Department of Microbiology, Institute of Medical Sciences, Banaras Hindu University, Varanasi, India.
BMC Microbiol. 2010 Jan 15;10:11. doi: 10.1186/1471-2180-10-11.
Enteric protozoa and sporozoa have emerged as important opportunistic parasites and can cause fatal infections in AIDS patients. The line of treatment being different for them necessitates an accurate and prompt identification of these to avoid empirical treatment. In this study which is the first of its kind from India we did a comprehensive evaluation of different techniques, comparing them on the basis of the attributes like yield, cost, time taken, expertise and infrastructure. For the first time combination of Calcoflour White and DAPI, a nuclear stain, were used to identify Microsporidia spp. Thus, a diagnostic protocol was devised for rapid, sensitive and cost effective identification of the opportunistic enteric protozoa.
The organisms isolated from the stool samples of the cases (450 HIV patients) were predominantly Cryptosporidium spp., Microsporidia spp. and Cyclospora spp. Interestingly, the control group (200 relatives of the patients who were HIV negative) showed a high incidence (21%) of Cryptosporidium spp. We found a significant increase in the sensitivity of microscopy in detecting Cryptosporidium spp. and Cyclospora spp. after formol ether concentration. Kinyoun's staining was better compared to Modified safranin staining for Cryptosporidium spp. identification. Although ELISA had a sensitivity of 93.25% and specificity of 97% for Cryptosporidium spp. detection, we ranked Kinyoun's staining better than ELISA because it is not affordable to most of our patients. For detecting Cyclospora cayetanensis, autoflourescence was the easiest and most cost effective method followed by Safranin technique. Combination of Calcoflour White stain and DAPI gave good results for the identification of Microsporidia spp. We assessed the above techniques and graded the attributes in the following descending order: cost effectiveness, sensitivity, ease of use and interpretation, time taken for the procedure and batch testing.
Thus, we conclude that a combination of minimum three procedures should be carried out for the screening of stool specimens of HIV positive patients. Kinyoun's staining should be made mandatory for every diarrheal stool sample from HIV patients. Also every laboratory should assign its own value to the attributes and apply Multiattribute utility theory or the Analytical hierarchy process to decide the most appropriate methodology.
肠道原生动物和孢子虫已成为重要的机会性寄生虫,可在艾滋病患者中引起致命感染。由于它们的治疗方法不同,因此需要准确、及时地识别这些寄生虫,以避免经验性治疗。在这项研究中,我们首次从印度进行了全面评估,比较了不同技术,根据产量、成本、所需时间、专业知识和基础设施等属性进行了比较。我们首次将Calcoflour White 和 DAPI(一种核染色剂)联合用于鉴定微孢子虫属。因此,制定了一种快速、敏感且具有成本效益的诊断方案,用于鉴定机会性肠道原生动物。
从病例(450 名 HIV 患者)的粪便样本中分离出的生物体主要是隐孢子虫属、微孢子虫属和环孢子虫属。有趣的是,对照组(200 名 HIV 阴性患者的亲属)的隐孢子虫属发生率(21%)很高。我们发现福尔马林乙醚浓缩后,显微镜检测隐孢子虫属和环孢子虫属的灵敏度显著提高。与改良萨夫兰染色相比,金永氏染色更适合隐孢子虫属的鉴定。尽管 ELISA 对隐孢子虫属的检测具有 93.25%的灵敏度和 97%的特异性,但我们认为金永氏染色优于 ELISA,因为它对我们大多数患者来说都负担不起。对于检测环孢子虫属,自发荧光是最简单、最具成本效益的方法,其次是 Safranin 技术。Calcoflour White 染色和 DAPI 的组合可很好地鉴定微孢子虫属。我们评估了上述技术,并按以下降序对属性进行评分:成本效益、灵敏度、易用性和解释性、程序所需时间和批处理测试。
因此,我们得出结论,对于 HIV 阳性患者的粪便标本筛查,应至少进行三种程序的组合。金永氏染色应成为每位 HIV 患者腹泻粪便样本的强制性检查。此外,每个实验室都应为本实验室的属性赋予自己的价值,并应用多属性效用理论或层次分析法来决定最合适的方法。