Parasitological diagnosis combining an internally controlled real-time PCR assay for the detection of four protozoa in stool samples with a testing algorithm for microscopy

Molecular detection of gastrointestinal protozoa is more sensitive and more specific than microscopy but, to date, has not routinely replaced time-consuming microscopic analysis. Two internally controlled real-time PCR assays for the combined detection of Entamoeba histolytica , Giardia lamblia , Cryptosporidium spp. and Dientamoeba fragilis in single faecal samples were compared with Triple Faeces Test (TFT) microscopy results from 397 patient samples. Additionally, an algorithm for complete parasitological diagnosis was created. Real-time PCR revealed 152 (38.3%) positive cases, 18 of which were double infections: one (0.3%) sample was positive for E. histolytica , 44 (11.1%) samples were positive for G. lamblia , 122 (30.7%) samples were positive for D. fragilis , and three (0.8%) samples were positive for Cryptosporidium . TFT microscopy yielded 96 (24.2%) positive cases, including five double infections: one sample was positive for E. histolytica / Entamoeba dispar , 29 (7.3%) samples were positive for G. lamblia , 69 (17.4%) samples were positive for D. fragilis , and two (0.5%) samples were positive for Cryptosporidium hominis / Cryptosporidium parvum . Retrospective analysis of the clinical patient information of 2887 TFT sets showed that eosinophilia, elevated IgE levels, adoption and travelling to (sub)tropical areas are predisposing factors for infection with non-protozoal gastrointestinal parasites. The proposed diagnostic algorithm includes application of real-time PCR to all samples, with the addition of microscopy on an unpreserved faecal sample in cases of a predisposing factor, or a repeat request for parasitological examination. Application of real-time PCR improved the diagnostic yield by 18%. A single stool sample is sufficient for complete parasitological diagnosis when an algorithm based on clinical information is applied.     

结构生物学在溶组织内阿米巴致病机制研究中的应用

溶组织内阿米巴（Entamoeba histolytica,Eh）是寄生于人类肠道的原生动物,可引起阿米巴性结肠炎和肠外脓肿。结构生物学在原子水平上研究生命大分子结构和功能,是一种重要的生物学研究方法,经过半个多世纪的理论和技术发展,已应用于生物学和基础医学的各个研究领域。本文综述了目前结构生物学的研究技术,并以阿米巴穿孔素A和半胱氨酸蛋白酶抑制剂为例介绍结构生物学在Eh致病机制研究中的应用,总结了今后结构生物学在Eh研究领域可能的应用方向和研究热点。     

AIDS-ASSOCIATED PARASITIC DIARRHOEA

Since the advent of human immunodeficiency virus infection, with its profound and progressive effect on the cellular immune system, a group of human opportunistic pathogens has come into prominence. Opportunistic parasitic infection can cause severe morbidity and mortality. Because many of these infections are treatable, an early and accurate diagnosis is important. This can be accomplished by a variety of methods such as direct demonstration of parasites and by serological tests to detect antigen and/or specific antibodies. However, antibody response may be poor in these patients and therefore immunodiagnostic tests have to be interpreted with caution. Cryptosporidium parvum, Isospora belli, Cyclospora cayetanensis, Microsporidia, Entamoeba histolytica and Strongyloides stercoralis are the commonly detected parasites. Detection of these parasites will help in proper management of these patients because drugs are available for most of these parasitic infections.     

