Putative SNPs in Ovar-DRB1 and GALNTL6 Genes Conferring Susceptibility to Natural Infection of Haemonchus Contortus in Southern Indian Sheep.

AIM: To explore associations between phenotypic traits and polymorphisms in the DRB1 and GALNT6 gene in Nellore, Deccani and Kenguri sheep naturally infected with Haemonchus contortus. MATERIALS AND METHODS: Blood and faecal samples were collected to evaluate fecal worm egg counts (FEC), packed cell volume (PCV), hemoglobin (Hb), eosinophilia and for DNA isolation. RESULTS: Animals were grouped into susceptible and resistant groups based on EPG counts. FEC and circulating eosinophilia were higher in a susceptible group. Log FEC was negatively correlated (P < 0.01) with PCV, and Hb estimates. The second exon of DRB1 and intron variant of GALNTL6 genes were amplified from DNA samples of resistant and susceptible sheep. Characterization of Ovar-DRB1 amplicon by RFLP revealed two genotypes ('bb' and 'ab'). The genotype frequencies differed significantly between both groups (P < 0.05). The 'bb' genotypes had higher (P < 0.05) log FEC value than 'ab' genotypes and 'b' allele was linked with susceptibility to haemonchosis in sheep. The mean FEC of Nellore sheep was high indicating susceptibility of the breed and also in which the frequency of 'b' allele was more compared to the other two breeds. OVAR-DRB1 genotypes associated with FEC did not affect PCV and Hb. PCR-RFLP assay developed to determine the genotypes with respect to SNP rs424521894 of GALNTL6 revealed monomorphic nature at the locus in the breeds studied. CONCLUSION: MHC polymorphism could be used as a genetic marker for the selection of sheep resistant to H. contortus. However, a more intensive study, involving controlled infections and other GALNTL6 SNPs may be enforced to make any decisive assertion.

Co-infections of major tick-borne pathogens of dogs in Andhra Pradesh, South India.

Implementing hemoprotozoan control strategies in dogs has become difficult because of the co-infections. A multiplex polymerase chain reaction (PCR) was carried out for simultaneous detection of the co-infections of Babesia gibsoni, B. vogeli, Hepatozoon canis and Ehrlichia canis from dogs (N = 442) in Andhra Pradesh, South India. The co-infection combinations were classified as (i) B. gibsoni + B. vogeli + E. canis + H. canis (BEH), (ii) B. gibsoni + B. vogeli + E. canis (BE), (iii) B. gibsoni + B. vogeli + H. canis (BH) and (iv) E. canis + H. canis (EH) groups. The parasite-specific multiplex PCR amplified 18S rRNA gene of B. gibsoni, B. vogeli and H. canis and VirB9 gene of E. canis. The age, gender, breed, medium, living condition and region of dogs were studied as risk factors for co-infections using logistic regression model. Among the co-infections, the incidence was 1.81%, 9.28%, 0.69% and 0.90% for BEH, BE, BH and EH infections, respectively. Young age (< one year), females, mongrels, rural dogs, kennel dogs and presence of ticks were the identified risk factors for overall prevalence of tick-borne pathogens. The incidence of infection was less in rainy season, especially in dogs with a previous acaricidal treatment. The study concludes that the multiplex PCR assay could simultaneously detect natural co-infections in dogs, emphasizing the need for the assay in epidemiological studies to reveal the real pattern of pathogens and select pathogen-specific treatment protocols.

Prevalence of taeniosis in pig farming community in Andhra Pradesh, India.

Taeniosis is one of the neglected zoonoses in India. Facts on taeniosis, compared to cysticercosis are scanty in India. Hence, this study is aimed to determine the occurrence of taeniosis in human beings in Andhra Pradesh, India. A total of 1380 stool samples were collected from people associated with pig farming and or had the habit of pork eating in certain districts (n = 7) of Andhra Pradesh. Prevalence of human taeniosis was determined using microscopic examination of stool samples and proglottds. The overall prevalence of taeniosis was found to be 0.79%. Morphology of gravid segments revealed lower number of lateral branches indicating T. solium segments. Age and gender of human were not associated with the occurrence of taeniosis. The low prevalence of taeniosis in human indicates good hygiene and sanitation practices in humans, and awareness about the disease and its transmission. Further studies involving more sensitive techniques on stool and serum samples are warranted.

Cypermethrin and ivermectin resistance in field populations of Rhipicephalus sanguineus sensu lato (Latrielle, 1806) collected from dogs in south India.

