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1.
A total of 292 bovine Staphylococcus aureus isolates obtained from the 1950s (86 isolates), 1992 (107 isolates), and 2000 (99 isolates) were examined for antimicrobial susceptibility and phage typing. The same types of S. aureus (80, 52, 3A, 3A/3C, 42E, 77) were found among the isolates from all three time periods, representing 43.3% of the typeable isolates. This indicates that the Danish S. aureus population related to bovine mastitis has remained relatively unchanged over the last 50 years. The occurrence of antimicrobial resistance has remained low in Denmark in comparison to other countries in Europe.  相似文献   

2.
OBJECTIVE: To determine whether particular antimicrobial susceptibility profiles of bovine mastitis-causing Staphylococcus aureus isolates were associated with specific S aureus genotypes. SAMPLE POPULATION:357 S aureus isolates recovered from milk samples submitted for diagnostic bacteriologic testing from 24 dairy herds. PROCEDURES: Antimicrobial susceptibility of S aureus isolates was assessed by determining minimum inhibitory concentrations (MICs) to 14 antimicrobial agents. After digestion of S aureus genomic DNA by SmaI, electrophoretic patterns were obtained via pulsed-field gel electrophoresis (PFGE) and used to classify isolates into types. Gels were analyzed, and data were used to prepare dendrograms. RESULTS: 308 of 357 (86%) S aureus isolates were susceptible to all antimicrobials evaluated. Forty-nine S aureus isolates were resistant to 1 or more antimicrobials; of these isolates, 37 were resistant only to penicillin, 9 were resistant to penicillin and erythromycin, 2 were resistant to tetracycline, and 1 was resistant to erythromycin. Isolates were assigned to 7 PFGE types. An association was found between PFGE type and antimicrobial susceptibility profile. Organisms with resistance to at least one of the tested antimicrobial agents were identified in only 4 of the 7 types of S aureus. CONCLUSIONS AND CLINICAL RELEVANCE: Antimicrobial resistance was uncommon among the mastitis-causing S aureus isolates identified in the milk samples. A limited number of genotypes were associated with mastitis in these herds. Antimicrobial resistance phenotypes were associated with particular S aureus PFGE types; this association may have implications for future treatment and control of S aureus-associated mastitis in cattle.  相似文献   

3.
PCR-assays for the detection of staphylococcal enterotoxins A-E, and H, toxic shock toxin 1, and exfoliative toxins A and B were evaluated against phenotypic methods, and performed well. Four hundred and fourteen isolates of Staphylococcus aureus from Danish cases of bovine mastitis were screened for genes encoding these superantigens. One hundred isolates from Danish human carriers were also included in the study. In contrast to the frequent presence of genes encoding and in vitro expression of superantigens among the human carrier isolates, only one of 414 isolates from bovine mastitis carried the genes encoding enterotoxin C and toxic shock toxin-1. These results further support the hypothesis that the bovine and human S. aureus reservoirs constitute two separate sub-populations of the species S. aureus. The results also show that these superantigens are generally not present in Danish S. aureus isolates from bovine mastitis, and thus play no essential role in the pathogenesis of bovine S. aureus mastitis.  相似文献   

4.
Gram-negative bacteria are responsible for approximately one-third of the clinical cases of bovine mastitis and can elicit a life-threatening, systemic inflammatory response. Lipopolysaccharide (LPS) is a membrane component of Gram-negative bacteria and is largely responsible for evoking the inflammatory response. Antibiotic and anti-inflammatory therapy for treating Gram-negative infections remains suboptimal. Bactericidal/permeability-increasing protein (BPI) is a neutrophil-derived protein with antimicrobial and LPS-neutralizing properties. Select peptide derivatives of BPI are reported to retain these properties. The objective of this study was to evaluate the antimicrobial activity of a human BPI-derived synthetic peptide against clinical bovine mastitis isolates of Gram-negative bacteria. A hybrid peptide was synthesized corresponding to two regions of human BPI (amino acids 90-99 and 148-161), the former of which has bactericidal activity and the latter of which has LPS-neutralizing activity. The minimum inhibitory (MIC) and bactericidal (MBC) concentrations of this peptide against various genera of bacteria were determined using a broth microdilution assay. The MIC's were determined to be: 16-64 microg/ml against Escherichia coli; 32-128 microg/ml against Klebsiella pneumoniae and Enterobacter spp.; and 64-256 microg/ml against Pseudomonas aeruginosa. The MBC's were equivalent to or 1-fold greater than corresponding MIC's. The peptide had no growth inhibitory effect on Serratia marcescens. The antimicrobial activity of the peptide was retained in the presence of serum, but severely impaired in milk. Further functional evaluation of the peptide demonstrated its ability to completely neutralize LPS. Together, these data support additional investigations into the therapeutic application of BPI to the treatment of Gram-negative infections in cattle.  相似文献   

