A preliminary survey of Chlamydia psittaci genotypes from native and introduced birds in New Zealand

AIM: To describe the Chlamydia psittaci genotypes in samples from native and introduced birds from New Zealand by analysis of the sequence variation of the ompA gene.

METHODS: DNA was extracted from samples collected from a non-random sample of birds; either swabs from live asymptomatic birds or birds with clinical signs, or formalin-fixed, paraffin-embedded (FFPE) samples from historical post-mortem cases. The presence of C. psittaci in all samples had been confirmed using a quantitative PCR assay. The C. psittaci ompA gene was amplified and sequenced from samples from 26 native and introduced infected birds comprising 12 different species. These sequences were compared to published available C. psittaci genotypes.

RESULTS: Genotypes A and C of C. psittaci were identified in the samples. Genotype A was identified in samples from nine birds, including various native and introduced species. Genotype C was identified in samples from 16 different waterfowl species, and a mixed infection of both genotypes was found in a kaka (Nestor meridionalis). In native birds, C. psittaci infection was confirmed in seven new host species.

CONCLUSIONS AND CLINICAL RELEVANCE: Two genotypes (A and C) of C. psittaci were found in samples from a wider range of both native and introduced species of birds in New Zealand than previously reported. Both genotypes have been globally associated with significant disease in birds and humans. These initial results suggest the host range of C. psittaci in New Zealand birds is under-reported. However, the prevalence of C. psittaci infection in New Zealand, and the associated impact on avian and public health, remains to be determined. There are biosecurity implications associated with the importation of birds to New Zealand if there is a limited diversity of C. psittaci genotypes present.     

动物鹦鹉热衣原体疫苗研究现状

鹦鹉热衣原体为多种人畜共患病的病原体。衣原体免疫学机制研究表明其主要引发Th1细胞调节的细胞免疫和体液免疫。减毒活疫苗、灭活疫苗的使用在一定程度上控制了其发病率。基因工程疫苗,如重组亚单位苗、核酸疫苗、细茵载体苗等也都在深入研究中。同时,人们也在筛选特异性的抗原,构建各种疫苗载体及选择免疫佐剂和免疫途径等方面做了大量的工作。     

鹦鹉热衣原体疫苗研究进展

文章叙述了鹦鹉热衣原体的生物特性以及感染的宿主范围;鹦鹉热衣原体减毒活疫苗温度敏感株的培育及致病机理的研究;灭活疫苗灭活条件的研究,最佳免疫量,不同免疫途径的研究和我国对绵羊和猪鹦鹉热衣原体灭活疫苗的研究;以及鹦鹉热衣原体主要外膜蛋白基因工程亚单位疫苗和禽衣原体DNA疫苗的研究情况。     

山东省部分地区家禽鹦鹉热衣原体感染情况调查

为了解山东地区家禽鹦鹉热衣原体(Cps)感染现状,本研究采用间接血凝法(IHA)对2013年~2014年采集自山东潍坊、淄博、济南、临沂、烟台等地区的1 020份鸡、鸭血清样品进行Cps抗体的检测,并对检测数据进行了统计分析;结果显示:IHA测得总阳性率为29.51%(301/1 020);鸡阳性率为25.26%(197/780);鸭阳性率为43.33%(104/240);各地区间阳性率存在一定差异。本调查结果表明,家禽Cps感染在山东地区具有较高的感染率。     

1株鹦鹉热衣原体的冻干、鉴定和分子分型研究

从国家兽医微生物菌(毒)种保藏中心领取1支鹦鹉热衣原体CVCC2410进行冻干和鉴定。将衣原体经卵黄囊接种SPF鸡胚进行复壮和传代,收获卵黄囊膜研磨后分装、冻干,并对冻干衣原体进行了无菌检验、真空度测定、染色特性和形态观察、特异性检验、16S rRNA鉴定、分子分型、衣原体含量测定。结果表明冻干衣原体CVCC2410 2203为C型鹦鹉热衣原体,含量为10^5.50ELD₅₀/0.2mL。本研究完善了该株衣原体的信息,为制定鹦鹉热衣原体的入库标准奠定基础。     

Chlamydia psittaci: a suspected cause of reproductive loss in three Victorian horses

Chlamydia psittaci was detected by PCR in the lung and equine foetal membranes of two aborted equine foetuses and one weak foal from two different studs in Victoria, Australia. The abortions occurred in September 2019 in two mares sharing a paddock northeast of Melbourne. The weak foal was born in October 2019 in a similar geographical region and died soon after birth despite receiving veterinary care. The detection of C. psittaci DNA in the lung and equine foetal membranes of the aborted or weak foals and the absence of any other factors that are commonly associated with abortion or neonatal death suggest that this pathogen may be the cause of the reproductive loss. The detection of C. psittaci in these cases is consistent with the recent detection of C. psittaci in association with equine abortion in New South Wales. These cases in Victoria show that C. psittaci, and the zoonotic risk it poses, should be considered in association with equine reproductive loss in other areas of Australia.     

