Molecular and microscopic detection of natural and experimental infections of Toxocara vitulorum in bovine milk.

Toxocara vitulorum is an Ascarid nematode infecting the small intestine of buffalo and cattle particularly neonate calves, with the postnatal route through milk is the main infection source. However, little is known about shedding rates and the optimum detection methods of T. vitulorum larvae in the milk of the infected bovine hosts. In this study, we aimed to evaluate the use of two methods, microscopy and PCR, and their detection limits both under the experimental and natural infection situations. In doing this, T. vitulorum eggs extracted from naturally occurring adult female worms were successfully subjected to experimental embryonation, and larvae were implemented in experimental infection of milk in ascending infection doses of 0, 1, 5, 10, 20, 50 larvae/2-ml milk samples. With the except of negative control, microscopy-based examination detected larvae in all samples, albeit with means, ranges, and the total number of larvae were detected in exponential rates relative to larvae densities in milk samples. PCR technique corresponded well to microscopy in detecting genomic DNA of T. vitulorum larvae in all milk samples down to a single larva/sample. On the other hand, and by applying the same methodology approach on 50 naturally-occurring bovine colostrum/milk samples, 13 (26%) and 20 (40%) samples were tested positive for T. vitulorum infection by microscopy and the PCR-based detection, respectively. Of these, 11 out of 26 buffalo samples (42.30%) and 2 out of 24 cow samples (8.33%) were tested positive by microscopy, while 16 (61.54%) and 3 (12.50%) of buffalo and cow samples were tested positive by PCR, respectively. By applying the Agreement Coefficient, substantial agreement (0.77) between molecular and microscopy detection was detected from all tested samples. In conclusion, larvae of T. vitulorum were unequivocally detected by microscopy and molecular methods in milk samples both under the experimental and natural field situations. Nevertheless, slightly higher rates by PCR than microscopy were obtained when detecting naturally-infected milk samples. To the best of our knowledge, this is the first in situ detection of larvae of T. vitulorum in the milk of the naturally infected animals.

Assessment of the inhibitory effects of disinfectants on the embryonation of Ascaridia columbae eggs.

This study was set out with the aim of assessing the effects of the most commonly available commercial disinfectants on embryogenesis of A. columbae eggs. In addition to the distilled water treatment as a control group, four disinfectants were tested that included formalin, povidone iodide, TH4, and Virkon-S, in three independent experiments. In the first experiment, an overnight incubation with the working concentration of disinfectants resulted in significant inhibition of 80%, 85%, and 98% of embryonic development at day 9 post-treatment with formalin, povidone iodide, and TH4, respectively. This inhibition was continued through days 12 and 15 with the three tested disinfectants. Virkon-S did not affect embryogenesis with larval development comparable to that of the control group. In the second more dissected experiment, contact times of 10, 20, 30, and 60 minutes were set out for each disinfectant with embryogenesis inhibition results echoed that of the first experiment, with all disinfectants but Virkon-S inhibited larval development in significant proportions of eggs. Again, Virkon-S was very neutral in its effect on embryogenesis. When pigeon fecal matters were mixed with eggs and were subjected to disinfectants, discrepancies to results of the first two experiments were observed with only formalin inhibited embryogenesis in considerable proportion of eggs. Thus, and with the exception of Virkon-S, disinfectants tested at levels similar to those applicable in poultry in-houses and farms exhibited potent ovicidal activities on free eggs. To our knowledge, this is the first study involving the application of the commonly used in-poultry houses disinfectants to inactivate or delay the embryogenesis of bird Ascarids. The future perspective will potentially involve the in-field applications of the efficient disinfectants to eliminate or reduce the dissemination of infections with bird Ascarids in the chicken, turkey, pigeons, and other poultry houses.