobial susceptibility testing was performed with the disc diffusion test. Logistic regression with a random herd effect to control for clustering was used for statistical analysis.

Results

During the study period, 3058 clinical mastitis samples from 190 farms and 5146 subclinical mastitis samples from 274 farms were investigated. Positive results were found in 57% of the samples (4680 out of 8204), and the proportion did not differ according to year (p > 0.05). The proportion of bacteriologically negative samples was 22.3% and that of mixed growth was 20.6%. Streptococcus uberis (Str. uberis) was the bacterium isolated most frequently (18.4%) from cases of clinical mastitis, followed by Escherichia coli (E. coli) (15.9%) and Streptococcus agalactiae (Str. agalactiae) (11.9%). The bacteria that caused subclinical mastitis were mainly Staphylococcus aureus (S. aureus) (20%) and coagulase-negative staphylococci (CNS) (15.4%). The probability of isolating S. aureus from milk samples was significantly higher on farms that had fewer than 30 cows, when compared with farms that had more than 100 cows (p < 0.005). A significantly higher risk of Str. agalactiae infection was found on farms with more than 600 cows (p = 0.034) compared with smaller farms. The proportion of S. aureus and CNS isolates that were resistant to penicillin was 61.4% and 38.5%, respectively. Among the E. coli isolates, ampicillin, streptomycin and tetracycline resistance were observed in 24.3%, 15.6% and 13.5%, respectively.

Conclusions

This study showed that the main pathogens associated with clinical mastitis were Str. uberis and E. coli. Subclinical mastitis was caused mainly by S. aureus and CNS. The number of S. aureus and Str. agalactiae isolates depended on herd size. Antimicrobial resistance was highly prevalent, especially penicillin resistance in S. aureus and CNS.     

In vitro activity of difloxacin against canine bacterial isolates.

The in vitro activity of difloxacin against canine bacterial isolates from clinical cases was studied in the United States and The Netherlands. Minimal inhibitory concentrations (MIC), the postantibiotic effect, the effect of pH on antimicrobial activity, and the bacterial killing rate tests were determined according to standard techniques. The MICs of American and Dutch isolates agreed in general. The MICs of the American gram-negative isolates ranged from 0.06 to 2.0 microg/ml, and the MICs of the Dutch gram-negative isolates ranged from 0.016 to 8.0 microg/ml. A few European strains of Proteus mirabilis and Klebsiella pneumoniae had relatively high MICs. Bordetella bronchiseptica also was less susceptible to difloxacin. The MICs of the American gram-positive cocci ranged from 0.125 to 4.0 microg/ml, and the MICs of Dutch isolates ranged from 0.125 to 2.0 microg/ ml. Difloxacin induced a concentration-dependent postantibiotic effect that lasted 0.2-3 hours in cultures with Escherichia coli, Staphylococcus intermedius, Streptococcus canis, Proteus spp., and Klebsiella pneumoniae. There was no postantibiotic effect observed against canine Pseudomonas aeruginosa. Decreasing the pH of the medium increased the MIC of Proteus mirabilis for difloxacin. The MICs of Escherichia coli and Klebsiella pneumoniae were lowest at neutral pH and were slightly increased in acid or alkaline media. At a neutral pH, most tested bacterial species were killed at a difloxacin concentration of 4 times the MIC. Similar results were obtained when these same bacteria were tested against enrofloxacin. A Klebsiella pneumoniae strain in an acidic environment was readily killed at difloxacin or enrofloxacin MIC, but at neutral pH the drug concentration had to be raised to 4 times the MIC for a bactericidal effect. After 24 hours of incubation at pH 7.1, difloxacin and enrofloxacin had similar bactericidal activity for all bacteria tested except Staphylococcus intermedius. Against S. intermedius, difloxacin was more bactericidal than enrofloxacin.     

凉州区奶牛乳房炎主要病原分离鉴定及药敏试验

乳房炎是奶牛最常见的疾病,随着奶牛产业的蓬勃发展,乳房炎已经成为制约该产业的瓶颈之一。从甘肃省武威市凉州区5个奶牛养殖场采集了1 000头荷斯坦奶牛共计3 991份奶样,对临床型和非临床型(隐性)乳房炎分别通过临床症状和兰州隐性乳房炎诊断液(LMT)进行诊断。结果显示:该地区奶牛临床型乳房炎发病率18.20%,亚临床型乳房炎发病率36.50%,总阳性率54.70%,总阳性乳区率54.72%。从阳性奶样中分离纯化得到的主要病原菌为链球菌属、金黄色葡萄球菌和大肠杆菌,且在不同类型乳房炎中的分布情况不同,临床型中链球菌属最多(54.37%),亚临床型中金黄色葡萄球菌最多(48.56%)。对主要病原菌进行药敏试验,结果表明治疗凉州区奶牛乳房炎的最佳药物应为喹诺酮类和庆大霉素类。