Morphological changes in brucellosis in guinea pigs due to experimental infection with Brucella suis

Sixty-four guinea-pigs were infected with the germs of Brucella suis. biotype 2 via the vaginal, peroral, intranasal, conjunctival and subcutaneous routes. In the subcutaneous infection the length of the trial was 151 days and in the other methods of infection 56 days. As found, guinea-pigs are highly sensitive and brucellosis lesions were detected in all cases. Brucellosis was chronical and spread locally in the guinea-pigs. In the haematogenic spreading of the disease, lesions were constantly recorded in liver, spleen and lungs. In the cases of subcutaneous infection lesions were also observed in regional lymph nodes at the places of infection. The lesions had amorphous structure and regular borders. The lesions were characterized by pronounced exudation and the exudate was subject to caseous necrosis, followed by colliquation. Secondary central colliquation was regularly observed in older granulomas, so that colliquation is considered as a characteristic morphological trait of brucellosis process in guinea-pigs. Histiocytary granuloma constituted the microscopic basis of the lesions. In older granulomas the central necroses were wide and contained a large amount of nuclear detritus, owing to the karyorhexis of cells, mainly neutrophiles. In the necroses of caseous type lipoid droplets were found and calcification was observed on the 151st day. Big cells of Langhans type occurred sometimes in the histiocytary layer of granulomas; they were present most frequently in the lesions affecting lymph nodes. Interstitial pneumonia was a characteristic symptom in liver and big cells of Sternberg type were found in the granulomas. Follicular tumor was constantly observed in spleens. Morphologically detectable lesions first developed in liver after about 7 days from conjunctival infection and subcutaneous infection, after 21 days from vaginal and intranasal infection, and after 28 days from peroral infection. In the case of subcutaneous infection, granulomas were found in the regional lymph nodes on the 14th day. The most expressive lesions in organs, mainly liver, were found from the 49th to 56th day from infection.     

Evaluation of cattle for experimental infection with and transmission of Brucella suis biovar 4

OBJECTIVE: To determine whether cattle can become persistently infected with Brucella suis biovar 4, whether the organism can be transmitted vertically or horizontally, and whether tests for bovine brucellosis are diagnostic. DESIGN: Observational study. ANIMALS: 24 pregnant cows and their calves and 6 bulls. PROCEDURE: Cows and bulls were housed separately in groups of 6 with each group consisting of 3 cattle experimentally infected with B suis biovar 4 and 3 na?ve animals. Cattle were observed for clinical signs daily; blood samples were collected weekly. Clotted blood from each sample was submitted for bacterial culture. Serum was tested with an indirect ELISA and the standard tube agglutination test (STAT), buffered plate agglutination test, brucellosis card test (BCT), and complemen't fixation test (CFT). Tissues collected at necropsy were submitted for bacterial culture and histologic examination. RESULTS: All 15 inoculated cattle seroconverted on 2 or more serologic tests, and bacteria were isolated from 4 inoculated cows at necropsy. There was no bacteriologic evidence of vertical or horizontal transmission, and none of the cattle developed clinical abnormalities or gross or histologic lesions. Results of the indirect ELISA were positive for all inoculated cattle. The other tests gave variable results; the CFT, STAT, and BCT yielded negative results for at least 1 of the 4 cattle from which the organism was isolated. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that cattle-to-cattle transmission of B suis biovar 4 is unlikely. Serologic tests for bovine brucellosis should be used cautiously when attempting to identify cattle with rangiferine brucellosis, as they do not discriminate between the 2 diseases and vary in their ability to detect exposed cattle.     

Brucella suis biotype 1 infection in a dog

Brucella suis biotype 1 was isolated from the semen of a dog with hindlimb weakness and a large, firm, left epididymis. A semen sample was oligospermic, with many neutrophils, the numbers of which decreased in serial sampling. A card agglutination test for B abortus and a rapid slide agglutination test for B canis were positive. The modified 2-mercaptoethanol slide agglutination test for B canis and the agar gel immunodiffusion test, using B canis cell wall antigen, were negative. At necropsy, chronic granulomatous inflammation was found in, and B suis biotype 1 was isolated from, the left epididymis and prostate gland.     

Brucella suis biovar 3 infection in a Kentucky swine herd

Sows from a large farrow-to-finish operation in western Kentucky had late-term abortions. Boars and breeding-age sows were tested serologically for brucellosis, and 83 of 125 were classified as reactors. No brucellae were isolated from the tissues of 6 unbred reactor sows, but Brucella suis biovar 3 was recovered from 5 aborted fetuses. Epidemiological studies failed to determine the source of the infection.     

