Assessment of consistency of minimum post-mortem intervals estimated by thermal summation-based methods in medico-legal cases associated with blowflies.

BACKGROUND: This study is based on blow fly samples collected from 8 medico-legal cases in Tamil Nadu, India. The fly life stages were identified and the consistency of minimum post-mortem intervals (PMImin) estimated by different thermal summation-based methods was assessed. METHODS: PMImin of 8 medico-legal cases was estimated using six different thermal summation constants and lower developmental temperatures that are based on C. megacephala and C. rufifacies developmental data. Limits of agreement (LoA), intra class correlation coefficient (ICC) between PMImin values and margin of error of mean of difference between PMImin values were calculated. RESULTS: Intra-class correlation between the PMImin values estimated using different thermal summation constants based on C. megacephala ranged between 0.89 and 0.98 and coefficient of determination ranged between 0.93 and 0.98. Intra-class correlation between the PMImin values estimated using different thermal summation constants based on C. rufifacies ranged between 0.91 and 0.99 and coefficient of determination ranged between 0.95 and 0.99. The mean difference of PMImin values estimated using different thermal summation methods based on C. megacephala ranged between 1.8 hr and 6.6hr and margin of error ranged between 2.51 and 6.93hr. The mean difference of PMImin values estimated using different thermal summation methods based on C. rufifacies ranged between 3.33 and 31.33hr and margin of error ranged between 4.66 and 32hr. CONCLUSION: Consistency of PMImin values estimated by different thermal summation methods was good to excellent. Thermal summation constants useful in estimation of PMImin with lowest mean difference and margin of error were described.

Prognostic markers and their discriminant score in predicting the outcome of Babesia gibsoni infection.

BACKGROUND: We aimed to identify prognostic markers and their discriminant score in predicting the lethal outcome of canine Babesia gibsoni. METHODS: Blood samples were collected from 108 client-owned dogs with clinical signs commensurate with babesiosis to analyze haematological, biochemical, haemostatic, antioxidant profile and thiobarbituric acid reactive substance levels. Samples were screened for Babesia infection (microscopic and molecular techniques). Babesiosis-affected dogs were classified into survivors and non-survivors, and 30 healthy dogs were used in the control group. RESULTS: Haemoglobin, thrombocytes, catalase, urea, creatinine, alanine aminotransferase (ALT), alkaline phosphatase, lactate and reticulocytes were highly correlated to survival. Receiver operating characteristics analysis revealed urea, ALT and lactate as specific prognostic markers for the disease. The formula for calculation of discriminant scores (Di) for lethal outcome of the disease was generated with cut-off score 0.141. The scoring system was 79% sensitive and 83% specific in predicting the lethal outcome of the disease. CONCLUSIONS: A scoring system developed from the prognosticating markers may aid in predicting the outcome of Babesia gibsoni infection on the day of presentation itself enabling intensive care for those animals with a cut-off score more than 0.141.

Retraction Note to: Antigen based detection of cystic echinococcosis in buffaloes using ELISA and Dot-EIA.

[This retracts the article DOI: 10.1007/s12639-016-0762-9.].

Dirofilaria hongkongensis - A first report of potential zoonotic dirofilariosis infection in dogs from Tamil Nadu.

Dirofilariosisis, is a mosquito borne anthropo-zoonotic disease caused by the filarial nematodes of the genus Dirofilaria. The current paper reports a species of Dirofilaria, which has not been recorded in Tamil Nadu, in addition to differentiating it from the existing species of Dirofilaria spp. through morphological and molecular techniques. Micrometry of the microfilaria detected by Knott's technique as well as in Giemsa stained blood smears revealed that the length was on an average 280 µm to 310 µm whereas the width was 5 to 8 µm. The microfilariae were unsheathed with blunt heads and tapering tails. For molecular confirmation, the blood sample was subjected to genomic DNA isolation. PCR amplification using specific primers for D. hongkongensis was carried out using thermal cycler with the isolated genomic DNA as template. Molecular analyses revealed the band size of about 230 bp which confirmed the presence of D. hongkongensis. Phylogenetic analysis was done using UPGMA and MLM method. Dirofilaria is recognized as a zoonotic entity throughout the world, and dirofilariasis should be considered an emerging zoonosis in southern India. The control of ubiquitous mosquito population is essential to prevent the transmission of microfilaria from animals to arthropods and from arthropods to humans.

