3种动物专用抗菌药在鸡肠道沙门菌的流行病学临界值的建立

为建立3种动物专用抗菌药（氟苯尼考、安普霉素和达氟沙星）在鸡肠道沙门菌的流行病学临界值（ECOFFs）,本研究选取2017年分离自广东省各地兽医站和养殖场的166株鸡肠道沙门菌,采用琼脂稀释法测定3种动物专用抗菌药的最小抑菌浓度（MIC）,利用非线性回归分析法及NRI（normalized resistance interpretation）和ECOFFinder程序分析处理MIC数据,并最终确定氟苯尼考、安普霉素和达氟沙星对鸡肠道沙门菌的ECOFFs。结果显示,氟苯尼考、安普霉素和达氟沙星对沙门菌的MIC范围分别为2~>512、4~>512和0.015~64 μg/mL;MIC₅₀和MIC₉₀分别为256和>512、16和>512、0.5和16 μg/mL。氟苯尼考和安普霉素的MIC值呈明显的单峰或双峰分布,且界限明显,而达氟沙星的MIC值出现了不连续多峰分布。综合3种分析方法,最终确定氟苯尼考、安普霉素和达氟沙星的ECOFFs分别为16、16和0.125 μg/mL。本研究初步得到了3种药物在鸡肠道沙门菌的ECOFFs,为沙门菌耐药判定标准的制定提供了参考。     

副猪嗜血杆菌对喹诺酮类药物流行病学临界值的建立

为了解副猪嗜血杆菌对喹诺酮类药物的耐药现状,制定副猪嗜血杆菌对喹诺酮类药物的流行病学临界值,本试验根据CLSI-VET中规定的方法,对ScienceDirect、PubMed、中国知网等数据库中副猪嗜血杆菌对喹诺酮药物的耐药数据进行收集,共收集到605株环丙沙星、74株氧氟沙星、322株左氧氟沙星、211株达氟沙星、276株萘啶酸、143株洛美沙星、638株恩诺沙星及262株马波沙星的最小抑菌浓度数据,进行数据修正后利用统计软件ECOFFinder进行整理拟合,得出MIC拟合直方分布图,确定了副猪嗜血杆菌对8种喹诺酮类药物的流行病学临界值和耐药率。8种药物中可用于兽医临床使用的药物为达氟沙星、恩诺沙星、马波沙星、环丙沙星;萘啶酸作为第1代喹诺酮抗菌药,已基本退出市场;另外3种药物不可用于兽医临床。通过调查研究得出副猪嗜血杆菌对氧氟沙星、萘啶酸、洛美沙星、左氧氟沙星、环丙沙星、达氟沙星、马波沙星、恩诺沙星药物的流行病学临界值分别为8、4、4、0.25、0.25、0.0625、0.0625和0.03125 μg/mL,说明目前野生型副猪嗜血杆菌对环丙沙星、达氟沙星、马波沙星和恩诺沙星这4种兽医临床可用药敏感性较高,临床推荐使用。文章得出的数据在国际交流频繁的时代普遍适用,在国内外缺乏相应药效学折点和临床折点时可作为敏感和耐药的判定标准,并为进一步制定临床折点,指导临床用药、延长喹诺酮药物的临床使用寿命奠定了基础。     