北京市及周边省份家禽鹦鹉热嗜性衣原体流行状况的调查

对北京市周边6省份怀疑感染鹦鹉热嗜性衣原体的鸡鸭血清样品374份、病料81份，分别使用IHA诊断试剂盒、ELISA试剂盒以及抗酸染色试剂和荧光抗体诊断试剂进行了检查，以评价北京市及其周边地区家禽鹦鹉热嗜性衣原体的流行性。结果，上述4种试剂检测出的阳性率依次为24．9％、77．9%、18．5%和38．2％；北京市10份SPF鸡血清的抗体全部为阳性；患病肉鸡、肉鸭气囊样品，蛋鸡输卵管样品的检出率较高。表明，北京市及其周边地区家禽已经感染了鹦鹉热嗜性衣原体，酶联免疫吸附法和荧光抗体染色法能分别提高抗体和抗原的检出率。     

Infection of the Bovine Female Genital Tract with Chlamydia psittaci as a Possible Cause of Infertility

Contents: Chlamydia psittaci was isolated in embryonated chicken eggs via the yolk sac route from ten (16,7%) vaginal andlor endometrial mucosal scrapings of 60 slaughter cows. Simultaneous fecal shedding of chlamydiae was found in four animals. Chlamydial infections of the genital tract were frequent (p < 0,01) when there were endometrial inflammatory lesions together with the failure to detect other bacterial pathogens in the uterus.     

布鲁氏菌和鹦鹉热衣原体双重PCR检测方法的建立与应用

为建立同时检测布鲁氏菌和鹦鹉热衣原体的双重PCR方法,本研究据GenBank上已发表的具有属间特异性的布鲁氏菌bp26基因和鹦鹉热衣原体23S rRNA基因,利用 Primer Premier 5.0软件各设计1对特异性引物,扩增的目的片段长度分别为219和356 bp。通过优化反应条件,建立了能同时检测布鲁氏菌和鹦鹉热衣原体的双重PCR方法。该方法具有较好的特异性和可重复性,对2种基因单重PCR检测敏感性均达到3.1×10²拷贝/反应,双重检测的灵敏度为3.1×10³拷贝/反应。利用该双重PCR方法对流产牛抗凝全血、血清、流产胎儿及奶液共172份临床疑似布鲁氏菌感染的样品进行检测,检测到布鲁氏菌阳性样品53份,鹦鹉热衣原体阳性样品2份,以上这2种病原的阳性检出率分别为30.8%和1.2%,且检测到2种病原混合感染的阳性样品2份,阳性检出率为1.2%。临床应用结果表明,该方法可用来对布鲁氏菌和鹦鹉热衣原体进行同步、快速、灵敏的检测。     

First detection of Chlamydia psittaci from a wild native passerine bird in New Zealand

AIMS: To undertake disease surveillance for Chlamydia psittaci in native birds as part of a pilot study to examine pathogen diversity on Hauturu-o-Toi/Little Barrier Island. To retrospectively review the Massey University post-mortem database to determine previous cases of avian chlamydiosis in New Zealand.

METHODS:Mistnetting of forest birds was conducted across an elevational gradient on Hauturu-o-Toi/Little Barrier Island. Minitip culture swabs were used to collect cloacal samples from native birds. These swabs were screened for Chlamydia family DNA using two PCR methods. Positive results were sequenced. A retrospective review of the Massey University post-mortem database of all avian cases from 1990 to 2011 was conducted.

RESULTS:Ten native birds including four bellbirds (Anthornis melanura), three rifleman (Acanthisitta chloris), two hihi (Notiomyces cincta), and one whitehead (Mohoua albicilla) were sampled and one otherwise healthy female hihi was positive by both PCR screening methods for Chlamydophila. Sequencing confirmed 99–100% genetic similarity to C. psittaci. A retrospective review of the Massey University post-mortem database revealed no previous diagnoses of avian chlamydiosis in wild native New Zealand birds although it has been detected in captive parrots, and wild and captive exotic pigeons.

CONCLUSIONS:This is the first report of the detection of C. psittaci from a wild native bird in New Zealand. The bird was a Passeriforme from an endangered species that was captured free-living on Little Barrier Island. The incidence of avian chlamydiosis in native birds in New Zealand appears to be very low, based on the retrospective review of the post-mortem database.

CLINICAL RELEVANCE: It is unlikely that avian chlamydiosis is a significant problem for hihi population health. The detection of this organism has greater significance for other more susceptible species on Little Barrier Island and for human health, particularly for conservation workers involved in wildlife translocations. It further suggests that passerine birds may be a reservoir for C. psittaci in New Zealand ecosystems.     