Innovative way to dispense pheromones for off-host control of Rhipicephalus sanguineus sensu lato ticks.

Vapour patches dispensing pheromones were evaluated as lures to increase the attractiveness of sticky tick traps for Rhipicephalus sanguineus sensu lato (s.l.). Sex pheromone (SP), assembly pheromone (AP) and a combination of SP + AP at optimal concentrations were impregnated in vapour patches. The responses of the different stages of R. sanguineus s.l. (i.e. larvae, nymphs and adults) to the pheromones were evaluated using a Petri dish bioassay. The impregnated vapour patches were retained as such for a period of two mo and their efficacy was reassessed. In a subsequent field trial, pheromone impregnated vapour patches were placed as lures in bamboo (Bambusa vulgaris) sticky traps designed for the control of ticks in dog kennels. In vitro AP impregnated vapour patches were effective in attracting the different life stages of R. sanguineus s.l. whereas SP was effective in attracting the unfed and fed male stages of R. sanguineus s.l. The field trial revealed that questing and engorged larvae, nymphs and females of R. sanguineus s.l. were attracted more towards AP impregnated vapour patches than SP and AP + SP impregnated vapour patches. Fed and unfed male ticks were lured effectively by SP alone. The combination of SP + AP revealed no potent additive or synergistic effect.

Comparison of in-vitro bioassays for evaluation of the response of different stages of Rhipicephalus sanguineus sensu lato to calcium alginate encapsulated pheromone beads.

Tick sex pheromone (SP), assembly pheromone (AP) and their combination (SP + AP) were encapsulated in calcium alginate beads. In vitro bioassays, namely Petri dish and olfactometer assays, were employed to estimate the level of attraction of the various stages of Rhipicephalus sanguineus sensu lato, namely unfed and engorged (fed) larvae, nymphs, females, and males to the different pheromones. The study revealed that only the Petri dish assay was suitable to evaluate the response of larval stages whereas the olfactometer bioassay could also be used for evaluating the response of all other stages. Attraction to pheromone encapsulated calcium alginate beads of all tick stage was higher in the Petri dish assay than in the olfactometer assay.

Hypodermosis in cattle translocated to Tamil Nadu from Punjab.

Fifteen apparently healthy Kandari cross-bred cattle aged about 4 years were purchased from Rurki, Patiala district of Punjab by a private dairy farmer in Erode, Tamil Nadu. Four animals showed eruptions on the lateral thoracic and dorsal abdominal regions of the body after 15-day period of quarantine. Manual palpation of the eruptions resulted in the emergence of larval stages of dipteran flies, identified by their morphology as Hypoderma from these animals. Molecular identification based on mitochondrial cytochrome oxidase-1 (COX-1) gene confirmed it to be Hypoderma lineatum. Treatment with oral ivermectin did not have any curative effects, with exacerbation of disease being noticed, as more than 500 eruptions subsequently emerged in each animal, which had to be culled. Consequences of long distance migration of host on parasite epidemiology are discussed. Awareness must be created among livestock farmers to prevent their economic loss while purchasing cattle from different parts of the country.

Evaluation of multiplex PCR assay for detection of Babesia spp, Ehrlichia canis and Trypanosoma evansi in dogs.

A multiplex PCR test was evaluated to detect the DNA of three important dog haemoparasites by comparing with singular PCR counterpart on clinical blood samples of dogs in and around Chennai, Tamil Nadu, India. Initial screening of samples was done by microscopic examination of peripheral blood smear and singular PCR and those found exclusively positive for Babesia spp, Ehrlichia canis and Trypanosoma evansi and concurrent infections were used to standardize multiplex PCR. Amplicons of 619 bp, 377 bp and 227 bp corresponding to Babesia spp (18S rRNA gene), E. canis (VirB9 gene), and T.evansi (VSG gene) respectively were amplified, without any non-specific amplification. The laboratory sensitivity (91.7% to 100%) and specificity (100%) of the multiplex PCR were calculated using 'true positive' and 'true negative' dog blood samples obtained in the initial screening process. Clinical blood samples from 287 dogs were screened using singular PCR and multiplex PCR tests for the presence of genome of Babesia spp, E. canis and T. evansi. The multiplex PCR was found to have high level of diagnostic specificity (97.5%-100%) in the detection of all three dog blood parasites and high level of diagnostic sensitivity (95%) in the detection of T. evansi from field level clinical blood samples compared to the singular PCR. However, the diagnostic sensitivity of the multiplex PCR was found to be low to moderate (40.45%-66.7%) in detection of Babesia spp and E. canis from field level clinical blood samples. The strength of agreement between singular and multiplex PCR assays was 'moderate' (0.445), 'good' (0.708) and 'very good' (0.968) in detection of DNA of Babesia spp, E. canis and T. evansi. The multiplex PCR was found to be 10 fold less sensitive in comparison with the singular PCR counterpart.

