Combination of cell culture and qPCR to assess the efficacy of different anticoccidials on XXXX sporozoites.

@entity7846 @entity2697 @entity12917 @entity7844 @entity281 @entity71346 @entity291
UNASSIGNED: Three in vitro studies were designed to develop an assay for anticoccidial efficacy by use of laboratory (Houghton) and field (T-376) @entity12917 strains. In study (1), minimum inhibitory concentrations (MICs) of monensin (Mon), @entity7846 ( @entity71346 ), @entity7844 ( @entity2697 ), and lasalocid (Las) were determined that are able to inhibit more than 50  % of sporozoites in host cell (Madin-Darby @entity291 kidney (MDBK)) penetration and more than 95  % of Houghton sporozoites development to mature merozoites (treatment time 24  h) using quantitative real-time PCR (qPCR). MICs were 0.5, 2.5, 1, and 0.5   g/ml for Mon, @entity71346 , @entity2697 , and Las, respectively. Applying the previous MIC on T-376 strain revealed a different sensitivity profile. @entity71346 reduced T-376 gene copies by only 89.3  % after 96  h of @entity281 . In study (2), Houghton strain sporozoites were incubated with MIC of the different tested ionophores for 2 and 4  h, respectively; afterwards, their ability to invade MDBK cells was determined using phase-contrast microscopy and qPCR. Treatment of sporozoites with ionophores for 4  h resulted in significant inhibition of invasion compared with non-treated parasites as assessed both by microscopy as well as qPCR. Inhibition rates for Mon, @entity71346 , @entity2697 , and Las were 90.2, 75.0, 88.3, and 82.6  % using phase-contrast microscopy and 83.9, 81.4, 85.8, and 75.4  % using qPCR, respectively. T-376 sporozoite invasion into MDBK cells was reduced to 48.9  % by @entity71346 . Study (3) was conducted to determine inhibition exerted by toltrazuril (Tol). Tol at 5   g/ml reduced reproduction of Houghton strain by 95  %, whereas T-376 was only reduced by 86.5  %. The presented experiments indicate that infectivity inhibition of sporozoites incubated for 4  h with anticoccidials and development inhibition after 96  h of @entity281 by qPCR are suitable means to assess sensitivity of @entity12917 strains to anticoccidials.