Comparison of the omp I gene of Chlamydia psittaci between isolates in Victorian koalas and other animal species

Objective The objective of this study is to compare the strain of chlamydia causing genital infection in koalas from Victoria with isolates from other animal species.
Design Polymerase chain reaction and restriction enzyme analysis has been used to compare various Chlamydia psittaci isolates from a range of animals and disease syndromes. The isolates used in this study include isolates from three birds, three from aborted sheep, one from polyarthritis, one from bovine abortion, one from feline pneumonitis, three porcine isolates from faeces, polyarthritis and abortion, and three urogenital isolates from Victorian koalas.
Procedure Two polymerase chain reactions were performed, each amplifying a different region of the omp I gene. The first polymerase chain reaction amplified a 144 bp segment of the gene which was then digested with the restriction enzyme Eco R I. The second polymerase chain reaction amplified a larger 1070 bp region of the omp I gene which was digested with two restriction enzymes Alu I and Nde II.
Results and conclusions The results obtained have confirmed that variation in DNA sequence of various animal chlamydia isolates does occur. They have also shown that it is possible to classify isolates, based on their restriction enzyme profiles, into distinct groups.     

Methods of detection of Chlamydia psittaci in domesticated and wild birds

OBJECTIVE: To study the occurrence of Chlamydia psittaci in domesticated and wild birds and compare the sensitivity of molecular detection with cell culture isolation. DESIGN: Study of cell culture isolation and PCR detection of C psittaci in avian samples. PROCEDURE: Samples were obtained from 485 birds. Domesticated birds were selected at random from pet shops, private aviaries and zoos, while wild birds were captured locally, sampled, and immediately released. Swabs were collected from choanal slit, conjunctiva and cloaca of each bird and pooled. Samples were divided into equal portions for use in PCR dot-blot and cell culture detection. PCR and dot-blot detection was based on the ompB gene. RESULTS: Prevalence of infection varied markedly between flocks of captive birds. It was highest where there were frequent changes in the flock members or where there were many birds confined in small areas. C psittaci was not detected in wild birds or water birds. The sensitivity of cell culture compared to PCR dot-blot detection was 68%. All samples positive by cell culture were also positive by PCR. CONCLUSIONS: PCR-dot blot detection of C psittaci in birds appears to be more sensitive than cell culture isolation in this study. C psittaci infection of birds may occur in clinically normal captive birds.     

Chlamydophila psittaci infections in humans during an outbreak of psittacosis from poultry in Germany

In 2005, an outbreak of severe respiratory disease in a mixed poultry flock that was infected with Chlamydophila (C.) psittaci led to dissemination of the infection to at least 100 small poultry farms in 11 districts of Central Germany. At the same time, a total of 24 persons in contact with poultry from one of the flocks reported flu-like symptoms to their physician, thus suggesting zoonotic transmission. Within 3 weeks, seven individuals had to be hospitalized, with three of them requiring intensive care. Analysis of ompA sequences from chlamydial isolates and directly from clinical samples revealed the presence of both genotype A and E/B of C. psittaci at the source of the outbreak and in contact flocks. Genotype A was also detected in the three severely ill patients. The findings of the present study demonstrate the high zoonotic potential of avian chlamydiae. To ensure speedy eradication of psittacosis in poultry flocks and effective treatment of infected humans, fast, sensitive and species-specific detection of the causative agent is essential, as well as close collaboration between regional public health services, attending physicians and the diagnostic laboratories involved.     

动/物/的/鹦/鹉/热/衣/原/体/调/查

本文报道从１９８０年～１９８９年１０年中,在新疆阿勒泰、阿克苏、巴州等１５个地州市、８６个县、４２６个乡（镇）、１５个种畜场的１９种动物和１２７８９９头（只）动物鹦鹉热衣原体病调查,并通过衣原体、弓形体和布鲁氏菌病对比调查说明,新疆布病控制后,衣原体是引起山、绵羊流产的主要原因之一。并从阿克苏、塔城等四个地州的山、绵羊流产胎儿中成功在分离出１９株鹦鹉热衣原体,鉴定方法采用电镜、药敏试验、鸡胚毒力试     