动物源肠球菌对饲用抗菌促生长剂流行病学临界值的建立

【目的】建立动物源肠球菌对抗菌促生长剂的流行病学临界值,并基于此了解2019-2021年动物源肠球菌对饲用抗菌促生长剂的耐药情况。【方法】测定2019-2021年从北京、河北、四川、山西、陕西和内蒙古养殖场动物样本中分离得到的肠球菌对7种抗菌促生长剂（吉他霉素、黄霉素、恩拉霉素、那西肽、阿维拉霉素、维吉尼亚霉素和杆菌肽）的最小抑菌浓度（MIC）值。根据CLSI-VET中规定的方法,利用统计软件ECOFFinder对MIC分布数据进行非线性回归模拟,以95%置信区间下的流行病学临界值计算动物源肠球菌对7种抗菌促生长剂的耐药率。【结果】确定了动物源肠球菌对吉他霉素、黄霉素、恩拉霉素、那西肽、阿维拉霉素、维吉尼亚霉素和杆菌肽的流行病学临界值分别为8、8、8、0.25、8、8和32 μg/mL。由此计算的药物耐药率为64.38%、34.48%、80.06%、14.69%、19.06%、50.69%和24.96%,其中动物源肠球菌对恩拉霉素、吉他霉素和维吉尼亚霉素的耐药情况最为严重,而对那西肽最敏感。在2019-2021年的耐药率变化情况统计中,黄霉素和杆菌肽的耐药率下降趋势较为明显。【结论】本研究制定了动物源肠球菌对7种养殖业常用抗菌促生长剂的流行病学临界值,可作为动物源肠球菌对饲用抗菌促生长剂敏感和耐药的初步判定标准,为进一步制定耐药折点,监测动物源肠球菌对抗菌促生长剂的耐药情况变化提供数据基础和科学依据。     

苍术油对四种细菌最小抑菌浓度的研究

本研究通过试管2倍稀释法测定了苍术油对大肠杆菌(44102)、金黄色葡萄球菌(26003)、枯草芽孢杆菌(63501)和沙门氏菌的最小抑菌浓度(MIC)。结果:苍术油对大肠杆菌、金黄色葡萄球菌和枯草芽孢杆菌的最小抑菌浓度分别为:117.2μg/ml、625μg/ml和312.5μg/ml,而对沙门氏菌没有任何抑菌作用。研究证明苍术油对部分细菌具有良好的抑制作用。     

金黄色葡萄球菌gyrA、gyrB、grlA、grlB和norA基因突变与环丙沙星耐药性的关系

采用药敏纸片法测定了临床分离的160株金黄色葡萄球菌对14种抗菌药物的耐药性，同时进行了MRSA的检测，并采用琼脂稀释法测定纸片法筛选出的45株金黄色葡萄球菌对环丙沙星的最低抑菌浓度（MIC）。根据金黄色葡萄球菌gyrA、gyrB、grlA、grlB和norA基因的核苷酸序列设计引物、以筛选出的45株金黄色葡萄球菌染色体DNA为模板．对gyrA、gyrB、grlA、grlB和norA基因片段进行PCR扩增，回收PCR产物．并与pMD18-T载体连接．转化DH5a感受态大肠杆菌．提取质粒并测序。进一步分析考查金黄色葡萄球菌gyrA、gyrB、grlA、grlB和norA基因及其启动子区突变与环丙沙星耐药性的关系。结果表明．grlA中的2个突变（位于80和84密码子）及gyrA中的2个突变（位于84和88密码子）与环丙沙星耐药性明显相关。     

福建白羽肉鸡鸡毒支原体的分离鉴定及最小抑菌浓度测定

为了解福建白羽肉鸡中鸡毒支原体（MG）对临床常用药物的敏感性,本研究从2个商品代白羽肉鸡养殖场采集了疑似患慢性呼吸道病（CRD）的鸡喉拭子40份,接种于适宜的支原体培养基上进行纯化。通过对菌株生物学特性分析、细菌形态学观察、PCR测序及动物回归试验进行鉴定,并利用最低抑菌浓度（MIC）测定了分离株对临床常用抗菌药的敏感性。结果显示,本试验成功获得2株MG,命名为MX和HY。将分离株接种于含有酚红的支原体液体培养基中能分解葡萄糖产酸使培养基颜色由红变黄;固体培养基于低倍镜下观察到典型的"荷包蛋"状菌落;分离株经过Dienes染色后的菌落中心呈致密深蓝色,边缘淡蓝,且30 min后不褪色,并具有较强吸附红细胞的能力,呈现典型MG培养特征;动物回归试验和分离株pvpA基因测序结果显示,2株MG分离株均具有较强致病性,证实其与R株的亲缘关系最近;药敏试验结果显示,MX和HY对泰妙菌素、氟苯尼考和泰乐霉素表现出了高度敏感性,其次是盐酸恩诺沙星、盐酸大观霉素,而对具有与泰乐霉素类似分子结构和抗菌谱的畜禽专用抗生素替米考星敏感性较差,此差异可能与临床上长期应用该药有关。研究结果表明,福建地区商品代白羽肉鸡中存在致病性MG流行,泰妙菌素可以作为临床防治MG的首选药物。     

