Effects of flumethrin on the germinative cells of engorged females of Rhipicephalus annulatus.

Chemical acaricides are widely used for the effective control of ticks in India. The synthetic pyrethroids, are one of the most popular chemical acaricides with selective neurotoxic potential. Flumethrin is a type II synthetic pyrethroid used extensively in veterinary practice in India. The present study was undertaken to evaluate the cytotoxic effects of flumethrin on the engorged females of Rhipicephalus annulatus using entomological parameters, histology, electron microscopy and relative quantification of receptors of dopamine and GABAB mRNAs. Adult immersion test (AIT) using flumethrin (100 ppm), revealed twenty per cent mortality of ticks, hundred per cent inhibition of fecundity and complete blocking of hatching of the laid eggs. Microscopic analysis of the structure of the ovaries after 24 h of treatment with flumethrin (90 ppm) revealed changes, viz., reduction in size with the presence of amorphous material inside stage I oocytes, wrinkled boundary and chromatin fragmentation of nucleus of stage II oocytes, vacuoles around the germinal vesicle, thickening of the nuclear membrane and chromatin clumping of stage III oocytes and reduction in size and shape of mature stage IV and V oocytes. Also, a large number of vacuoles were observed throughout the pedicel cell region of stage II and III oocytes. Ultrastructurally, irregular nuclear membrane, swelling as well as crystolysis of mitochondria and detachment of external and internal layers of the basal lamina of oocytes were the major structural alterations confirming direct damaging effects of flumethrin on the germinative cells. The relative quantification of the expression of dopamine D1, dopamine D2 and GABAB receptors by quantitative real-time PCR (qRT PCR), revealed the upregulation of dopamine D1 receptor and downregulation of receptors of dopamine D2 and GABAB in the ovary of treated ticks.

In vitro efficacy of amitraz, coumaphos, deltamethrin and lindane against engorged female Rhipicephalus (Boophilus) annulatus and Haemaphysalis bispinosa ticks.

The present study compares the in vitro efficacy of four chemical acaricides, viz. amitraz, coumaphos, deltamethrin and lindane, against Rhipicephalus (Boophilus) annulatus and Haemaphysalis bispinosa ticks based on adult immersion tests. Amitraz, at 350 ppm, elicited 29.2 ± 4.17% mortality against R. (B.) annulatus, 100% inhibition of fecundity and absence of hatching of eggs laid by treated ticks. The same compound at 300 ppm caused 62.5 ± 12.5% mortality against H. bispinosa, 96.7% inhibition of fecundity and complete blocking of eclosion. The LC50 value of amitraz against susceptible H. bispinosa was 181 ppm. Deltamethrin at 400 ppm, elicited 25.0 ± 4.81% adult R. (B.) annulatus mortality, 97.5% inhibition of fecundity and absence of egg hatching. Complete blocking of egg hatching was observed even at 30 ppm. However, deltamethrin (at 50 ppm) elicited 75.0 ± 10.76% mortality against H. bispinosa, 65.8% inhibition of fecundity and very low egg hatching (10%). The LC50 for deltamethrin against susceptible H. bispinosa was 33.8 ppm. Coumaphos at 50 ppm, caused mortality of 70.8 ± 4.17% with R. (B.) annulatus whereas 100% mortality was observed against H. bispinosa. The LC50 values of coumaphos against R. (B.) annulatus and H. bispinosa were 9 and 8.75 ppm, respectively. Complete inhibition (100%) of fecundity was observed even at 30 ppm against both parasites. Complete blocking of egg hatching was also observed even at 10 ppm of coumaphos. Lindane at 1000 ppm caused mortality of 87.5 ± 7.98% against R. (B.) annulatus and 83.3% mortality against H. bispinosa at 100 ppm. The LC50 values of lindane against R. (B.) annulatus and H. bispinosa were 157 and 8.61 ppm, respectively. Complete inhibition of fecundity was observed with R. (B.) annulatus treated with lindane above 200 ppm and with H. bispinosa at a concentration above 50 ppm. Complete blocking of egg hatching was observed in R. (B.) annulatus, even at 100 ppm. Lindane caused 100% blocking of egg hatching at 1 ppm in the case of H. bispinosa.

In vitro effects of caffeic acid, nortriptyline, precocene I and quercetin against Rhipicephalus annulatus (Acari: Ixodidae).

In the present study, the acaricidal effects of caffeic acid, nortriptyline, precocene I and quercetin against Rhipicephalus annulatus (syn. Boophilus annulatus) Say (Acari: Ixodidae) were evaluated. Adult immersion technique (24 ticks immersed for 2 min in one dilution of the compound) was used for the assessment of the effects of caffeic acid (0.39-100 mg/mL), nortriptyline (0.625-50 mg/L), precocene I (0.004488-5 mg/mL) and quercetin (6.25-100 mg/mL) against R. annulatus. Adult tick mortality, reproductive index, inhibition of fecundity and hatching were calculated. Caffeic acid, nortriptyline, precocene I and quercetin revealed very low adult mortality and inhibition of fecundity, even at the highest concentration tested. Quercetin (>50 mg/mL) caused blocking of hatching of eggs.

Ultrastructural analysis of oocytes of Rhipicephalus (Boophilus) annulatus during postengorgement period as a tool to evaluate the cytotoxic effects of amitraz and deltamethrin on the germinative cells.

The present study utilizes the ultrastructural analysis of the fully engorged female Rhipicephalus (Boophilus) annulatus ticks, as a tool to evaluate the cytotoxic potential of deltamethrin and amitraz on the germinative cells. The ultrastructural analysis of the ovary of the normal (untreated) R (B.) annulatus revealed, oocytes in different stages of development, attached to the ovary wall by pedicel cells. The attachment site of oocyte to the pedicel cell was characterized by indentations of the plasma membrane. The oocyte was bound by three cell membranes viz., plasma membrane, chorion and basal lamina. The stages of oocytes were differentiated ultrastructurally based on the features of their outer membrane and the number and size of lipid and yolk droplets. Detailed day wise analysis of ultrastructural changes in the ovary during the post-engorgement period revealed the occurrence of the degenerative changes from day five onwards. These appeared first in the oocytes followed by the germinal epithelium. The ovary of ticks treated with methanol (control), revealed similar topographies as that of a normal ovary except for the presence of very few oocytes with ring shaped nucleoli. Ultrastructurally, treatment with deltamethrin produced more prominent and extensive morphological alterations when compared to amitraz. In the case of ticks treated with amitraz, the oocytes of stage IV and V showed wavy and disrupted outer boundaries along with the loss of integrity of the yolk droplets. Uneven nuclear membranes of stage II oocytes and cristolysis of mitochondria of mature oocytes were the other changes noticed. Ticks treated with deltamethrin revealed prominent modifications such as, detachment of the basal lamina, wrinkled boundary, inconsistent nuclear membrane, ring shaped nucleoli and chromatin clumping in the case of the early stage oocytes (I and II), whereas swelling and cristolysis of mitochondria were seen in mature oocytes. The study further indicated that, in addition to the previous proven neurotoxic effects, these compounds act directly on the ovary of tick.