5.
Mastitis is one of the most important diseases in dairy cattle of which Staphylococcus aureus is a major pathogen. Despite an apparently good antimicrobial susceptibility in vitro, the cure of diseased animals from this bacteriological infection is often disappointing, which results in cases of recurrent clinical- and chronic subclinical infections. It has been suggested that these recurrent and chronic Staphylococcus infections can be attributed to the growth of bacteria in biofilm. The objective of this study was to compare the susceptibility for antimicrobial agents of S. aureus isolates obtained from bovine mastitis growing under different conditions. These conditions include a conventional conventional microbroth dilution assay in which minimal inhibitory concentration values are determined, the MBEC assay which measures both the susceptibility in biofilm and the susceptibility of sequester cells released from the biofilm. A comparison of the susceptibility for antimicrobial agents of a number of representative S. aureus isolates grown in broth (representing in vitro growth conditions) or milk (representing in vivo growth conditions) is also made. The results indicate that S. aureus isolates obtained from bovine mastitis are highly resistant to antimicrobial agents when growing in biofilms.  相似文献   

6.
The antibacterial effect of lactoferrin (Lf) was tested on isolates of Escherichia coli (E. coli), Staphylococcus aureus (S. aureus), and coagulase-negative staphylococci (CNS) as well as on Pseudomonas aeruginosa (P. aeruginosa) and Klebsiella pneumoniae (K. pneumoniae), originally isolated from bovine mastitis. Concentrations of Lf used were 0.67 mg/ml, 1.67 mg/ml, and 2.67 mg/ml. Growth of udder pathogens was monitored by turbidometry either in broth culture or in whey prepared from normal milk. We focused on 3 different growth variables: lag time, slope, and maximum absorbance of bacterial growth curves. Growth inhibition was seen in the broth but hardly at all in whey. The isolates of E. coli and CNS did not grow sufficiently well in whey to draw any conclusions. The most effective inhibitory activity of Lf was seen against E. coli and P. aeruginosa. All 5 E. coil isolates had similar growth patterns. Inhibition of growth by Lf was concentration-dependent. The concentration of 0.67 mg/ml in broth and whey was generally too low for a significant inhibitory effect.  相似文献   

7.
Antibacterial activity of bovine lactoferrin hydrolysates (LFH) on microorganisms isolated from bovine mastitis, and superoxide (O(2)(-)) production of bovine neutrophils were evaluated. Antibacterial effects of LFH were measured in vitro against Staphylococcus aureus, coagulase-negative staphylococci, Streptococci, Enterococci, Escherichia coli, Klebsiella pneumoniae, yeast-like fungi and Prototheca zopfii isolated from clinical cases of bovine mastitis. To compare susceptibilities against LFH, minimal inhibitory concentration (MIC) values were determined by a micro-plate assay method. Most organisms were sensitive to LFH. Prototheca zopfii was highly sensitive to LFH; the growth of the microorganism was inhibited completely even at 1 mug/ml. Staphylococcus aureus and Escherichia coli were resistant to LFH. The production of O(2)(-) by bovine neutrophils was used to evaluate the effect of LFH administration on functional activity. Increase in O(2)(-) production by bovine neutrophils occurred upon addition of LFH to neutrophils. These results demonstrate that LFH possesses antibacterial activity against pathogens that cause mastitis and activates neutrophil superoxide production.  相似文献   

