羊布鲁氏菌病4种血清学检测方法的临床应用比较

目的荧光偏振试验(FPA)、间接ELISA方法 (IELISA)、试管凝集试验(SAT)、虎红平板凝集试验(RBT)4种临床检测方法检测内蒙古地区羊布鲁氏菌病。方法 FPA方法、IELISA方法、SAT法及RBT法同时检测在内蒙古各地区随机采集的2 727份羊血清样本。结果 FPA、IELISA、SAT及RBT法对2 727份羊血清的阳性检出率分别为7.59%、7.45%、6.31%和6.20%。集约化养殖场的阳性检出率为4.80%,散户养殖场的9.41%,内蒙古地区依旧存在布鲁氏菌病的威胁。结论 FPA法检测羊布鲁氏菌病适合于临床诊断,在田间也能快速准确的诊断布鲁氏菌病。     

204例布氏杆菌病实验室检测结果分析

目的通过实验室检测,发现布氏杆菌感染者,掌握疫情动态,为预测布氏杆菌病流行趋势、制订防治对策提供依据。方法根据布氏杆菌病实验室诊断标准,利用虎红平板凝集试验(RBPT)和试管凝集试验(SAT)相结合的试验方法,对布氏杆菌病疑似病例和有接触史等的人员进行布氏杆菌抗体检测。结果在470例监测对象中,布氏杆菌抗体阳性病例204例,阳性率43.4%。在阳性病例中,男女性别比为1.62∶1;40~69岁年龄组占75.49%;畜牧养殖人员占40.69%,屠宰作业者占32.35%。结论应加强对养殖业和畜牧业人员的预防控制工作,提高其自我防护意识。     

大熊猫布鲁氏菌病、弓形虫病以及心丝虫病的血清学调查研究

为摸清大熊猫布鲁氏菌病、弓形虫病以及心丝虫病的血清学感染资料,采用虎红平板凝集试验、试管凝集试验以及外膜蛋白BCSP3基因PCR扩增检测布鲁氏菌病;同时采用弓形虫间接血凝试验和犬心丝虫抗原快速诊断对样品进行检测。结果表明,6只大熊猫RBPT检测为阳性,进一步采用SAT和血液细菌BCSP31基因PCR扩增结果均为阴性,排除了布鲁氏菌感染;大熊猫蜀兰弓形虫抗体检测呈阳性,复查也为阳性,表明存在弓形虫感染;所有大熊猫心丝虫抗原检测结果均为阴性,表明无心丝虫感染。本次检测的3种疾病中,仅发现1只熊猫(蜀兰)存在弓形虫感染,布鲁氏菌病和心丝虫病均为阴性,表明目前成都大熊猫繁育研究基地内大熊猫疾病的预防工作成果显著。     

四种检测沙眼衣原体方法的诊断符合率分析

目的评价直接免疫荧光染色(DFA)、聚合酶链式反应(PCR)、酶联免疫吸附试验(ELISA)和金标免疫层析试验(GICA)4种方法检测沙眼衣原体(Chlamydia trachomatis,CT)的实验诊断符合率。方法同时采用DFA、PCR、ELISA和GICA4种方法检测临床泌尿生殖道样本CT感染标志物。2种及2种以上方法结果阳性判为真阳性,仅有1种方法结果阳性或全部方法结果阴性则判为真阴性。分别计算4种法的灵敏度、特异性、阳性预测值和阴性预测值、ROC曲线下面积以及各方法之间的Kappa值。结果 223份临床泌尿生殖道标本中阳性32例,阴性191例。DFA、PCR、ELISA、GICA 4种方法的敏感度依次为93.7%、96.8%、90.6%和28.1%,特异性为98.9%、98.9%、98.4%和100%,阳性预测值为93.7%、93.9%、90.6%和100%,阴性预测值为98.9%、99.4%、98.4%和89.2%;ROC曲线下面积为0.964、0.964、0.945和0.641。前3种方法之间Kappa值0.82~0.95,与GI-CA法之间Kappa值均小于0.5。结论 DFA法、ELISA法、PCR法作为临床CT感染的实验室检测是较理想的方法;GICA法因灵敏度低且ROC曲线下面积仅0.641,作为实验诊断方法不太理想。     

