Department of Animal Biotechnology, Madras Veterinary College, Tamil Nadu Veterinary and Animal Sciences University, Chennai, India, 600007.
Parasitol Res. 2010 Sep;107(4):1013-7. doi: 10.1007/s00436-010-1971-2. Epub 2010 Jul 6.
Coccidosis is one of the most commonly prevalent and economically important parasitic diseases of poultry worldwide. Chicken coccidia are protozoan parasites of the genus Eimeria. This study aimed at analysing the molecular prevalence of seven species of Eimeria infecting chickens in Tamil Nadu, India. Tissue samples (caecum, rectum and upper and mid intestines) collected from chickens exhibiting symptoms of coccidiosis were used for DNA extraction, followed by amplification of the internal transcribed spacer (ITS) region of Eimeria genome with genus-specific primers and speciation in nested polymerase chain reaction (PCR) with species-specific primers. Of 43 tissue samples examined, 25 were positive in ITS PCR and all the seven species could be identified. However, the prevalence of each species varied. In broilers, Eimeria necatrix was present in all infected chickens with Eimeria brunetti, Eimeria tenella, Eimeria maxima and Eimeria acervulina present in more than 50% of infected chickens, while Eimeria praecox and Eimeria mitis were only present in 11% to 16%. Although only 7 samples were positive among layers, the prevalence was largely similar, but with a higher prevalence of E. praecox and E. mitis and a lower prevalence of E. tenella. Multiple infections were most common, with 2-6 Eimeria species infecting the same chickens. In order to estimate the preponderance of each infecting species of Eimeria, a random cloning technique was adopted. The genus-specific ITS PCR product was cloned in a TA vector and ten clones were randomly picked and used as template for amplification of all the seven genera of Eimeria. If the specific species of Eimeria is preponderant, then the frequency of the clones showing that species-specific PCR amplification would be higher. Using this method, the most preponderant species present in the rectum, mid and upper intestines of layers was assessed to be E. acervulina, E. brunetti and E. necatrix. E. acervulina was present in 60-90%, E. necatrix in 10-30% and E. brunetti in 10-20% of the clones screened, indicating that these species could be the most preponderant Eimeria species. Intervention strategies should aim at these species. This new method of estimating preponderance of infecting Eimeria species could be used to assess the relative importance of each species at the farm or region level instead of relying only on prevalence estimates.
球虫病是全世界家禽中最常见和最重要的寄生虫病之一。鸡球虫是艾美耳属的原生动物寄生虫。本研究旨在分析印度泰米尔纳德邦感染鸡的七种艾美耳球虫的分子流行情况。从表现出球虫病症状的鸡中采集盲肠、直肠和上、中肠组织样本，用于 DNA 提取，然后用属特异性引物扩增艾美耳球虫基因组的内部转录间隔区（ITS），并用种特异性引物进行嵌套聚合酶链反应（PCR）进行种化。在检查的 43 个组织样本中，25 个在 ITS-PCR 中呈阳性，所有七种均可鉴定。然而，每种的流行情况有所不同。在肉鸡中，所有感染鸡都存在艾美尔球虫，而艾美尔球虫、柔嫩艾美尔球虫、巨型艾美尔球虫和堆型艾美尔球虫在超过 50%的感染鸡中存在，而早熟艾美尔球虫和微小艾美尔球虫仅存在于 11%至 16%的感染鸡中。虽然在蛋鸡中仅 7 个样本呈阳性，但流行情况大致相似，但早熟艾美尔球虫和微小艾美尔球虫的流行率较高，而柔嫩艾美尔球虫的流行率较低。最常见的是混合感染，有 2-6 种艾美耳球虫感染同一鸡。为了估计感染艾美耳球虫的每种优势种的比例，采用了随机克隆技术。属特异性 ITS-PCR 产物克隆在 TA 载体中，随机挑选 10 个克隆作为模板，用于扩增所有 7 种艾美耳球虫。如果特定的艾美耳球虫优势种，则显示该种特异性 PCR 扩增的克隆频率会更高。使用这种方法，评估了蛋鸡直肠、中肠和上肠中最优势的种为堆型艾美尔球虫、柔嫩艾美尔球虫和毒害艾美尔球虫。堆型艾美尔球虫存在于 60-90%的克隆中，毒害艾美尔球虫存在于 10-30%的克隆中，柔嫩艾美尔球虫存在于 10-20%的克隆中，表明这些种可能是最优势的艾美耳球虫种。干预策略应针对这些种。这种估计感染艾美耳球虫优势种的新方法可用于评估农场或地区水平上每种种的相对重要性，而不仅仅依赖于流行率估计。