氨基糖苷类药物诱导大肠埃希菌生物被膜对抗菌药物敏感性变化规律

采用浓度梯度递增法对3株不同生物被膜形成能力的大肠埃希菌进行4种氨基糖苷类药物(阿米卡星、链霉素、庆大霉素和安普霉素)诱导,考察大肠埃希菌生物被膜对其余8种不同抗菌药物最小抑菌浓度和最小膜清除浓度的变化规律。结果表明,在氨基糖苷类药物诱导压力下,3株不同生物被膜形成能力的菌株对其余8种抗菌药物的最小膜清除浓度比最小抑菌浓度至少增加8倍以上,与诱导前相比,大肠埃希菌形成生物被膜后,其最小膜清除浓度增加的速率比最小抑菌浓度快。     

Bacteriostatic Test in vitro of Traditional Chinese Medicine Radix Dichroa Powder on Common Pathogens

The purpose of this study was to indicate the antibacterial effect of traditional Chinese medicine Radix dichroa powder (RDP) in vitro. Minimal inhibitory concentration (MIC) of RDP for 12 pathogens were determined by two fold dilution method and agar dilution method, and the dose-effect relationship of antibiotic effect was studied, and the antimicrobial effect was also observed by Oxford-cup method. The results from two fold dilution method indicated that RDP had stronger inhibited effect to Streptococcus and Bacillus subtilis (MIC were from 15.6 to 62.5 mg/mL) than that of S. aureus (MIC was about 250 mg/mL), Enterobacteria (MIC was about 500 mg/mL) and fungi (MIC > 500 mg/mL); The results of agar dilution method showed that RDP had some antibacterial activities on Proteusbacillus vulgaris, Candida albicans And aspergillus niger, MIC was about 500 mg/mL, while it had weak antibacterial effect on other pathogens (MIC > 500 mg/mL). The results of Oxford-cup method showed that RDP (500 mg/mL) had some degree antimicrobial effect on pathogens, and the bacteria-inhibiting ring diameter of RDP not exceeded 10 mm, while drug-action ring was observed obviously around the Oxford-cup, and the number of bacteria was significantly decreased within the ring. In conclusion, RDP had some degree antibacterial effect on common pathogens, but the effects were not strong because of charicteristics of RDP itself and lower effective content.     

Comparative Analysis on Antibiotic Resistance Characteristics of Listeria spp. and Enterococcus spp. Isolated From Laying Hens and Eggs in Conventional and Organic Keeping Systems in Bavaria,Germany