布鲁菌omp19原核表达和间接酶联免疫吸附试验检测布鲁菌方法的建立

本研究原核表达并纯化了布鲁菌脂蛋白Omp19,并通过蛋白免疫印迹法对其免疫原性进行验证。采用Omp19作为包被抗原,建立检测羊种布鲁菌的间接酶联免疫吸附试验(iELISA),检测90份羊血清样本,并与标准血清凝集试验(SAT)比较,用SPSS软件进行数据分析。结果显示,iELISA与SAT的阳性符合率为95.12%,阴性符合率为67.35%,Kappa值为0.607 7。对2种方法进行McNemar卡方检验,结果有显著性差异(P0.05)。本研究建立的iELISA与SAT的总符合率达80%,可作为羊种布鲁菌病血清学诊断的备用方法。     

明胶颗粒凝集试验联合ELISA法在梅毒检测中的应用价值

目的分析明胶颗粒凝集试验(TPPA)联合酶联免疫吸附试验(ELISA)在梅毒检测中的应用价值。方法选择我院2015年1月至2018年12月收治的128例疑似梅毒患者为研究对象,将其中43例临床确诊患者纳入梅毒组,余下85例为非梅毒组,全部受试者均接受ELISA法联合TPPA检测,比较单独ELISA法和ELISA联合TPPA检测的结果。结果梅毒组患者中ELISA法诊断阳性35例,阴性8例;ELISA联合TPPA诊断阳性42例,阴性1例;ELISA法假阳性率14.63%(6/41),明显高于ELISA联合TPPA法的2.33%(1/43),差异有统计学意义(χ~2=4.163,P=0.041)。ELISA联合TPPA诊断梅毒的特异性、敏感性、阴性预测值及阳性预测值分别为98.82%、97.67%、98.82%、97.67%,均显著高于单独ELISA检测的92.94%、81.40%、90.80%、85.37%,差异均有统计学意义(χ~2=143.650、6.081、5.575、4.163,P0.001、=0.014、=0.018、=0.041)。结论 ELISA适用于大批量梅毒筛查,TPPA法的特异性及敏感性高,可作为梅毒血清抗体检测的确认试验。二者联合检测能够提高梅毒诊断效能,具有较高的应用价值。     

布氏菌病血清学筛选试验的比较

<正> 试管凝集试验(TAT)已成为布氏菌病血清学诊断的标准方法。是公认的关于布病血清学的定量试验。虽然TAT是标准方法,但因其繁琐,所以不适用于布病大量标本的初步试验。TAT对于现场或用于流行病学调查不方便。已报导了几种快速筛选试验,包括玻片凝集试验(TAT),酸性平板抗原凝集试验(CAT)和微量凝集试验(MAT)。在报告中,对SAT、MAT和CAT的敏感性和特异性、TAT真阳性和真阴性的结果界限进行了严格测定。     

牛布鲁氏菌病阳性血清国家标准品的研制

为制备标定凝集试验用的牛布鲁氏菌病阳性血清国家标准品,采集4份田间自然感染牛布鲁氏菌病的阳性血清作为候选物,经过筛选后,选择了3#血清用于标准品制备。对该标准品进行了物理性状、无菌检验、真空度测定、剩余水分检验、均匀性检测和稳定性试验,检测结果均符合要求。以国际标准品为参比,测定了该标准品的RBT、SAT和CFT效价,结果分别为1∶160“+”、1∶2 400“++”、1∶800“++”,与协作标定结果完全一致。该标准品以国际标准品溯源的国际单位含量为4 000 IU/mL。     

