数字PCR技术在动物疫病检测中的应用     

数字PCR技术在动物疫病检测中的应用 《畜牧与饲料科学》2021,42(6):124-128

为了支持动物疫病流行地区的控制及消除计划,以及在动物疫病非流行地区进行有效筛查,必须使用更加准确灵敏的诊断手段。数字PCR可实现绝对定量及痕量核酸的检测,与传统检测方法相比具有不依赖参考物或标准品、对抑制剂具有较高耐受性、灵敏度和精准度更高的优点。数字PCR作为一种定量分析的核酸检测新方法,在动物疫病检测中得到了应用,围绕数字PCR技术的原理、应用、存在的问题及发展趋势等进行综述及分析,为数字PCR在动物疫病检测中的进一步应用提供参考。  相似文献   

A serological and molecular study on Francisella tularensis in rodents from Hamadan province,Western Iran     

《Comparative immunology, microbiology and infectious diseases》2020

Introduction and purposeTularemia is a zoonotic disease, the most important hosts of which are rodents. Endemic regions and reservoirs of F. tularensis are not well-researched areas in Iran. The present study aimed to study F. tularensis infection in the rodent populations of western Iran.Materials and methodsSamples were collected in different areas of Kabudar Ahang County in Hamadan province (west of Iran) from 2014 to 2017. Tularemia serological and molecular tests were conducted using the tube agglutination test and Real-time PCR method tracking the ISFtu2 gene. Positive serum samples were evaluated for cross-reactivity with brucellosis.ResultsA total of 433 rodents, collected from 33 localities, were included in the study. The most abundant species belonged to the Persian jird (Meriones persicus; 75.5%), and Libyan jird (Meriones libycus; 10.1%). Among the studied samples, three (0.74 %) were seropositive and five (1.15%) were PCR positive. Seropositive samples were two M. persicus and one M. libycus, and PCR positive rodents were four M. persicus and one M. vinogradovi. Tularemia seropositive samples showed no cross-reactivity with brucellosis.ConclusionGiven the presence of infection in rodents with tularemia agent in the studied area, it is crucial to elucidate the risks of rodent exposure to tularemia for physicians, health personnel and the general population.  相似文献   

Serological and molecular detection of infection with Mycobacterium leprae in Brazilian six banded armadillos (Euphractus sexcinctus)     

《Comparative immunology, microbiology and infectious diseases》2020

Leprosy was recognized as a zoonotic disease, associated with nine-banded armadillos (Dasypus novemcinctus) in the Southern United States of America in 2011. In addition, there is growing evidence to support a role for armadillos in zoonotic leprosy in South America. The current study evaluated twenty specimens of the six-banded armadillo (Euphractus sexcinctus), collected from rural locations in the state of Rio Grande do Norte (RN), Brazil for evidence of infection with Mycobacterium leprae. Serum was examined using two "in-house" enzyme-linked immunosorbent assays (ELISAs) and via two commercially available (ML flow and NDO-LID®) immunochromatographic lateral flow (LF) tests, for detection of the PGL-I and/or LID-1 antigens of the bacterium. The presence of M. leprae DNA in liver tissue was examined using the multi-copy, M. leprae-specific repetitive element (RLEP), as target in conventional and nested PCR assays. Molecular and anti-PGL-I-ELISA data indicated that 20/20 (100 %) of the armadillos were infected with M. leprae. The corresponding detection levels recorded with the LF tests were 17/20 (85 %) and 16/20 (85 %), for the NDO-LID® and ML flow tests, respectively. Our results indicate that, in common with D. novemcinctus, six banded armadillos (a species hunted and reared as a food-source in some regions of Brazil, including RN), represent a potential reservoir of M. leprae and as such, their role in a possible zoonotic cycle of leprosy within Brazil warrants further investigation.  相似文献   

Molecular survey of Dirofilaria species in stray dogs,red foxes and golden jackals from Vojvodina,Serbia     

《Comparative immunology, microbiology and infectious diseases》2020

Cardiopulmonary dirofilariosis in dogs and other carnivores is caused by Dirofilaria immitis, while Dirofilaria repens usually causes a subcutaneous infection. The importance of red foxes and golden jackals in the epidemiology of dirofilariosis remains unknown. Thus, the aim of this study was to conduct a cross-sectional molecular survey of Dirofilaria species in stray dogs, red foxes and golden jackals from the endemic region of Vojvodina, Serbia, in order to determine and update data on their prevalence and provide insight into the epidemiological importance of wild canids.A total of 59 blood samples from stray dogs, 94 from red foxes and 32 from golden jackals were collected and screened by real-time PCR targeting a 115-bp fragment of the mitochondrial 12S gene of filarioids and by conventional PCR assay targeting a 484–524-bp fragment of 5.8S-ITS2-28S locus of filarioids.The cross-sectional molecular survey detected the filarioid mitochondrial 12S gene fragment in stray dogs (27.1 %), red foxes (8.5 %) and golden jackals (6.3 %) in the same endemic region of Vojvodina, Serbia. Only D. immitis was detected in stray dogs, while both D. immitis and D. repens were detected in populations of red foxes and golden jackals. These results outline a possible interaction of D. immitis infection between the dog population and the wild canid populations, while D. repens was found to circulate mostly in golden jackals and red foxes populations.  相似文献   

