Leishmania infantum-derived lipophosphoglycan as an antigen in the accurate serodiagnosis of canine leishmaniasis

Lipophosphoglycan (LPG) is the major surface glycoconjugate of Leishmania protozoan and has an important biological role in host-parasite interactions both in the midgut epithelium of the sand fly vector and in the vertebrate macrophages. Canine leishmaniasis (CanL) is a chronic infectious disease predominantly caused by Leishmania infantum. An early and accurate immunodiagnosis of the disease is crucial for veterinary clinical practice and for disease control. In this work, we evaluated L. infantum LPG as an antigen in an indirect enzyme-linked immunosorbent assay (ELISA) for CanL immunodiagnosis (LPG-ELISA) by testing serum samples from 97 naturally infected dogs with diverse clinical presentations ranging from subclinical infection to severe disease, as evaluated by veterinarian infectologists. Serum samples from healthy dogs from non-endemic areas (n = 68) and from dogs with other infectious diseases (n = 64) were used as controls for assay validation. The performance of the LPG-ELISA was compared with that of an ELISA using the soluble fraction of L. infantum total lysate antigen (TLA). LPG-ELISA presented a superior performance in comparison to TLA-ELISA, with 91.5% sensitivity, 98.5% specificity and 99.7% accuracy. A distinguishing feature of the LPG-ELISA compared to the TLA-ELISA was its higher ability to identify subclinical infection in clinically healthy dogs, in addition to the absence of cross-reactivity with other canine infectious diseases. Finally, LPG-ELISA was compared to TR DPP visceral canine leishmaniasis test, the immunochromatographic test recommended by the Brazilian Ministry of Agriculture. LPG-ELISA exhibited higher values of specificity (98.5% versus 93.1%) and sensitivity (91.5% versus 90.6%) compared to TR DPP. In conclusion, L. infantum-derived LPG was recognized by antibodies elicited during CanL in different infection stages and was shown to be a suitable antigen for specific clinical settings of veterinary diagnosis and for public health usage.

脂磷酸聚糖（Lipophosphoglycan, LPG）是利什曼原虫的主要表面糖缀合物，在白蛉媒介的中肠上皮以及脊椎动物巨噬细胞的宿主-寄生虫互作过程中发挥关键生物学功能。犬利什曼病（CanL）是一种主要由婴儿利什曼原虫（Leishmania infantum）引起的慢性传染病，早期精准的免疫诊断对于兽医临床实践与疾病防控均具有重要意义。本研究以婴儿利什曼原虫来源的LPG为抗原，构建了用于犬利什曼病免疫诊断的间接酶联免疫吸附试验（LPG-ELISA），并对其检测性能展开评估：共纳入97份经兽医感染病学家确认的自然感染犬血清样本，受试犬的临床表现多样，涵盖从亚临床感染到重症疾病的不同感染阶段；同时设置两组对照样本，分别为来自非流行地区的健康犬血清68份，以及患有其他传染病的犬血清64份，用于验证该检测方法的性能。随后，本研究将LPG-ELISA的检测性能与以婴儿利什曼原虫全裂解物可溶性组分抗原（TLA）为包被原的ELISA（简称TLA-ELISA）进行对比，结果显示LPG-ELISA的整体性能优于TLA-ELISA，其灵敏度为91.5%、特异性为98.5%、准确率为99.7%。相较于TLA-ELISA，LPG-ELISA的显著优势在于能够更高效地识别临床健康犬的亚临床感染，且不会与其他犬类传染病发生交叉反应。最后，本研究将LPG-ELISA与巴西农业部推荐的TR DPP内脏犬利什曼病免疫层析检测试剂盒进行对比，结果表明LPG-ELISA的特异性（98.5% vs 93.1%）与灵敏度（91.5% vs 90.6%）均高于TR DPP。综上，婴儿利什曼原虫来源的LPG可被犬利什曼病不同感染阶段产生的抗体识别，其作为抗原适用于兽医诊断的特定临床场景以及公共卫生应用场景。