电化学发光法检测梅毒螺旋体特异性抗体的应用研究

目的对罗氏电化学发光免疫法(ECLIA)检测梅毒螺旋体特异性抗体的临床价值进行评估。方法收集梅毒疑似病例血清标本132份,分别用ECLIA、梅毒螺旋体明胶凝集试验(TPPA)和免疫印迹法(WB)进行检测,以WB为金标准,计算并比较ECLIA和TPPA的灵敏度和特异性,进而比较化学发光免疫法检测低S/CO值和高S/CO值的灵敏度和特异性差异。结果针对132份血清标本,ECLIA敏感性为100.00%,特异性为83.33%,阳性预测值为96.43%,阴性预测值为100.00%,总符合率为96.97%。TPPA敏感性为93.52%,特异性为87.50%,阳性预测值为97.12%,阴性预测值为75.00%,总符合率为92.42%。ECLIA检测1≤S/CO3组与S/CO≥3组的敏感性均为100.00%,特异性分别为86.96%和95.24%,结论 ECLIA检测具有较高的敏感性,适合临床大样本筛查,对S/CO值低的标本应结合TPPA、WB及临床资料确诊。     

HPV L1 壳蛋白与hTERC 基因联合检测诊断新疆维、汉妇女宫颈病变CIN2+的临床价值

目的:评价HPV L1壳蛋白与h TERC基因联合检测诊断新疆维、汉妇女宫颈病变CIN2~+的临床价值。方法:选择HPV感染阳性或TCT阳性或两项同时阳性的新疆维、汉妇女465例纳入研究。所有研究病例均行阴道镜下活检送病理组织学检查,并通过免疫细胞化学法检测HPV L1蛋白的表达、通过F1SH技术检测h TERC基因的表达。通过诊断试验评价与接收者工作特征曲线评价其联合检测诊断新疆维、汉妇女宫颈病变CIN2~+的价值。结果:HPV L1壳蛋白与h TERC基因联合检测用于区分CIN2~+与正常或慢性炎症患者及CIN1患者时,其诊断CIN2~+的ROC曲线下面积为0.5左右(维吾尔族0.577,汉族0.572);用于区分CIN2~+与CIN1患者时,其ROC曲线下面积可达0.7左右(维吾尔族0.706,汉族0.699)。此时,其诊断新疆维族妇女CIN 2~+的灵敏度、特异度、正确率、假阳性率、假阴性率、阳性预测价值和阴性预测价值分别为92.37%、48.84%、81.61%、15.38%、32.26%、84.62%和67.74%;诊断汉族妇女CIN2~+的灵敏度、特异度、正确率、假阳性率、假阴性率、阳性预测价值和阴性预测价值分别为93.69%、46.15%、81.33%、16.80%、28.00%、83.20%和72.00%。维、汉之间的灵敏度、特异度、正确率、假阳性率、假阴性率、阳性预测价值和阴性预测价值均无显著差异(P0.05)。结论:HPV L1壳蛋白与h TERC基因联合检测诊断维、汉妇女CIN 2~+时,其适用人群为具有CIN的患者。此时其诊断维、汉族妇女宫颈高度病变CIN 2~+的灵敏度、正确率和阳性预测价值较高,特异度和阴性预测价值较低,且均不存在民族差异。     

A serological and bacteriological survey of brucellosis in wild boar (Sus scrofa) in Belgium