Death of a neonatal lamb due to Clostridium perfringens type B in New Zealand     

JS Munday  H Bentall  D Aberdein  M Navarro  FA Uzal  S Brown 《New Zealand veterinary journal》2020,68(4):242-246

ABSTRACT

Case history and clinical findings: A flock of 20 sheep was kept within three paddocks on a single property. None of the animals in the flock had been vaccinated against any disease for at least three years. Abdominal bloating and haemorrhagic diarrhoea were observed in Lamb 1 at 24 hours-of-age. The lamb subsequently died within an hour of the onset of clinical signs. Lamb 2 was 3-days-old when observed to be recumbent with opisthotonus. The lamb was treated with dextrose, vitamins B1 and B12, and penicillin G, but died 4 hours later.

Pathological findings: Examination of Lamb 1 revealed markedly increased gas within the peritoneum and within dilated loops of intestine. The intestines were dark red and contained large quantities of haemorrhagic fluid. Histology of the intestines revealed peracute mucosal necrosis with minimal accompanying inflammation. The intestinal lumen contained cell debris, haemorrhage, and myriad large Gram-positive bacilli. The intestines of Lamb 2 did not appear bloated or reddened. However, multiple fibrin clots were visible within the pericardial sac. Histopathological examination revealed small foci of necrosis within the mucosa of the distal intestine. The necrotic foci were often associated with large numbers of large Gram-positive bacilli.

Immunohistochemsitry and molecular biology: Intestinal samples from Lamb 1 were processed for Clostridium perfringens immunohistochemistry, which revealed large numbers of intralesional, positively immunostained rods. Fragments corresponding to the expected sizes for genes encoding alpha, beta, and epsilon C. perfringens typing toxins were amplified by PCR from DNA extracted from formalin-fixed sections of intestine.

Diagnosis: Lamb dysentery due to C. perfringens type B.

Clinical relevance: C. perfringens bacteria have a worldwide distribution, but disease due to C. perfringens type B has only been diagnosed in a small number of countries and has never been reported in New Zealand or Australia. C. perfringens type B produce both beta toxin and epsilon toxins, therefore both haemorrhagic enteritis and systemic vascular damage can develop. As many animals are exposed to C. perfringens without developing disease, there must be additional unknown factors that resulted in disease in these particular sheep. Vaccines that specifically protect against C. perfringens type B are available and may be recommended for use in smaller non-commercial flocks, as in the present case.  相似文献   

Serum amyloid A (SAA) mRNA expression in chicken and quails in response to bacterial stress     

《Comparative immunology, microbiology and infectious diseases》2020

Monitoring of acute phase proteins such as serum amyloid A at gene expression level may provide quick information about immune status of the host and its susceptibility towards common infections. Present study was carried out to evaluate and compare the mRNA expression of SAA gene in Rhode Island Red chicken (RIR) and Japanese quails using real time PCR analysis in response to inactivated Salmonella gallinarum culture. The results showed that expression of SAA gene was approximately 17–33 folds higher in case of birds administered with bacterial culture when compared to un-inoculated controls and expression was higher and quicker in case of quails than RIR chicken. The SAA genes from chicken and quail were cloned and upon sequence analysis it was observed that deduced amino acid sequence of SAA from chicken and quails were having approximately seven percent variation which might have significance in function of this protein in these species.  相似文献   

Outbreak of anaplasmosis associated with novel genetic variants of Anaplasma marginale in a dairy cattle     

