Folle Ana Maite, Kitano Eduardo S, Lima Analía, Gil Magdalena, Cucher Marcela, Mourglia-Ettlin Gustavo, Iwai Leo K, Rosenzvit Mara, Batthyány Carlos, Ferreira Ana María
Cátedra de Inmunología, Facultad de Ciencias/Facultad de Química, Universidad de la República (UdelaR), Montevideo, Uruguay.
Laboratório Especial de Toxinologia Aplicada, Center of Toxins, Immune-Response and Cell Signaling (CeTICS), Instituto Butantan São Paulo, Brazil.
PLoS Negl Trop Dis. 2017 Jan 3;11(1):e0005250. doi: 10.1371/journal.pntd.0005250. eCollection 2017 Jan.
The larva of cestodes belonging to the Echinococcus granulosus sensu lato (s.l.) complex causes cystic echinococcosis (CE). It is a globally distributed zoonosis with significant economic and public health impact. The most immunogenic and specific Echinococcus-genus antigen for human CE diagnosis is antigen B (AgB), an abundant lipoprotein of the hydatid cyst fluid (HF). The AgB protein moiety (apolipoprotein) is encoded by five genes (AgB1-AgB5), which generate mature 8 kDa proteins (AgB8/1-AgB8/5). These genes seem to be differentially expressed among Echinococcus species. Since AgB immunogenicity lies on its protein moiety, differences in AgB expression within E. granulosus s.l. complex might have diagnostic and epidemiological relevance for discriminating the contribution of distinct species to human CE. Interestingly, AgB2 was proposed as a pseudogene in E. canadensis, which is the second most common cause of human CE, but proteomic studies for verifying it have not been performed yet. Herein, we analysed the protein and lipid composition of AgB obtained from fertile HF of swine origin (E. canadensis G7 genotype). AgB apolipoproteins were identified and quantified using mass spectrometry tools. Results showed that AgB8/1 was the major protein component, representing 71% of total AgB apolipoproteins, followed by AgB8/4 (15.5%), AgB8/3 (13.2%) and AgB8/5 (0.3%). AgB8/2 was not detected. As a methodological control, a parallel analysis detected all AgB apolipoproteins in bovine fertile HF (G1/3/5 genotypes). Overall, E. canadensis AgB comprised mostly AgB8/1 together with a heterogeneous mixture of lipids, and AgB8/2 was not detected despite using high sensitivity proteomic techniques. This endorses genomic data supporting that AgB2 behaves as a pseudogene in G7 genotype. Since recombinant AgB8/2 has been found to be diagnostically valuable for human CE, our findings indicate that its use as antigen in immunoassays could contribute to false negative results in areas where E. canadensis circulates. Furthermore, the presence of anti-AgB8/2 antibodies in serum may represent a useful parameter to rule out E. canadensis infection when human CE is diagnosed.
细粒棘球绦虫广义复合体的幼虫可引发囊型包虫病（CE）。它是一种全球分布的人畜共患病，具有重大的经济和公共卫生影响。用于人类CE诊断的最具免疫原性和特异性的棘球属抗原是抗原B（AgB），它是包虫囊肿液（HF）中的一种丰富脂蛋白。AgB蛋白部分（载脂蛋白）由五个基因（AgB1 - AgB5）编码，这些基因产生成熟的8 kDa蛋白（AgB8/1 - AgB8/5）。这些基因在棘球属物种中似乎存在差异表达。由于AgB的免疫原性取决于其蛋白部分，细粒棘球绦虫广义复合体内AgB表达的差异可能在区分不同物种对人类CE的贡献方面具有诊断和流行病学意义。有趣的是，AgB2在加拿大棘球绦虫中被认为是一个假基因，加拿大棘球绦虫是人类CE的第二大常见病因，但尚未进行蛋白质组学研究来验证这一点。在此，我们分析了从猪源（加拿大棘球绦虫G7基因型）的有生殖力的HF中获得的AgB的蛋白质和脂质组成。使用质谱工具对AgB载脂蛋白进行了鉴定和定量。结果表明，AgB8/1是主要的蛋白质成分，占总AgB载脂蛋白的71%，其次是AgB8/4（15.5%）、AgB8/3（13.2%）和AgB8/5（0.3%）。未检测到AgB8/2。作为方法学对照，平行分析在牛的有生殖力的HF（G1/3/5基因型）中检测到了所有的AgB载脂蛋白。总体而言，加拿大棘球绦虫的AgB主要由AgB8/1和脂质的异质混合物组成，尽管使用了高灵敏度蛋白质组学技术，仍未检测到AgB8/2。这证实了基因组数据，支持AgB2在G7基因型中表现为假基因。由于重组AgB8/2已被发现对人类CE具有诊断价值，我们的研究结果表明，在加拿大棘球绦虫流行地区，将其用作免疫测定中的抗原可能会导致假阴性结果。此外，血清中抗AgB8/2抗体的存在可能是在诊断人类CE时排除加拿大棘球绦虫感染的一个有用参数。