ABSTRACT: BACKGROUND: Brucellosis is frequently reported among wild boar populations in Europe. The aim of the study was to assess the epidemiological situation in Belgium, regarding the steady increase of wild boar populations over the last decades. Several serological tests were used and compared with culture and IS711 polymerase chain reaction (PCR), to determine the most suitable combination of diagnostic tools for conducting a successful prevalence study in wildlife. RESULTS: An indirect enzyme-linked immunosorbent assay (iELISA) was used on 1168 sera from hunter-killed wild boar sampled between 2003 and 2007 in 4 natural regions of southern Belgium. Results gave an apparent prevalence of 54.88% (95% CI 52.03-57.73). Prevalence was significantly affected by age and by the year of study, but not by sex nor by the region of sampling. The relative sensitivities of the complement fixation test (CFT), the Rose Bengal test (RBT), and the slow agglutination test (SAT) versus the iELISA differed widely between tests, reaching 62.67%, 46.68%, and 34.77%, respectively. The relative specificities of the CFT, RBT and SAT versus the iELISA were respectively 99.01%, 92.49%, and 99.1%. From seropositive animals (iELISA), 9% were positive by culture and 24% by PCR when testing spleen and/or tonsils. Sensitivity of the PCR was higher on tonsils than on spleen. All bacterial isolates were identified as Brucella suis biovar 2. CONCLUSIONS: Brucellosis is widespread among wild boar in southern Belgium, with seroprevalences having increased over ten years, and constitutes a growing risk of spillback to outdoor-farmed pig herds. The iELISA showed a better sensitivity than the CFT, RBT and SAT. Serological tests must be associated with direct diagnosis and PCR proved more sensitive than culture under wildlife sampling conditions. Spleen and tonsils are lymphoid tissues usually sampled in multi-disease monitoring programs. They remain top-grade organs for direct diagnosis of brucellosis, with a preference for tonsils.     

The Rose Bengal Test in human brucellosis: a neglected test for the diagnosis of a neglected disease

Brucellosis is a highly contagious zoonosis affecting livestock and human beings. The human disease lacks pathognomonic symptoms and laboratory tests are essential for its diagnosis. However, most tests are difficult to implement in the areas and countries were brucellosis is endemic. Here, we compared the simple and cheap Rose Bengal Test (RBT) with serum agglutination, Coombs, competitive ELISA, Brucellacapt, lateral flow immunochromatography for IgM and IgG detection and immunoprecipitation with Brucella proteins. We tested 208 sera from patients with brucellosis proved by bacteriological isolation, 20 contacts with no brucellosis, and 1559 sera of persons with no recent contact or brucellosis symptoms. RBT was highly sensitive in acute and long evolution brucellosis cases and this related to its ability to detect IgM, IgG and IgA, to the absence of prozones, and to the agglutinating activity of blocking IgA at the pH of the test. RBT was also highly specific in the sera of persons with no contact with Brucella. No test in this study outperformed RBT, and none was fully satisfactory in distinguishing contacts from infected patients. When modified to test serum dilutions, a diagnostic titer >4 in RBT resulted in 87.4% sensitivity (infected patients) and 100% specificity (contacts). We discuss the limitations of serological tests in the diagnosis of human brucellosis, particularly in the more chronic forms, and conclude that simplicity and affordability of RBT make it close to the ideal test for small and understaffed hospitals and laboratories.     

A simple enzyme immunoassay for detecting brucellosis antibodies

Abstract A simple enzyme immunoassay was developed and evaluated for serological diagnosis of brucellosis in 25 patients with various forms of brucellosis and 292 control patients with other conditions and disorders. All brucellosis patients gave a positive test with the initial sample. In 3 acute, febrile brucellosis patients with follow-up sera taken during therapy a sharp drop in specific antibody was noted. There was a less pronounced antibody reduction in 1 chronic and 2 relapse patients and an antibody increase in 1 chronic and 1 relapse case. All control samples gave negative results. In addition, the assay was evaluated as a screening test with 315 sera from 'healthy' individuals living in a brucellosis focus and representing 15% of that population. 11.5% (34/293) of subjects with no reported history of brucellosis and 45% (10/22) of cases treated in the past gave a positive test result. The agreement in those samples between the assay and the serum agglutination test was 95.5%.     

Comparison of diagnostic tests for the detection of <Emphasis Type="Italic">Brucella</Emphasis> spp. in camel sera

Background

Brucellosis in livestock causes enormous losses for economies of developing countries and poses a severe health risk to consumers of dairy products. Little information is known especially on camel brucellosis and its impact on human health. For surveillance and control of the disease, sensitive and reliable detection methods are needed. Although serological tests are the mainstay of diagnosis in camel brucellosis, these tests have been directly transposed from cattle without adequate validation. To date, little information on application of real-time PCR for detection of Brucella in camel serum is available. Therefore, this study was performed to compare the diagnostic efficiency of different serological tests and real-time PCR in order to identify the most sensitive, rapid and simple combination of tests for detecting Brucella infection in camels.

