Development and validation of a diagnostic test for anthelmintic resistance monitoring in sheep flocks

Author(s)

Yousmel Alemán Gainza

• Document type: Doctoral Thesis
• Press: Jaboticabal. 2020-07-29.
• Institution: Universidade Estadual Paulista (Unesp). Faculdade de Ciências Agrárias e Veterinárias. Jaboticabal
• Defense date: 2020-05-29
• Advisor: Ana Carolina de Souza Chagas

Abstract

The larval development test (LDT) can be a diagnostic tool for anthelmintic resistance in flocks, allowing monitoring and management adjustments. The aim of this study was to develop and validate an in vitro diagnostic test of Haemonchus contortus resistance to commercial anthelmintics and to validate the results at the farm level. The efficacy of thiabendazole (TBZ), levamisole (LEV), ivermectin-monosaccharide (IVM-M), monepantel (MPT) and Zolvix® (ZLV) were evaluated in LDT in susceptible (Echevarria1991-HcEc91) and resistant (Botucatu-HcBot) isolates, in 24 and 96 well culture plates. Complementary tests were subsequently performed with ivermectin aglycone (IVM-A). To validate the LDT, the fecal egg count reduction test (FECRT) was performed in five sheep flocks with commercial anthelmintics from the same chemical groups (groups of 7 animals with FEC ≥ 200). LDT and fecal cultures were also carried out with the samples of the flocks (HcFlocks). H. contortus was the major parasite on all farms (average of 74%). The resistance factor (RF) defined for the isolates was greater than 3, indicating that the test was able to differentiate HcEc91 and HcBot for TBZ, LEV, MPT, ZLV, IVM-A, and in a less consistent way for IVM-M. The similarity of the dose-response curves between the plates and plate*concentration interaction (R2 from 98.4 to 99.0% for HcEc91/HcBot and from 99.4 to 99.0% for HcFlocks), as well as low differences in efficacy mean (0.02 to 2.26% for HcEc91/HcBot and 0.02 to 4.90% for HcFlocks) for all anthelmintics, except for MPT, indicated a reliable agreement of the LDT in both plates, both for the evaluation of HcEc91/HcBot as for HcFlocks. In the latter case, resistance detection by ZLV was clearer and more stable than by MPT. The adaptation of the test to 96-well plates resulted in savings of at least 51.9%. Through FECRT, resistance to all chemical groups was detected in all flocks, except for ZLV (40% resistant and 20% suspected of resistance). LDT results in flocks indicated resistance to TBZ (100%), LEV (80%), ZLV (20%), IVM-M (0%) and IVM-A (100%). Total agreement was obtained regarding the results of both tests for TBZ and IVM-A (k = 1.00). For LEV and ZLV, although there was a divergence of results in two flocks, this was not statistically different for both drugs (P = 0.077 and P = 0.197, respectively), with substantial agreement (k = 0.8 and 0.6, respectively). The data from the present study indicate that the results of the two tests are comparable, thus validating this test as an option for the diagnosis of resistance for small ruminant farmers. (AU)

• FAPESP's process: 16/07132-8 - Development and validation of a diagnostic kit of parasitic resistance to anthelminthics for resistance monitoring in sheep flocks
• Grantee: Yousmel Alemán Gainza
• Support Opportunities: Scholarships in Brazil - Doctorate