The abuse of acaricides for control of Rhipicephalus sanguineus sensu lato ticks led to a strong selection pressure for acaricide resistance. Data on acaricide resistance in R. sanguineus s.l. populations from India is meager though veterinarians frequently report treatment failures. The present study was aimed to characterize the level of resistance in R. sanguineus s.l. against most commonly used drugs, cypermethrin and ivermectin in Andhra Pradesh, south India. Fourteen adult female tick populations were collected from naturally infested dogs at veterinary clinics, residence and stray dog in nine state municipal corporations/municipalities. Information on the history of dog treatment with acaricides was obtained by interviewing dog owners. The larval packet test (LPT) and larval immersion test (LIT) were used on the larvae of ticks to test for resistance to cypermethrin and ivermectin, respectively. Mortality data of each tick population was analyzed by probit analysis. Corresponding to the most susceptible population, thirteen (92.6%) and six (42.9%) tick populations were regarded as resistant to cypermethrin and ivermectin, respectively. The phenotypic level of cypermethrin (resistance ratios at LC50% varied from 1.55 to 13.29) and ivermectin (resistance ratios at LC50% ranged from 1.16 to 4.79) resistance varied distinctly between the populations. Additionally, all the populations were tested using PCR to demonstrate the frequency of the point mutation in sodium channel gene that corresponds with resistance of R. sanguineus s.l. to cypermethrin. A nucleotide substitution (T2134C) on domain III segment VI of the sodium channel gene was noticed in phenotypically resistant tick populations on DNA sequencing. Ivermectin resistance in the brown dog ticks is reported for the first time from India.

In Vitro Anthelmintic Activity of Three Tropical Plant Extracts on Haemonchus contortus.

AIM: To evaluate the anthelmintic activity of Anacardium occidentale shell, Illicium verum fruit, and Artocarpus heterophyllus seed to substantiate their traditional use against helminths. MATERIALS AND METHODS: In vitro anthelmintic activity of hydroalcoholic extracts of three plants was evaluated against eggs, infective larvae (L3), adult, and lactate dehydrogenase (LDH) of Haemonchus contortus of naturally infected sheep. RESULTS: The three extracts exhibited significant (P < 0.001) dose-dependent anthelmintic responses by inhibiting egg hatching and causing paralysis of larvae and mortality of worms. The extracts were most effective on egg and adult stage of H. contortus than on L3 stage at a higher concentration of 6 mg/mL. Anacardium occidentale shell exhibited maximum activity with 100% paralysis of L3 larvae. Probit analysis revealed that the extracts of A. Occidentale shell induced 50% egg hatch inhibition (LD50 = 0.0255 mg/mL), larval paralysis (LD50 = 0.196 mg/mL), and adult worm mortality (LD50 = 1.0365 mg/mL) at a lower concentration (LD50) compared with those of I. verum fruit and A. heterophyllus seed extracts. Further, all extracts significantly (P < 0.01) inhibited the LDH activity catalyzing the oxidation of lactate in adult H. contortus, with maximum level of inhibition caused by A. occidentale shell extract. CONCLUSION: Phytochemical screening of the extracts revealed the presence of alkaloids, flavonoids, tannins, saponins, and amino acids that could be responsible for the anthelmintic effects noticed. The results warrant further in vivo evaluation of these plants for potential use as anthelmintic agents.

Studies on prevalence and histomorphology of Sarcocystis species infecting cattle in Andhra Pradesh, India.

The present study was carried out to determine the prevalence of bovine sarcocystosis and identify the different species of Sarcocystis cysts in cattle in different regions of Chittoor district, Andhra Pradesh. A total of 150 slaughtered cattle over a period of 1 year were examined both macroscopically and microscopically for the presence of Sarcocystis infection. Out of 150 slaughtered cattle examined, macroscopic sarcocysts were observed in nine cattle and were exclusively found in oesophagus. Examination of tissue samples collected from different predilection sites from each of 150 cattle by pepsin-HCl digestion method revealed presence of live banana shaped bradyzoites under light microscopy in 134 cattle. Six cattle that were positive by macroscopic examination were also positive by microscopic examination of tissues. The overall prevalence of Sarcocystis infection in cattle of Chittoor district was 91.33%. The prevalence of macroscopic and microscopic sarcocysts was 6.57 and 93.43% respectively. Statistically a significant relationship between the prevalence of infection among different age groups (P < 0.001) and no significant relationship (P > 0.001) between the prevalence of Sarcocystis infection in male (91.76%) and female (90.76%) cattle was observed. In both male and females the prevalence of Sarcocystis infection increased with age. Microscopically sections of tissues from cattle that were positive by tissue digestion technique revealed thin walled cysts (4.5 ± 0.5 µm) in 131 samples and thick walled cysts (0.5 ± 0.12 µm) in six cattle.

Flow-Through Assay for Detection of Antibodies Using Protein-A Colloidal Gold Conjugate as a Probe.

Flow-through assay (FTA) is a rapid, simple-to-perform, cost-effective, and user-friendly diagnostic test for monitoring infections in non-laboratory settings. It is mostly applied for antibody detection. FTA employing protein-A colloidal gold conjugate to detect antibodies against porcine cysticerci using cyst fluid and whole cyst antigens of Taenia solium metacestode is described here. Antibodies in the serum are captured by an antigen spotted onto a nitrocellulose membrane mounted on a flow-through device that serves as the antigen capture matrix. The bound antibodies are visualized by the addition of protein-A colloidal gold conjugate, which imparts a pink color. The test can be completed within 3 min at room temperature without any instrumentation. The sensitivity and specificity of the FTA are in agreement with ELISA.