8.
The aim of the research was to test whether exogenic virulence factors secreted by Staphylococcus aureus isolates are involved in mechanisms that allow the bacteria to modulate and evade phagocytosis by bovine polymorphonuclear neutrophils. The research was based on the comparison of the effects of supernatants, prepared from cultures of 30 S. aureus isolates, on the functional properties of bovine neutrophils in vitro. S. aureus isolates were collected from milk samples from cows with clinical mastitis. Supernatants, which were used to treat leukocytes, were prepared from 18 h S. aureus cultures. Exogenic virulence factors secreted by S. aureus isolates significantly influenced the phagocytosis parameters evaluated. Depending on their leukotoxic or superantigenic properties, supernatants could affect the ingestion process, and also showed an influence on the digestion efficiency and phagocytosis carried out by bovine polymorphonuclear neutrophils in vitro.  相似文献   

9.
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.  相似文献   

10.
Population diversity was evaluated in strains of Staphylococcus aureus isolated from bovine mastitis in Brazilian dairy herds by PCR-RFLP and sequencing of the 3'-terminal portion of the coagulase gene, and the susceptibility of strains to antimicrobials. The results showed great diversity in S. aureus population studied and the existence of predominant clones that account for most infections. No associations between the predominant types observed in the PCR-RFLP and the forms of presentation of the mastitis or to any of the different patterns of antimicrobial resistance were observed.  相似文献   

11.
This study was conducted to investigate the diversity of phage types and associations between penicillin resistance and phage types among 815 Staphylococcus aureus isolates from bovine mastitis in nine European countries and USA. All isolates were examined for susceptibility to antimicrobial agents and characterised by phage typing. Penicillin resistance was found among strains from all countries with an average occurrence of 32.4% (2-71.4%). A total of 76% of isolates were identifiable by phage typing and 144 different phage types were observed. The most predominant types were phage type 29 (11% of the 815 isolates), phage type 52 (5%), and phage type 80 (5%). Phage type 95 and 29/52/52A/80 were both distributed within seven countries. In the countries with the highest occurrence of penicillin resistance a reduced diversity of phage types and phage groups was observed. Phage group III was significantly associated with penicillin resistance in contrast to phage group I (P=0.0023) and phage complex-80 (P=0.0066). This study confirms that a large number of phage types of S. aureus cause bovine mastitis, but that some types predominate. In addition, these findings could indicate that the use of penicillin in the bovine environment has selected for specific types of S. aureus in countries with a high frequency of resistance.  相似文献   

12.
Staphylococcus aureus is the most common causative agent of bovine mastitis and vaccines developed to control this disease showed limited protection due in part to the lack of common antigens among the mastitis isolates. We isolated and identified two genes encoding proteins with glyceraldehyde-3-phosphate dehydrogenase (GAPDH) activity from a S. aureus strain isolated from bovine clinical mastitis. The GapB and GapC proteins share considerable homology to the GapB and GapC products of human strains of S. aureus. These two proteins could be distinguished by their different GAPDH activities and binding to bovine transferrin properties. Both gapB and gapC genes were conserved in 11 strains tested, and the GapC protein was present on the surface of all S. aureus strains.  相似文献   

13.
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14.
Strains of Staphylococcus species isolated from bovine mastitic milk at 66 dairy farms in Japan during the period from November 1988 to May 1989 were identified, and examined for their drug susceptibility and beta-lactamase production in order to clarify an epidemiological aspect of bovine mastitis caused by staphylococci. The results of bacteriological identification showed that the most predominant species was S. xylosus. Other major species isolated were S. aureus, S. sciuri and S. hyicus. Thirty-eight (71.7%) isolates of S. xylosus, 21 (45.7%) of S. aureus and 5 (71.4%) of S. epidermidis were positive for beta-lactamase production. Most of the beta-lactamase-producers of S. aureus were classified as high producers, although all of the beta-lactamase-positive S. xylosus isolates remained to be low producers. All isolates of S. aureus were sensitive to methicillin and cloxacillin at 6.25 micrograms/ml and 1.56 micrograms/ml, respectively, and none of methicillin-resistant S. aureus were detected. Isolates of other species were considered to be susceptible to 6 beta-lactams, in contrast to human isolates, but antibacterial activities of penicillin G and ampicillin were affected more strongly by beta-lactamase than those of methicillin, cloxacillin, cefazolin and cefoperazone.  相似文献   