Findings

A total of 895 serum samples collected from apparently healthy Sudanese camels was investigated. Sudan is a well documented endemic region for brucellosis with cases in humans, ruminants, and camels. Rose Bengal Test (RBT), Complement Fixation Test (CFT), Slow Agglutination Test (SAT), Competitive Enzyme Linked Immunosorbant Assay (cELISA) and Fluorescence Polarization Assay (FPA) as well as real-time PCR were used. Our findings revealed that bcsp31 kDa real-time PCR detected Brucella DNA in 84.8% (759/895) of the examined samples, of which 15.5% (118/759) were serologically negative. Our results show no relevant difference in sensitivity between the different serological tests. FPA detected the highest number of positive cases (79.3%) followed by CFT (71.4%), RBT (70.7%), SAT (70.6%) and cELISA (68.8%). A combination of real-time PCR with one of the used serological tests identified brucellosis in more than 99% of the infected animals. 59.7% of the examined samples were positive in all serological tests and real-time PCR. A subpopulation of 6.8% of animals was positive in all serological tests but negative in real-time PCR assays. The high percentage of positive cases in this study does not necessarily reflect the seroprevalence of the disease in the country but might be caused by the fact that the camels were imported from brucellosis infected herds of Sudan, accidentally. Seroprevalence of brucellosis in camels should be examined in confirmatory studies to evaluate the importance of brucellosis in this animal species.

Conclusion

We suggest combining bcsp31 real-time PCR with either FPA, CFT, RBT or SAT to screen camels for brucellosis.

     

Prozone effects in microscopic agglutination tests for leptospirosis in the sera of mice infected with the pathogenic Leptospira interrogans serovar Canicola

Mice experimentally infected with a pathogenic strain of Leptospira interrogans serovar Canicola produced false negative results (prozone effect) in a microscopic agglutination test (MAT). This prozone effect occurred in several serum samples collected at different post-infection times, but it was more prominent in samples collected from seven-42 days post-infection and for 1:50 and 1:100 sample dilutions. This phenomenon was correlated with increased antibody titres in the early post-infection phase. While prozone effects are often observed in serological agglutination assays for the diagnosis of animal brucellosis and human syphilis, they are not widely reported in leptospirosis MATs.     

A seroprevalence survey of Toxoplasma gondii in common wombats (Vombatus ursinus)

Serum was collected from 23 wild common wombats (Vombatus ursinus) on a pastoral property in the Southern Tablelands of New South Wales, Australia (231N 616E) between 3rd August 2001 and 25th March 2002. The serum was tested using three serological methods for antibodies to Toxoplasma gondii. Six animals (26.1%) were shown to have antibodies to T. gondii. The latex agglutination test proved to be less sensitive than the direct agglutination test or the modified agglutination test. This is the first serological survey of T. gondii in wombats. This is the first recorded use of the latex agglutination test on wombat serum. This study demonstrated the highest percentage of seropositive animals in any serological survey for T. gondii in marsupials.     

Laboratory diagnosis of brucellosis in a rural endemic area in northeastern Spain.

Sera obtained from 62 patients from four mountain counties in Catalonia (Northeastern Spain), in whom brucellosis had been diagnosed on the basis of clinical evidence and/or personal history, were analyzed using the rose Bengal test, standard serum agglutination test (SAT), Coombs' test, ELISA, and complement fixation. The diagnosis was further confirmed through blood cultures. Clinical evidence, epidemiology, and the results from serologic tests were used to assign patients to one of two groups: group 1 (n = 38) patients had primary infections, whereas group 2 (n = 24) patients had been previously exposed to the microorganism, i.e. re-infection of group 2 individuals occurred after long periods of time during which no active infection by Brucella had been detected. Receiving-operating charts (ROC) were used to determine the diagnostic value of the different tests and to establish discriminant values. Blood culture was a valuable diagnostic tool in group 1 (0.92 sensitivity) but was inappropriate in group 2 (0.08). The combination of positive rose Bengal test and agglutination >/=1/160 was valid for diagnosis in group 1. In group 2, agglutination <1/160 (including negative agglutination) did not rule out brucellosis. The combination of positive rose Bengal test and Coombs' test >/=1/320 was the best diagnostic criterion (0.8 specificity; 1 sensitivity). ELISA (for IgG, IgM, or both) did not improve diagnostic accuracy.     

