Subekti Didik T, Azmi Zul, Kurniawati Dyah A, Suwanti Lucia T, Mufasirin Mufasirin, Sunarno Sunarno
Veterinary Science Program, Faculty of Veterinary Medicine, Airlangga University, Campus C - JL. Mulyorejo, Surabaya, East Java Province 60115, Indonesia; Center for Biomedical Research, Research Organization for Health, National Research and Innovation Agency, Cibinong Science Center, JL. Raya Jakarta - Bogor Km. 46, Bogor, West Jawa Province 16911, Indonesia.
Center for Standardization of Animal Husbandry and Animal Health Instruments, Agency for Standardization of Agricultural Instruments, Indonesian Ministry of Agriculture, JL. Raya Pajajaran Kav. E No. 59, Bogor, West Jawa Province 16143, Indonesia.
Vet Parasitol. 2024 Aug;330:110236. doi: 10.1016/j.vetpar.2024.110236. Epub 2024 Jun 15.
Four strains (SB-PR, SB-RS, SB-RD, and SB-RM) of Trypanosoma evansi (T. evansi) were used in this study. SB-PR is known to be trypanocide-sensitive, while the others are trypanocide-resistant to suramin, diminazene diaceturate, and melarsomine hydrochloride, respectively. SB-RS, SB-RD, and SB-RM are derivatives of a single field isolate of SB-PR. Trypanocide resistance will not only increase costs and decrease production efficiency but will also affect effective treatment strategies. Therefore, studies on this topic are important to avoid inefficient production and ineffective treatment. This paper aims to presents a comparative molecular characterization of the trypanocide-resistant strains compared to the parent population. Comparative molecular characterization of these strains based on a protein profile analysis performed with sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), DNA fingerprinting of random amplified polymorphic DNA (RAPD), and the molecular characterization of expression-site-associated 6 (ESAG6), variant surface glycoprotein (VSG), and T. evansi adenosine transporter-1 (TevAT1) gene sequences. The results show three derived strains (SB-RS, SB-RD, and SB-RM) exhibit different banding patterns than SB-PR. According to the RAPD results, SB-RS and SB-RD are different strains with DNA fingerprint similarities of about 77.8 %, while the DNA fingerprint of SB-RM has a similarity of 44.4 % to SB-RS and SB-RD. No differences in VSG were found among the four strains; however, ESAG6 showed differences in both nucleotide and amino acid sequences, as well as in its secondary and 3D structure. In conclusion, all molecular analyses of the ESAG6 gene showed that SB-PR, SB-RS, SB-RD, and SB-RM are different strains. Furthermore, SB-PR, SB-RS, SB-RD, and SB-RM did not exhibit the TevAT1 gene, so the resistance mechanism was determined to be unrelated to that gene.
本研究使用了四种伊氏锥虫（T. evansi）菌株（SB-PR、SB-RS、SB-RD和SB-RM）。已知SB-PR对杀锥虫剂敏感，而其他菌株分别对苏拉明、双乙酰氨苯胂和盐酸美拉胂明具有杀锥虫剂抗性。SB-RS、SB-RD和SB-RM是单一野外分离株SB-PR的衍生物。杀锥虫剂抗性不仅会增加成本并降低生产效率，还会影响有效的治疗策略。因此，关于该主题的研究对于避免生产效率低下和治疗无效很重要。本文旨在呈现与亲本群体相比，抗杀锥虫剂菌株的比较分子特征。基于十二烷基硫酸钠聚丙烯酰胺凝胶电泳（SDS-PAGE）进行的蛋白质谱分析、随机扩增多态性DNA（RAPD）的DNA指纹图谱以及表达位点相关6（ESAG6）、可变表面糖蛋白（VSG）和伊氏锥虫腺苷转运蛋白-1（TevAT1）基因序列的分子特征，对这些菌株进行比较分子表征。结果表明，三种衍生菌株（SB-RS、SB-RD和SB-RM）呈现出与SB-PR不同的条带模式。根据RAPD结果，SB-RS和SB-RD是不同的菌株，DNA指纹相似度约为77.8%，而SB-RM的DNA指纹与SB-RS和SB-RD的相似度为44.4%。在这四种菌株中未发现VSG存在差异；然而，ESAG6在核苷酸和氨基酸序列以及二级和三维结构上均显示出差异。总之，对ESAG6基因的所有分子分析表明，SB-PR、SB-RS、SB-RD和SB-RM是不同的菌株。此外，SB-PR、SB-RS、SB-RD和SB-RM未表现出TevAT1基因，因此确定抗性机制与该基因无关。