Amebic colitis in an antigenically and serologically negative patient: usefulness of a small-subunit ribosomal RNA gene-based polymerase chain reaction in diagnosis

Specific identification of Entamoeba histolytica in clinical specimens is an essential confirmatory diagnostic step in the management of amebiasis. Here, we report an unusual case of amebic colitis in a 20-year-old female immigrant from South China. The patient had experienced diarrhea, crampy abdominal pain, and fever for approximately 3 weeks prior to admission to hospital and had treated herself at home with metronidazole. On admission, stool microscopy and serology for E. histolytica were negative. Because the clinical findings raised the suspicion of Clostridium difficile fulminant colitis, she underwent a subtotal colectomy. Histopathology revealed flask-shaped ulcers characteristic of amebic colitis. Consequently, E. histolytica DNA was detected by a sensitive small-subunit rRNA polymerase chain reaction (PCR) from feces, and the patient was successfully treated for amebiasis with metronidazole. This case exemplifies the relative insensitivity of serologic tests for the diagnosis of intestinal amebiasis and the difficulties encountered in detecting the parasite antigen in a patient partially treated with metronidazole. We conclude that when the possibility of invasive intestinal amebiasis is suspected, detecting the parasite DNA directly in the stool sample by PCR using E. histolytica-specific primers may be an alternative, noninvasive, and reliable tool for the specific diagnosis of the disease.     

上海市区不同人群口腔原虫感染的调查

调查上海市区不同人群的口腔原虫的感染情况。从齿垢物取标本作新鲜涂片法检查。在４５０人中齿龈内阿米巴和口腔毛滴虫的感染率分别为２１．７８％和１．１１％。口腔科门诊患者的齿龈内阿米巴的感染率高达２８．３７％，而正常人的感染率仅为１０．７１％，两组差异非常显著。提示这两种原虫的感染与年龄有关，其感染率随年龄的增长有上升趋势，以及两种原虫感染与口腔疾病关系密切。     

用限制性内切酶分析溶组织内阿米巴的致病性

用SH-3、SH-5、SH-6、SH-7和SH-85株溶组织内阿米巴的DNA增殖35个周期,将其基因DNA用内切酶HinfⅠ和EcoT22Ⅰ进行消化后,作琼脂糖凝胶电泳分析,显示,5株虫株DNA经35个循环周期后产生531-bp的产物;经HinfⅠ消化后,SH-3、SH-5、SH-6和SH-7的基因DNA产生3个相同的片段,而显示其均与非致病性虫株SAW142的电泳谐完全相同;而SH-8基因DNA的电泳谱与致病性虫株SAW408的电泳谱一致,而用EcoT22Ⅰ消化结果也显示SH-8的图谱与致病性虫株SAW408的相同,证实了从包囊携带者和有发热、腹泻而无脓血便患者粪便中分离的虫株均属于非致病性虫株,从有发热、腹泻、脓血便的急性阿米巴痢疾患者粪便中分离到的虫株属于致病性虫株,均与酶株群分析相一致。提示,应用多聚酶链反应和限制性内切酶消化基因DNA来检测溶组织内阿米巴的基因型是十分有意义的。     

衡阳市居民381人口腔齿龈内阿米巴感染的调查

本文对衡阳市居民381人口腔齿龈内阿米巴感染进行了调查。采用刮取齿龈软垢、生理盐水涂片镜检的方法。结果表明,人群平均感染率为33％,阳性人群中,大部分为口腔卫生不良或牙周炎及并发牙周脓肿的患者。     

多聚酶链反应在阿米巴病流行病学研究中的应用

用两对引物扩增（ＰＣＲ）溶组织内阿米巴带囊者粪便中的ＤＮＡ，以鉴别感染虫株的致病性，致病和非致病性ＰＣＲ均为阳性者１人（３．１％），仅一项阳性者分别为６人（１８．８％）和２５人（７８．１％）。７例致病性ＰＣＲ阳性者中，６人有腹泻症状（ＯＲ＝３１．５）。各种对照标本均呈ＰＣＲ阴性反应。以ＰＣＲ法对一小样本人群进行阿米巴病调查，致病和非致病虫株的感染率均为１０％。结果表明，在阿米巴病流行病学调查中，可     

重庆市487例肠道寄生虫检查

使用粪便直接涂片法与饱和盐水浮聚法检查肠道蠕虫卵。使用粪便直接涂片,碘液染色和快速铁苏木素染色检查肠道原虫的包囊,非致病的虫瘘线虫卵与钩虫卵的鉴别。