Comparative evaluation of recombinant BP26 protein for serological diagnosis of Brucella melitensis infection in goats

An enzyme-linked immunosorbent assay (ELISA) was developed for the serological diagnosis of caprine brucellosis with purified recombinant BP26 (outer membrane protein 28) of Brucella melitensis 16M produced in Escherichia coli. Brucella protein named CP28, BP26, or Omp28 has been identified as an immunodominant antigen in infected cattle, sheep, goats, and humans. The recombinant BP26 (rBP6) ELISA performed with serum samples (n = 1738) taken from organized farms and field goats from Northern India and tested in two different batches of 778 and 960 animals for brucellosis. Positive results were compared with the traditional serum agglutination test (SAT), complement fixation test (CFT) and dot-ELISA. The diagnostic sensitivity of rBP26 ELISA, SAT, CFT and dot-ELISA was 87.5%, 56.25%, 62.5% and 75% respectively. The specificity of the rBP26 ELISA, SAT, CFT and dot-ELISA was 90%, 75%, 80%, and 85% respectively. The results of rBP26 ELISA positive animals were further compared and evaluated by tissue PCR using B. melitensis BP26 gene as target. This resulted in 100% positive correlation between rBP26 ELISA and BP26 PCR. Thus, these results confirmed the importance of BP26 as a suitable antigen and rBP26 ELISA, if well standardized, proved to be a good screening test for the serological diagnosis of caprine brucellosis.     

血清PCT、CRP及IL-6联合检测诊断细菌性血流感染的临床价值分析

目的:探讨血清降钙素原(PCT)、C反应蛋白(CRP)及白介素-6(IL-6)联合检测诊断细菌性血流感染(BSI)的临床价值。方法:选取我院2015年8月到2016年10月收治的疑似细菌性BSI患者216例,入院后均送检血培养,根据培养结果将其分为阳性组(102例)和阴性组(114例)。统计细菌性BSI阳性率、革兰阳性菌感染率和革兰阴性菌感染率;检测血清PCT、CRP、IL-6水平,并比较两组患者的差异,同时绘制ROC曲线并计算出各指标及联合检测的灵敏度、特异度、阳性预测值、阴性预测值及约登指数值。结果:所有疑似BSI患者的细菌阳性检出率为47.22%,革兰阳性菌感染率与革兰阴性菌感染率对比无差异(P0.05);阳性组的血清PCT、CRP、IL-6水平均明显高于阴性组(P0.05);血清IL-6的AUC明显大于PCT和CRP(P0.05);PCT、CRP及IL-6联合检测的灵敏度、特异度、阳性预测值、阴性预测值及约登指数均明显高于单项检测(P0.05)。结论:血清PCT、CRP及IL-6对于BSI均有着一定诊断价值,而各指标联合检测诊断BSI的临床价值更高。     

炭凝集试验快速检测呼吸道合胞病毒的研究

用炭凝集试验（ＣＡＴ）检测呼吸道合胞病毒（ＲＳＶ）,结果表明该法是一种简便、快速、特异的诊断方法。用ＣＡＴ对１６株ＲＳＶ和８株其它病毒做试验,结果仅ＲＳＶ凝集,而其它病毒均阴性。用该法与细胞培养法检测８３份临床呼吸道感染幼儿鼻咽吸出物,结果ＣＡＴ法阳性率为６９．８８％（５３／８３）,细胞培养法为３９．７５％（３３／８３）,两者阳性检出率相差极显著。阻断试验证明ＣＡＴ是高度特异的。结果证明ＣＡＴ具有较高的敏感性与特异性,可用于临床ＲＳＶ标本的快速检测。