水牛牛巴贝斯虫体外培养的研究：Ⅱ.冷冻血清的应用和筛选

应用冷冻血清对１株采自自然感染的水牛牛巴贝斯虫进行了长达７２ｄ的体外连续培养，共继代２０次，培养７２ｈ红细胞染虫率最高达１４．１％，平均为８％～１０％。培养２０ｄ和３０ｄ的牛巴贝斯虫经液氮保存复苏后，接种于去脾水牛犊均引发了严重的牛巴贝斯虫病，从而说明已建立了水牛牛巴贝斯虫的体外连续培养，且经培养后的牛巴贝斯虫致病力没有改变。本试验利用６头份的冷冻健康水牛血清同时进行培养，结果发现，并非所有的健康水牛血清均适合于体外培养牛巴贝斯虫。这一发现对建立水牛牛巴贝斯虫的体外培养和研究水牛牛巴贝斯虫病均具有重要意义     

一例荚膜血清A型多杀性巴氏杆菌和牛肺炎支原体混合感染犊牛的诊治

新疆某牛场的犊牛出现咳喘、鼻镜干燥、精神不振、食欲废绝、呼吸困难甚至出现死亡的症状,通过对患病牛群的发病情况、剖检、细菌分离鉴定、PCR鉴定等试验方法进行检查。结果显示:病死牛肺部可见明显出血并且伴有肺部实质性肝变;牛支原体和多杀性巴氏杆菌PCR检测都为阳性,曼氏杆菌PCR检测为阴性。通过实验室诊断最终确定该牛场为多杀性巴士杆菌和牛肺炎支原体混合感染。本研究通过病理变化、实验室诊断等方法找到病因,为此类疾病的防控提供借鉴意义。     

牛副结核分枝杆菌LAMP快速检测方法的建立

为建立牛副结核分枝杆菌(MAP)的快速检测方法,本研究采用环介导等温扩增(LAMP)技术,以MAP的IS900基因序列为靶基因设计特异性引物,建立了MAP特异性的LAMP快速检测方法。经反应条件优化,检测结果显示,建立的LAMP检测方法具有良好的特异性,最低见测量为0.53 pg;与其他病原菌无任何交叉反应。临床样品检测与行业标准的符合率为100%。该LAMP检测方法可以用于MAP的常规检疫。     

Veterinarian injuries associated with bovine TB testing livestock in Michigan, 2001

Testing all the cattle in an entire state with a uniform procedure for each animal affords an opportunity to relate human injury data to a known number of animals handled while carrying out a standardized procedure. Our objective was to describe the type and incidence density of injuries associated with TB-testing a large number of cattle herds, and to delineate the various factors associated with the risk of injury. A survey was mailed to all veterinarians (N = 259) who had completed at least five official bovine TB (bTB) herd tests in Michigan in 2001. We collected data regarding basic demographics and health status, work experience, veterinary specialty, and practice information. Each veterinarian was also requested to complete a separate injury questionnaire for each injury received while TB testing livestock in 2001. Accurate addresses were found for 247 eligible veterinarians, 175 (71%) of whom returned the survey. Thirty-six veterinarians reported a total of 53 injuries (10 major, 12 minor and 31 self-treated). Hands (29%) and legs (21%) were the anatomic locations most frequently injured, with sprains/strains (30%) and abrasion/contusion (30%) the most common types of injuries sustained. The overall incidence density of injuries was 1.9 per 10,000 animals tested. Female gender (RR = 3.3), being employed by the government (RR = 4.5), and smoking (RR = 6.0) were significantly associated with a higher rate of injury. Significant colliniearities were found between some risk factors associated with an increased rate of injury and participants thought 81% of their injuries could have been prevented. These results are explained by the administrative structure of the bTB testing program in Michigan, and the changing demographics of the veterinary workforce.     

我国部分地区牛支原体肺炎和关节炎的病原体诊断

重庆等4省市从外省引进的肉牛群发生以严重肺炎和关节炎为主要表现的疾病。该病的病变主要集中于肺脏,其组织学变化主要是间质增生,纤维素渗出,干酪样坏死,淋巴细胞、嗜中性粒细胞和单核细胞浸润。PCR检测病牛肺脏显示牛支原体阳性,而丝状支原体丝状亚种、牛分支杆菌和多杀性巴氏杆菌为阴性。从肺等组织中分离到牛支原体以及大肠杆菌、沙门氏菌、链球菌、葡萄球菌和烟曲霉,没有分离到巴氏杆菌。结果显示本病是以牛支原体感染为主引起的牛支原体肺炎和关节炎,长途运输等应激因素是该病突发的重要诱因,其他细菌和/或真菌继发感染加重了病情。     

应用镰形扇头蜱及巴贝斯虫病水牛的染虫血感染黄牛试验

从非流行区购进无蜱寄生的健康黄牛５头与水牛犊１头，其中３头成年黄牛（２～８岁）在畜舍内用从流行区水牛身上采得的镰形扇头蜱Ｒｈｉｐｉｃｅｐｈａｌｕｓｈａｅｍａｐｈｙｓａｌｏｉｄｅｓｈａｅｍａｐｈｙｓａｌｏｉｄｅｓ成虫，置于牛耳壳内叮咬；１头运至流行区放牧，让蜱上身叮咬；１头黄牛犊（１０年龄）去蜱后２周，用液氮保存的患病水牛染虫血４ｍｌ（４×１０ ８个虫体）皮下注射；另一头去蜱水牛犊亦同时注射相同剂量的染虫血，作为对照。５头黄牛在试验期间其体温、精神、食欲正常，未出现任何临床症状，感染后１０～１２ｄ外周血液中出现少量不典型的牛巴贝斯虫，红细胞染虫率在０．１％以下，持续３～５６消失；对照牛则体温升高（３９．８～４１．１℃），出现贫血、黄疸等症状，红细胞压积（ＰＣＶ）降至２６％，外周血液中出现典型的牛巴贝斯虫，红细胞染虫率（ＰＰＥ）高达１２％。对采用蜱叮咬及注射来自病水牛的染虫血为何不能使黄牛感染发病，文中进行了讨论。     

