Dip. di Sanità Pubblica e Malattie Infettive, Università di Roma Sapienza, Piazzale Aldo Moro 5, Rome, Italy.
Vector Borne Zoonotic Dis. 2012 Feb;12(2):106-10. doi: 10.1089/vbz.2011.0706. Epub 2011 Oct 13.
Babesioses are hematic tick-borne diseases that induce malaria-like disorders in domestic, wild animals, and humans. Although indirect fluorescent antibody test (IFAT) and enzyme-linked immunosorbent assay (ELISA) commercial kits are available to test the presence of antibodies against most Babesia species, no kit exists to serologically diagnose the infections due to Babesia divergens, one of the most important zoonotic species. To fill this gap and to develop assays to detect animal and human infections, in vitro cultures (microaerophilous stationary phase system) of B. divergens were organized. Infected erythrocytes were adsorbed as corpuscular antigen (CA) on IFAT slides and ELISA microwells. The supernatant medium of the cultures (metabolic antigen, MA) was collected and employed in ELISA and western blot (WB) assays. B. divergens was also used to produce positive sera in Meriones unguiculatus and to infect a calf. Serological tests were set up with sera from experimentally/naturally infected animals, and possible cross-reactions were evaluated using heterologous sera from cattle positive to other piroplasms. Sera from clinically healthy people at risk of infection were also tested. As expected, assays based on the purified MAs from in vitro cultures proved more sensitive and specific than CA-IFAT and CA-ELISA. In fact, MA-ELISA provided satisfactory performances (even if 8.4%-15.7% cross-reactions were evidenced), and the WB developed proved totally sensitive and specific. WB indicated as immunodominant antigens two major protein bands at 33 and 37 kDa, which were also evidenced in 2.2% of the human sera tested, proving the parasite transmission to humans also in Italy.
巴贝斯虫病是一种血源性蜱传疾病，可引起家畜、野生动物和人类疟疾样疾病。虽然间接荧光抗体试验（IFAT）和酶联免疫吸附试验（ELISA）商业试剂盒可用于检测大多数巴贝斯虫属物种的抗体存在情况，但尚无试剂盒可用于血清学诊断由重要的人畜共患病种之一——分歧巴贝斯虫引起的感染。为了填补这一空白并开发用于检测动物和人类感染的检测方法，我们建立了分歧巴贝斯虫的体外培养（微需氧固定相系统）。感染的红细胞被吸附为IFAT 载玻片和 ELISA 微量滴定孔上的颗粒抗原（CA）。培养物的上清液（代谢抗原，MA）被收集并用于 ELISA 和 Western blot（WB）检测。分歧巴贝斯虫还用于在蒙古沙鼠中产生阳性血清并感染小牛。使用来自实验/自然感染动物的血清建立了血清学检测方法，并使用对其他梨形虫呈阳性的牛异源血清评估了可能的交叉反应。还检测了有感染风险的临床健康人群的血清。如预期的那样，基于体外培养物的纯化 MA 的检测方法比 CA-IFAT 和 CA-ELISA 更敏感和特异。事实上，MA-ELISA 提供了令人满意的性能（即使有 8.4%-15.7%的交叉反应），而开发的 WB 完全敏感和特异。WB 表明两种主要的 33 和 37 kDa 蛋白带为免疫优势抗原，在 2.2%的测试人类血清中也有证据，证明该寄生虫也在意大利传播给人类。