猪布鲁菌感染对巨噬细胞泛素蛋白酶体功能的影响

巨噬细胞(macrophage,M_φ)是布鲁菌的主要宿主细胞,为了探明布鲁菌感染与M_φ泛素-蛋白酶体功能之间的关系,本试验在应用蛋白酶体抑制剂Lactacystin与促进剂IFN-γ的基础上,对M_φ感染猪种布鲁菌S2株(Brucella suis,B.suis)后细胞上清中泛素、蛋白酶体及胞内B.suis数量进行了检测。结果显示,IFN-γ与Lactacystin对M_φ泛素的表达均有明显的促进作用,且IFN-γ效果显著优于Lactacystin;其次IFN-γ有效促进了M_φ蛋白酶体的表达,而Lactacystin显著抑制了M_φ蛋白酶体的表达。在此基础上,将B.suis感染不同状态的M_φ,在0.5~24h分别进行胞内B.suis计数,研究表明,一方面B.suis的感染促进了M_φ泛素及蛋白酶体的表达;另一方面,泛素蛋白酶体系统功能的增强,显著降低了B.suis的早期感染,但对于B.suis感染中后期作用不明显,而感染后期抑制M_φ蛋白酶体功能,却可显著降低B.suis的胞内增殖能力。     

Experimental infection of rabbits (Oryctolagus cuniculus) with Brucella suis biovar 1 isolated from wild hares (Lepus europaeus)

Brucella suis biovar 1 is the causative agent of brucellosis in several domestic and wild animals and it is a common agent of human brucellosis. European hares (Lepus europaeus) have been shown to be infected by B. suis biovar 1 and the transmission to other animals has been suggested. In this work, experimental rabbits (Cuniculus orictolagus) were infected with B. suis biovar 1 isolated from wild hares. Infected rabbits showed high serological response in 2 weeks after discharge and typical granulomatous lesions (2mm diameter) were found in liver, spleen and kidneys after 50 days. B. suis biovar 1 was cultured from the lesion of the organs mentioned above as well as from urine, placenta and fetuses. These data suggest that hares are a potential source for horizontal transmission of B. suis biovar 1 to other mammalians.     

猪链球菌2型对家兔的试验感染及其病理学观察

利用猪链球菌2型四川分离强毒菌株458#,通过腹腔注射接种日本大耳白家兔,观察其临床症状和病理学变化。结果表明,猪链球菌2型强毒株458#可引起家兔发病或死亡,并能从发病家兔的组织样品中分离到所接种毒株。感染家兔的特征性组织病理学变化表现为典型的弥漫性血管内凝血和微血栓形成。猪链球菌2型感染家兔动物模型的建立,对于评价猪链球菌2型的毒力,探索其发病机制具有重要的意义。     

Brucella suis identification and biovar typing by real-time PCR

Fast and accurate identification of Brucella suis at the biovar level is an important issue for public health laboratories because some of the biovars that infect suidae (boars and pigs) are pathogenic for humans while others are not. Since classical biovar typing methods are often time-consuming, hard to standardize and require high-level biosafety containment, methodological improvements are desirable. This article describes new single nucleotide polymorphism (SNP) signatures for the rapid identification and biovar characterization of B. suis. These SNPs were included together with previously described ones in real-time PCR assays applicable to low-biosafety conditions. Allelic profiles unique for each B. suis biovar were defined and the most relevant signatures were determined on a collection of 137 field strains of worldwide origin characterized previously. Biovars assigned with both present and classical methods were globally consistent except for some biovar 3 field strains which matched the allelic profile of biovar 1.     

Evaluation of an Immunochromatographic Test to the Diagnosis of Canine Brucellosis Caused by Brucella canis

This study evaluated the performance of an immunochromatographic test (ICT) for the diagnosis of canine brucellosis caused by Brucella canis, comparing its results with that of the rapid slide agglutination test with and without the use of 2‐mercaptoethanol and the agar gel immunodiffusion test (AGID). The microbiological culture, PCR and clinical examination were used as reference. According to the results obtained in clinical examination, blood culture, culture of semen and vaginal swab and PCR in blood, semen and vaginal swab, a total of 102 dogs were divided into three groups: B. canis‐infected dogs (Group 1), B. canis‐non‐infected dogs (Group 2) and dogs with suspected brucellosis (Group 3). The diagnostic sensitivity of RSAT, 2ME‐RSAT, AGID and ICT in Group 1 was, respectively, 75%, 37.5%, 27.8% and 89.58%. The diagnostic specificity of RSAT, 2ME‐RSAT, AGID and ICT in Group 2 was, respectively, 91%, 100%, 100%, and 100%. In dogs with suspected brucellosis, 9.67% were RSAT positive, none was positive by 2ME‐RSAT, 3.22% were AGID positive and 6.45% were ICT positive. The main drawback concerning canine brucellosis diagnosis is the lack of a highly sensitive serological assay to be used as a screening test to the rapid identification of infected animals. The ICT showed a high diagnostic specificity and a diagnostic sensitivity value greater than that observed in the RSAT, 2ME‐RSAT and AGID. However, 10.41% of infected dogs had negative results by ICT. These dogs were positive by microbiological culture and/or PCR, indicating active infection and consequently a higher potential of spreading Brucella. Although rapid and simple to perform, the ICT lacked sensitivity to be used as a screening test.     