15.
Following the rapidly expanding dairy enterprise, mastitis has remained the most economically damaging disease. The objective of this study was mainly to investigate the in vitro antibacterial activities of ethanol extracts of Combretum molle (R.Br.Ex.G.Don) Engl & Diels (Combretaceae) against antibiotic-resistant and susceptible Staphylococcus aureus and Streptococcus agalactiae isolated from clinical cases of bovine mastitis using agar disc diffusion method. The leaf and bark extracts showed antibacterial activity against S. aureus at concentrations of 3 mg/ml while the stem and seed extract did not show any bioactivity. Although both leaf and bark extracts were handled in the same manner, the antibacterial activity of the bark extract against the bacterial strains had declined gradually to a lower level as time advanced after extraction. The leaf extract had sustained bioactivity for longer duration. The susceptibility of the bacteria to the leaf extract is not obviously different between S. aureus and S. agalactiae. Also, there was no difference in susceptibility to the leaf extract between the antibiotic-resistant and antibiotic-sensitive bacteria. Further phytochemical and in vivo efficacy and safety studies are required to evaluate the therapeutic value of the plant against bovine mastitis.  相似文献   

16.
The aim of the study was to isolate gram-positive cocci from cows with mastitis and to determine their resistance to beta-lactamic antibiotics. Eight hundred and nine strains were isolated and identified as staphylococci (n=516), streptococci (n=199) and enterococci (n=94) from sub-clinical and clinical cases of bovine mastitis in Lithuania. The most common causative agents of udder disease included: S. epidermidis (n=176), S. aureus (n=176), S. agalactiae (n=134), S. hyicus (136) and E. hirae (n=68). Isolates were analysed for antimicrobial resistance to penicillin, ampicillin, amoxicillin, cephalothin, cephalexin, amoxicillin + clavulanate. The susceptibility patterns were analysed using the agar disk diffusion method. S. aureus showed the highest level of resistance to amoxicillin (81.3%), penicillin (76.7%) and ampicillin (78.4%). The corresponding values for CNS strains were 59.7%, 59.7% and 50.6% against penicillin, ampicillin and amoxicillin respectively. Streptococci were the most frequently resistant to amoxicillin (29.3%), and enterococci to penicillin (27%), amoxicillin (27.5%) and amoxicillin/clavulanic acid (23.8%). The resistance of all tested mastitis pathogens to aminopenicillins and penicillin highly correlated (r=0.83). Compared with other antibiotics, amoxicillin and clavulanic acid combination tended to be more effective (p<0.05) against all tested bacteria in vitro. However, S. aureus, in 38.1% of cases, was resistant to this combination of antimicrobials. This study demonstrates that S. epidermidis, S. aureus, S. hyicus, S. agalactiae and E. hirae remain the most frequent mastitis causative agents on Lithuanian cattle farms. The highest resistance in vitro to penicillins was demonstrated by S. aureus, S. hyicus and S. intermedius. Resistance to cephalosporins remains low, irrespective of bacterial species of gram-positive cocci.  相似文献   

17.
This report compares the in vitro activity of three cephalosporins (cephalothin, cefoxitin and ceftriaxone) against 57 Staphylococcus aureus strains isolated from cows with clinical mastitis on the basis of the minimal inhibitory (MIC) and minimal bactericidal concentrations (MBC). The majority of the S aureus strains showed resistance to cefoxitin and ceftriaxone and sensitivity to cephalothin. The highest MICs and MBCs were found for cefoxitin and ceftriaxone. Antimicrobial tolerance (MBC/MIC greater than or equal to 32:1) was observed in relation to cephalothin and ceftriaxone. The data suggest that these cephalosporins may not be effective for the treatment of staphylococcal bovine mastitis. The precise definition of their antimicrobial efficacies requires more detailed in vitro and in vivo studies.  相似文献   