By investigating the prevalence and antimicrobial resistance characteristics of Gram‐positive bacteria from organic and conventional keeping systems of laying hens, it was to be determined to what extent these properties are influenced by the different systems. For this purpose, a total of 799 cloacal swabs and 800 egg samples were examined. Prevalences for all selected bacteria from cloacal swabs were much the same for both organic and caged birds: Listeria spp. 1.3%_[org] versus 1.6%_[con]; Enterococcus spp. 95.5%_[org] versus 97.5%_[con]. Egg contents and eggshells were generally contaminated to a lesser extent, primarily with Enterococcus spp. Listeria isolates were susceptible to almost all tested antibiotics, only three Listeria innocua from conventional keepings were resistant to clindamycin; one isolate additionally to imipenem. High percentages of Enterococcus faecalis were resistant to doxycycline and macrolides. Enterococcus faecium proved to have high resistance rates to clindamycin, fosfomycin and erythromycin; 9.1% were even resistant to the reserve antibiotic synercid. Further, Enterococcus spp. showed higher resistance rates to doxycycline, erythromycin, fosfomycin and rifampicin. No glycopeptide resistant enterococci were detected. A correlation between keeping system and resistance/susceptibility rates could be demonstrated. In detail, E. faecalis from organic laying hen husbandries showed significant lower resistance prevalences to tylosin, streptomycin and doxycycline; susceptibility rates were higher for enrofloxacin and ciprofloxacin. Rifampicin and imipenem were more effective in isolates from conventional keepings (P < 0.05). The amounts of resistant isolates of the Enterococcus raffinosus from organic farms were significantly lower, the amounts of sensitive isolates were significantly higher than from conventional farms concerning eight antibiotics (P < 0.05). When comparing the susceptibility/resistance rates, as well as the mean minimum inhibitory concentrations values, the consistent tendency is that bacteria from organic layer flocks are more susceptible to antimicrobials. These results show that organic livestock farming plays a part in contributing to reduced antibiotic resistance.     

莱芜黑山羊胃肠道血管活性肠肽免疫反应阳性细胞的研究

比较观察血管活性肠肽(VIP)在莱芜黑山羊胃肠道内的分布,用免疫组织化学方法显示胃肠壁各层及各肠段的分布情况。结果表明,血管活性肠肽以贲门部数量最多,结肠、空肠、幽门、盲肠、直肠、胃底次之,十二指肠、回肠、食管中未发现VIP细胞。VIP细胞形态多样,呈椭圆形和锥体形,常有突起伸至其他细胞附近,也有突起直达上皮表面。     

Use of 2-hydroxy-4-(methylthio)-butanoic acid to inhibit Salmonella and Listeria in raw meat for feline diets and palatability in domestic cats

While the raw pet food market continues to grow, the risk of bacterial contamination in these types of diets is a major concern, with Salmonella enterica and Listeria monocytogenes being the most frequently associated pathogens in raw pet food product recalls. dl-Methionine is included in some commercial feline kibble and canned diets to improve protein quality; however, an alternative to this is a liquid methionine supplement, 2-hydroxy-4-(methylthio)-butanoic acid (HMTBa), which is also an organic acid. 2-Hydroxy-4-(methylthio)-butanoic acid has previously demonstrated similar efficacy to formic acid against pathogens in a liquid environment and may be a good candidate to inhibit S. enterica and L. monocytogenes in raw ground meat. First, the minimum inhibitory concentration and minimum bactericidal concentration of HMTBa against these pathogens under laboratory growth conditions were determined by measuring growth of pathogens over 36 h when exposed to 10 concentrations of HMTBa (0.10% to 1.00%) mixed with tryptic soy broth. 2-Hydroxy-4-(methylthio)-butanoic acid included at ≥0.50% was bactericidal to S. enterica and L. monocytogenes (P < 0.05). Next, five levels of HMTBa (0.50% to 1.25%) were included in raw ground meat mixtures inoculated with cocktails of S. enterica or L. monocytogenes, and contamination levels were determined at four timepoints: immediately, and after refrigerated storage (4 °C) at 24, 48, and 72 h after removal from freezer (24 h at −20 °C). 2-Hydroxy-4-(methylthio)-butanoic acid included as 1.25% of the meat mixture reduced S. enterica and L. monocytogenes compared with the control (P < 0.05); however, it did not result in total kill of either of these pathogens. Following this, feeding behaviors of seven domestic cats were assessed when offered a raw chicken diet treated with or without 1.25% HMTBa for 5 d each, after which a 2-d 2-choice preference test was conducted. Cats demonstrated a preference for raw diets without HMTBa, but still readily consumed diets with 1.25% HMTBa, suggesting that such a diet was still palatable to them.