表达绿色荧光蛋白猪种布鲁菌的构建

为了更直观的观察猪种布鲁菌在宿主细胞/宿主体内的活动规律,本研究首先应用PCR技术扩增pEGFP-N1质粒的N1基因,与pMD18-T载体连接后,成功构建PMD18-N1质粒;进而应用XbalⅠ与SacⅡ限制性内切酶,将PMD18-N1质粒与pBBR1MCS-6质粒分别进行双酶切,并将N1片段反向连接于PBBR1MCS-6载体上,经菌液PCR鉴定与双酶切鉴定,证明成功构建了PBBR1MCS-N1质粒;在此基础上,将PBBR1MCS-N1质粒电转化至猪种布鲁菌感受态细胞,通过含氯霉素的TSA培养基筛选、PCR鉴定、荧光鉴定及菌株的传代稳定性试验与培养特性的观察,最终获得了可稳定表达绿色荧光的猪种布鲁菌。     

Trichuris suis population dynamics following a primary experimental infection

Trichuris suis population kinetics was studied by experimentally infecting 40 pigs with 5000 T. suis infective eggs. Six pigs were sacrificed every 2 weeks from 1 to 9 weeks post-inoculation (p.i.) and the remaining 10 pigs were sacrificed 11 weeks p.i. to estimate worm burdens. An equal number of uninfected control pigs were sacrificed at the same time points for comparison. Egg excretions from each pig were evaluated every week from 5 to 11 weeks p.i. Peripheral blood eosinophilia and basophilia were also evaluated every 2 weeks throughout the experimental period. After an initial almost 100% establishment of T. suis an expulsion phase followed approximately 9 weeks p.i., resulting in an aggregated distribution of the worms in the pig population, as it is characteristic for most helminth infections. By 11 weeks p.i. almost all worms had been expelled. Egg excretion peaked 7 weeks p.i. and a significant non-linear relationship between worm burdens and egg excretion was found. The predilection site for T. suis was the caecum and proximal colon and the relative distribution of worms in the large intestine did not change over time until expulsion. Peaking peripheral blood eosinophil and basophil levels were observed in the T. suis infected pigs 5 and 7 weeks p.i., respectively, reflecting the host activated immune response against the parasite. We here describe the course of a primary T. suis infection in pigs by detailed worm counts, demonstrating an effective expulsion that results in an almost 100% clearance of infection as previously indicated by monitoring faecal egg excretion.     

Passive immunization of pigs against experimental infection with Streptococcus suis serotype 2

The safety and protective efficacy of a horse antiserum raised against inactivated whole cell preparations of Streptococcus suis serotype 2 was investigated in pigs by experimental challenge. The antiserum was evaluated in two similar experiments each comprising 12 4-week-old pigs treated with 6 ml of antiserum the day before challenge and four pigs used as challenge controls. Pigs were infected by subcutaneous injection with approximately 10(11) colony forming units of S. suis serotype 2. Clinical disease in the pigs that could be attributed to infection with S. suis was reduced from 88 to 35% (P = 0.015). The percentage of pigs with lesions that could be associated with S. suis was reduced from 88 to 22% (P = 0.002) and isolation of S. suis serotype 2 was reduced from five (63%) out of eight pigs in the combined challenge control groups to 3 (13%) out of 23 pigs in the combined treatment groups. These results indicate that passive immunization of pigs may be a way to reduce or control S. suis serotype 2 infections in pigs.     

Respiratory chlamydial infection based on experimental aerosol challenge of pigs with Chlamydia suis

An experimental study of aerogeneous challenge in pigs was conducted in order to reveal characteristic features of porcine respiratory chlamydiosis. Eight conventionally raised pigs were exposed to a pathogenic strain of Chlamydia (C.) suis, four controls were mock infected. Besides pathological changes, the acute-phase and humoral immune responses, as well as the dissemination and transmission of the challenge strain was monitored in the course of infection. The data from clinical investigations, LPS-binding protein assay, antibody ELISAs, confocal laser scanning and light microscopy, immunohistochemical staining and PCR provided extensive evidence of the pathogenic potential of C. suis for the porcine respiratory system. This model appears suitable for further pathophysiological and immunological investigations of chlamydial respiratory infections and can also be recommended for studies of Chlamydia-associated infections of the human lung.