《Comparative immunology, microbiology and infectious diseases》2017

During early lactation, dairy cows may present a transient immunosuppressive state and develop anaplasmosis caused by Anaplasma marginale. In this study, clinical anaplasmosis in dairy cattle in the Thrace region of Turkey was investigated with respect to within-herd prevalence, vertical transmission, and genetic diversity. In March and September 2015, thirty lactating cows showed primary clinical signs of anaplasmosis, including fever, anaemia, decreased milk yield, anorexia, and laboured breathing. Symptoms disappeared in most cows after administration of long-acting oxytetracycline, but nine of them (30%) died. Following diagnosis based on clinical signs, microscopy and molecular findings, blood samples were collected from apparently healthy lactating cows (n = 184), pregnant heifers (n = 39) and newborn calves (n = 24). DNA was extracted from each sample and analyzed for the presence of major surface proteins (MSPs) of A. marginale, followed by sequencing to assess diversity of isolates. Microscopic examination of erythrocytes revealed A. marginale inclusion bodies in symptomatic cows. Examination of thin blood smears showed 3.8% of the lactating, clinically asymptomatic, cows to be infected with A. marginale, while nPCR detected 31.0% positive. A. marginale infection was not detected in pregnant heifers by either method. Congenital infection was found in one calf by nPCR. This is the first report of transplacental transmission of A. marginale in Turkey. The MSP4 sequence analyses showed high genetic diversity among the isolates, presenting 97.6-99.6% homology at the amino acid level. The sequences of MSP1a amplicons revealed genetic diversity providing three new tandem repeats.  相似文献   

Molecular prevalence,risk factors assessment and haemato-biochemical alterations in hepatozoonosis in dogs from Punjab,India     

《Comparative immunology, microbiology and infectious diseases》2017

Hepatozoonosis caused by Hepatozoon canis is an important tick-borne disease of dogs in tropical and sub-tropical regions throughout the world. In the present study evaluation of blood samples collected from 225 dogs presented at Small Animal Clinics, GADVASU, Ludhiana, Punjab (India) was done for the presence of H. canis by PCR based assay targeting a portion of 18S rRNA gene. Of the total samples subjected to PCR, an amplicon of 666 bp was detected in 13.78% samples whereas, routine blood smear examination revealed gamonts in 5.78% samples. Furthermore, prevalence of H. canis infection was found to be significantly associated with season, being highest in summer and lowest in winter while other risk factors e.g. age, sex and breed showed non-significant association. In terms of various clinico-pathological parameters, significant drop in haemoglobin, total red blood cell count, packed cell volume and lymphocytes were recorded in positive cases whereas the total white blood cell count was non-significantly increased. The haematological alterations in the positive cases were lymphopenia, anaemia, thrombocytopenia, relative neutrophilia, neutrophilic leucocytosis, eosinophilia, monocytosis and lymphocytosis while the biochemical profile revealed hypoproteinemia and increased levels of blood urea nitrogen and creatinine (in positive cases) pointing towards renal failure.  相似文献   

利用QuantStudioTM 3D数字PCR分析转基因玉米MON863含量   总被引：1，自引：0，他引：1  

胡佳莹  姜羽  杨立桃 《农业生物技术学报》2016,(8):1216-1224

QuantStudioTM 3D数字PCR (QuantStudioTM 3D digital PCR,3D-dPCR)是一种基于超高密度亲疏水微孔芯片实现数字PCR分液原理的新型核酸绝对定量平台,在转基因生物定量领域具有极大的应用前景.本研究基于3D-dPCR平台,以转基因玉米(Zea mays)MON863混合样品为例,建立基于单重和双重数字PCR体系的转基因生物(genetically modified organisms,GMOs)含量分析方法.与传统qRT-PCR比较发现,在缺乏样品纯度、纯合度信息的情况下,数字PCR能够较好地排除这些因素的影响,测定准确的量值.研究结果表明,QuantStudioTM 3D数字PCR是一种适用于转基因生物含量分析的精确定量方法,还可反映转基因玉米种子的基因型.本研究基于3D-dPCR建立的转基因玉米MON863单重和双重定量方法为转基因检测提供了新的方法和参考.  相似文献   

Prevalence,molecular characterization and risk factor analysis of Ehrlichia canis and Anaplasma platys in domestic dogs from Paraguay     

《Comparative immunology, microbiology and infectious diseases》2019

This is the first study to investigate the prevalence and risk factors associated with Ehrlichia canis and Anaplasma platys positivity in dogs from Paraguay. Conventional PCR assays for the E. canis 16SrRNA gene and A. platys p44 gene were carried out in blood samples from 384 dogs from Asunción city, Paraguay. Sequencing and phylogenetic analysis were performed in selected positive E. canis and (16SrRNA gene) and A. platys (16S and p44 genes) samples. The overall prevalence of E. canis and A. platys in dogs in Paraguay was 10.41% (40/384) and 10.67% (41/384), respectively. Older dogs without veterinary care had higher odds for E. canis positivity and a higher number of dogs in the same household, as well as absence of anti-tick treatment were considered risk factors for A. platys. Ehrlichia canis and A. platys circulate in the dog population from Asunción, and are described for the first time in Paraguay.  相似文献