Improved detection of Mycobacterium tuberculosis and M. bovis in African wildlife samples using cationic peptide decontamination and mycobacterial culture supplementation

In South Africa, mycobacterial culture is regarded as the gold standard for the detection of Mycobacterium tuberculosis complex (MTBC) infection in wildlife even though it is regarded as “imperfect.” We compared a novel decontamination and mycobacterial culture technique (TiKa) to the conventional mycobacterium growth indicator tube (MGIT) system using known amounts of bacilli and clinical samples from MTBC-infected African buffaloes (Syncerus caffer), white rhinoceros (Ceratotherium simum), and African elephants (Loxodonta africana). Use of the TiKa-KiC decontamination agent on samples spiked with 10,000 to 10 colony forming units (cfu) of M. bovis (SB0121) and M. tuberculosis (H37Rv) had no effect on isolate recovery in culture. In contrast, decontamination with MGIT MycoPrep resulted in no growth of M. bovis samples at concentrations < 1,000 cfu and M. tuberculosis samples < 100 cfu. Subsequently, we used the TiKa system with stored clinical samples (various lymphatic tissues) collected from wildlife and paucibacillary bronchoalveolar lavage fluid, trunk washes, and endotracheal tube washes from 3 species with known MTBC infections. Overall, MTBC recovery by culture was improved significantly (p < 0.01) by using TiKa compared to conventional MGIT, with 54 of 57 positive specimens versus 25 of 57 positive specimens, respectively. The TiKa mycobacterial growth system appears to significantly enhance the recovery of MTBC members from tissue and paucibacillary respiratory samples collected from African buffaloes, African elephants, and white rhinoceros. Moreover, the TiKa system may improve success of MTBC culture from various sample types previously deemed unculturable from other species.     

Mycobacterium fortuitum abortion in a sow

Two aborted Chester White pig fetuses were presented to a veterinary diagnostic laboratory in Illinois. Postmortem examination identified no gross abnormalities. Histologic evaluation revealed multifocal necrosis of chorionic epithelial cells, coalescing areas of mineralization in the placenta, and focal accumulations of viable and degenerate neutrophils in the lung. Intra- and extracellular acid-fast bacilli were identified in the lesions in both the placenta and lungs. Bacterial culture of stomach contents yielded heavy growth of Mycobacterium fortuitum, a rapidly growing nontuberculous mycobacterium (NTM), which was further confirmed through whole-genome sequencing. NTM are opportunistic pathogens commonly found in the soil and in contaminated water supplies. In animals, M. fortuitum is typically introduced through cutaneous wounds leading to infections limited to the skin, with systemic infection being uncommon. To our knowledge, abortion caused by M. fortuitum has not been reported previously.     

Occurrence of Mycobacterium spp. in ornamental fish in Italy

The occurrence of Mycobacterium spp. in freshwater and marine ornamental fish was studied in Italy from June 2002 to May 2005. Two surveys were carried out, one of aquarium fish sent to the Laboratory for diagnosis, and the other of prevalence of infection by mycobacteria in ornamental fish imported into Italy. Bacterial isolation was carried out from the spleen, kidney and liver, and the isolates were subsequently identified by biochemical tests. In the first survey, 387 fish were examined and Mycobacterium spp. were isolated from 181 (46.8%) fish. In the second survey 127 batches of ornamental fish from different countries were examined. Mycobacterium spp. were isolated from 38 (29.9%) batches. The following species were found: M. fortuitum, M. peregrinum, M. chelonae, M. abscessus, M. marinum, M. gordonae, M. nonchromogenicum and M. interjectum. There was a high prevalence of infection independent of the presence of macroscopic lesions. Mycobacterium fortuitum and M. chelonae were more prevalent than M. marinum in the samples examined.     

28株牛源分枝杆菌的耐药基因突变分析

为明确分离自宁夏、甘肃和新疆地区的28株牛源分枝杆菌的耐药基因突变情况,通过采用16S rRNA、MPT64和多位点PCR方法进行基因分型鉴定,L-J比例法对其进行药敏试验,随后用DNA测序技术对10株耐药菌和4株敏感菌进行耐药基因检测及突变分析。分型鉴定结果表明,28株分离株均属于结核分枝杆菌复合群(MTBC),其中26株为牛分枝杆菌,2株为人型结核分枝杆菌。药敏试验结果表明,28株MTBC中18株对4种一线抗结核药物敏感,10株耐药。耐药菌株中,1株对利福平耐药;7株对异烟肼、利福平和链霉素三重耐药;2株对异烟肼、利福平、链霉素和乙胺丁醇四重耐药。耐药基因突变位点分析结果表明,rpoB 378位点的突变率为10.0%;katG 463位点的突变率为100.0%;rrs 311位点的突变率为11.1%;embB 378位点的突变率为100.0%;oxyR-ahpC和rpsL位点无突变。提示宁夏、甘肃和新疆地区奶牛场中MTBC主要以牛分枝杆菌为主,且存在耐多药情况,耐药突变位点以katG 463和embB 378为主。