Seroprevalence of Antibodies to Chlamydophila psittaci in Zoo Workers in Brazil

To evaluate the prevalence of antibodies to Chlamydophila psittaci 364 serum samples were collected from veterinarians, biologists, animal scientists, veterinary students, animal keepers and others employees in 20 zoos, and from veterinary practitioners in 10 Brazilian states. Subjects ranged from 15 to 64 years of age, with 268 (74%) males and 96 (26%) females. Chlamydial antibodies were determined by the complement fixation test (CFT) and specific anti‐C. psittaci IgG antibodies were determined by the microimmunoflurescence (MIF) test. Complement fixation test showed 23.9% (87/364) and MIF test showed 4.7% (17/364) positive serum samples. Titres ranged from 16 to 256 in both assays, demonstrating evidence of recent or current infection. Although chlamydial antibodies were detected in workers of seventeen zoos, MIF test only detected specific C. psittaci antibodies in seven of them. Previous psittacosis infection was suspected in eight workers of two zoos, five of whom reported having pneumonia, while employed at the zoos. However, diagnosis was not established in any of these cases in the past. Results indicated the occurrence of infection and previous contact of Brazilian zoo workers with C. psittaci, as well as the zoonotic potential of psittacosis in this risk population. Other studies are necessary to evaluate the risk factors of infection in this population. This seroepidemiological survey confirmed the need to adopt preventive measures to control avian chlamydiosis and protect the health of zoo workers in the country.     

10种预测的鹦鹉热衣原体Ⅲ型分泌系统效应蛋白的定位分析

旨在分析预测的10种鹦鹉热衣原体（Chlamydia psittaci，Cps）Ⅲ型分泌系统效应蛋白在细胞内的定位，为下一步研究其结构与功能奠定基础。本研究利用计算机辅助的生物信息学方法（Effective T3和BPBAac tool）预测Cps T₃SS效应蛋白。利用分子克隆技术构建重组质粒pGEX-6P-1/目的基因，诱导表达、纯化相应重组蛋白。皮下免疫BALB/c小鼠，制备各重组蛋白的多克隆抗体，ELISA测定其效价。以制备的多克隆抗体为一抗，应用间接免疫荧光法检测假定的效应蛋白在感染细胞中的定位。本研究通过应用Effective T3、BPBAac tool交叉验证，预测了14个Cps T₃SS效应蛋白编码基因：CPSIT_0357、CPSIT_0429、CPSIT_0461、CPSIT_0463、CPSIT_0490、CPSIT_0594、CPSIT_0785、CPSIT_0844、CPSIT_0846、CPSIT_0019、CPSIT_0020、CPSIT_0968、CPSIT_0969、CPSIT_0942，并进行分析。构建了14个靶基因的重组质粒，经IPTG诱导，除CPSIT_0357、CPSIT_0429、CPSIT_0968、CPSIT_0969外，其他10个重组菌分别表达了相对分子质量（M_r）与预测M_r相近的可溶性蛋白。经GST树脂纯化后，将重组蛋白免疫雌性BALB/c小鼠，血清抗体效价为（1：32 000）~（1：64 000）。IFA细胞内定位观察，发现CPSIT_0844和CPSIT_0846位于包涵体膜，而CPSIT_0461、CPSIT_0463、CPSIT_0490、CPSIT_0594、CPSIT_0785、CPSIT_0019、CPSIT_0020、CPSIT_0942位于包涵体内。本研究鉴定了2种定位于衣原体包涵体膜的效应蛋白和8种定位于衣原体包涵体中的效应蛋白。     

新型滴鼻灭活鹦鹉热衣原体疫苗的免疫效力评价

【目的】探究由霍乱弧菌菌影(Vibrio cholerae ghosts, VCG)和纳米壳聚糖凝胶(Gel)组合制作的一种新型灭活衣原体疫苗复合佐剂是否可增强鹦鹉热衣原体原体(elementary body, EB)灭活抗原诱导动物机体产生的免疫应答。【方法】将60只7日龄SPF鸡随机分为4个组,分别为：EB+VCG+Gel组、EB+VCG组、Gel组和EB组。通过滴鼻途径免疫鸡群,免疫2次,每次间隔14 d,免疫后检测鸡血清中IgG抗体水平、淋巴细胞增殖指数、CD4⁺/CD8⁺T细胞比例、细胞因子(白介素-4(IL-4)、IL-10、IL-12和干扰素-γ(IFN-γ))含量及攻毒后鸡喉头排菌量和肺脏的病理损伤程度。【结果】与Gel和EB组相比,EB+VCG+Gel和EB+VCG组IgG抗体水平、淋巴细胞增殖指数、IFN-γ含量和CD4⁺/CD8⁺T细胞比值显著或极显著升高(P<0.05;P<0.01)。此外,与Gel和EB组相比,攻毒后第12天,EB+VCG+Gel组喉头排菌量极显...     

猪鹦鹉热衣原体流产灭活疫苗试生产试验

按照猪鹦鹉热衣原体流产灭活疫苗制造和检验试行规程要求,在GMP生产车间进行鸡胚的孵化、衣原体的繁殖、衣原体灭活及疫苗乳化,共试生产三批猪鹦鹉热衣原体流产灭活疫苗,每批80万毫升。结果显示这三批疫苗生产中的中间品和成品的标准均符合猪鹦鹉热衣原体流产灭活疫苗制造和检验试行规程。