18.
本研究旨在了解甘肃地区奶牛乳房炎金黄色葡萄球菌的耐药性和耐甲氧西林金黄色葡萄球菌的感染情况,为奶牛乳房炎的防制提供理论依据。采用KB纸片扩散法,检测17株金黄色葡萄球菌对8种不同抗菌药物的敏感性;再用琼脂稀释法检测了苯唑西林、万古霉素对金黄色葡萄球菌的最小抑菌浓度(MICs);头孢西丁纸片扩散法和PCR扩增特异性mecA耐药基因对所有受试菌株进行全面的耐甲氧西林金黄色葡萄球菌检测。结果表明,菌株对青霉素、磺胺异恶唑具有较强抗性,而对环丙沙星、头孢唑啉、万古霉素和苯唑西林全敏感;头孢西丁纸片扩散法未能检测出表型为MRSA的阳性菌株,而PCR方法却检测出8株mecA基因阳性菌株,且这些菌株的苯唑西林MIC均小于2 μg/mL。菌株的耐药情况较严重,对甲氧西林敏感而携带mecA基因的菌株高频存在于被调查地区的奶牛场中。  相似文献   

19.
为获得抑制奶牛乳房炎源金黄色葡萄球菌(S. aureus)的乳酸菌(LAB),本研究从新疆巴音布鲁克牧区鲜牛乳和哈萨克族乳制品奶疙瘩样品中分离培养LAB,通过传统的分离鉴定与16S rDNA基因序列测序相结合的方法鉴定LAB种类,同时以临床奶牛乳房炎源金黄色葡萄球菌S. aureus N2为指示菌,采用双层琼脂扩散法检测分离株的抑菌能力。通过测定生长曲线确定分离株的生长稳定期,进而利用硫酸铵沉淀法透析提取稳定期内分离株的细菌素,并检测其细菌素抑菌效价。结果显示:从样品中筛选获得5株能够抑制指示菌生长的LAB,分别为希氏乳杆菌、干酪乳杆菌、粪肠球菌、戊糖片球菌和乳酸乳球菌亚种。生长规律曲线表明20 h^30 h为5株LAB的稳定期,此期培养液pH值维持在3.8~4.5。从培养20 h的5株LAB上清液中提取到了细菌素,经检测其具有抑菌活性,抑菌效价分别为457 IU/mL、1 023 IU/mL、676 IU/mL、1 862 IU/mL和1 023 IU/mL。本研究结果表明5株LAB通过在生长稳定期内维持较低酸性环境(pH<4.5),代谢产生细菌素对乳房炎源S. aureus发挥抑制生长作用。本研究为S. aureus性奶牛乳房炎的生物防治提供了实验依据。  相似文献   

20.
OBJECTIVE: To evaluate antimicrobial activity of bovine bactericidal permeability-increasing protein (bBPI)-derived synthetic peptides against mastitis-causing gram-negative bacteria. SAMPLE POPULATION: Bacterial isolates from the milk of cows with clinical mastitis. PROCEDURES: 3 peptides were synthesized with sequences corresponding to amino acids 65 to 99 (bBPI(6,599)) or 142 to 169 (bBPI(142,169)) or the combination of amino acids 90 to 99 and 148 to 161 (bBPI(9,099, 148,161)) of bBPI. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of these peptides against bacterial isolates from cows with mastitis were determined by use of a standardized broth microdilution assay. The ability of these peptides to retain their antimicrobial activity in serum and milk was also evaluated. Finally, bacterial lipopolysaccharide (LPS)-neutralizing activity of these peptides was assayed with the Limulus amebocyte lysate test. RESULTS: Of the 3 peptides tested, bBPI(9,099, 148,161) had the widest spectrum of antimicrobial activity, with MIC and MBC values ranging from 16 to 64 Mg/mL against Escherichia coli, Klebsiella pneumoniae, and Enterobacter spp and from 64 to 128 Mg/mL against Pseudomonas aeruginosa. None of the peptides had any growth-inhibitory effect on Serratia marcescens. The antimicrobial activity of bBPI(9,099, 148,161) was inhibited in milk, but preserved in serum. Finally, bBPI(142,169) and bBPI(9,099, 148,161) completely neutralized LPS. CONCLUSIONS AND CLINICAL RELEVANCE: bBPI(9,099, 148,161) is a potent neutralizer of the highly proinflammatory molecule bacterial LPS and has antimicrobial activity against a variety of gram-negative bacteria. The ability of bBPI(9099,148161) to retain antimicrobial activity in serum suggests a potential therapeutic application for this peptide in the management of gram-negative septicemia.  相似文献   

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