Prevalence and staining characteristics of Blastocystis isolates from food animals in Tamil Nadu.

The present study was undertaken to investigate the prevalence and staining characteristics of Blastocystis isolated from food animals. Smears of the duodenal and caecal mucosal scrapings, collected from food animals, were stained with Giemsa, Gram's, modified acid-fast and acridine orange. Blastocystis was identified in 295 samples, including faeces and intestinal contents of animals like small ruminants (95), poultry (170) and pigs (30). The prevalence in pigs was found to be high (94.4%) followed by poultry (29.4%) and small ruminants (14%). Various forms of Blastocystis such as vacuolar, granular and amoeboid forms were identified by using different stains. The parasites stained with Giemsa were identified by the presence of eosinophilic nucleus and basophilic cytoplasm. In organisms stained with Gram's stain, the cytoplasm of the vacuolar forms took up the counter stain safranine. Blastocystis appeared as a pink colored cyst against bluish green background with modified acid-fast staining. The study shows that there is a very high prevalence of Blastocystis among the food animals investigated. Simple parasitological procedures, including direct microscopical examination and staining with agents like Giemsa, Gram's and acridine orange can assist identification of the parasites from intestinal contents and faecal material.

Cat fur mite Lynxacarus radovskyi in India.

A Persian cat with the complaint of lustreless dry coat, salt and pepper appearance was brought to the dermatology unit of Madras Veterinary College, Chennai. Microscopical examination of the tape impression and hair coat brushings revealed Lynxacarus radovskyi mites with eggs adhering to the hair shaft. The cat was treated with Ivermectin @ 300 µg/kg body weight subcutaneously once a week for 5 weeks after getting owner's consent. The cat was free from the parasites 5 weeks post treatment. Fomite and close contact might aid the spread of infestation. Isolation coupled with full course of treatment favours complete recovery from the infestation.

Incidence of fur mite infestation in laboratory rodents.

Incidence of fur mites in laboratory rodents is reported. Among mice, infestation was found to be highest in male Swiss Albino mice followed by female Swiss Albino mice, male Balb C mice and female Balb C mice. Infestation in Guinea pigs was 50 %. None of the rats screened were found to be positive for fur mites. Species of fur mites identified were Myocoptesmusculinus in mice and Chirodiscoidescaviae in Guinea pigs. Clinical signs in affected animals are also reported.

Incidence of Onchocerca gibsoni in subcutaneous nodules of cross bred Jersey cows: case report.

Onchocerca gibsoni subcutaneous nodules in four cross bred Jersey cows aged 5-6 years brought for post mortem with calcified and caseated skin nodules in the lateral flank region is reported. Examination and dissection of these nodules revealed that they were "worm nests" of Onchocerca sp. of filarid nematodes. The worm nests were carefully dissected and filarid worms were extracted out. Multiple numbers of worms were recovered from each nodule ranging from 15 to 20. Female worms were found inside the worm nests and were found to be filled with microfilariae. The nodules weighed 2-3 g. Based on the morphological features the worms were identified as O. gibsoni.

Antigen based detection of cystic echinococcosis in buffaloes using ELISA and Dot-EIA.

Cystic echinococcosis is caused by the larval stage of the dog tapeworm, Echinococcus granulosus. The disease is recognized as one of the world's major zoonoses affecting human beings and domestic animals apart from its economic and public health importance. Development of the cysts in the intermediate host such as buffaloes occurs in the lungs, liver and other organs. In this study, detection of circulating antigen in the diagnosis of cystic echinococcosis in buffaloes was done using enzyme linked immunosorbent assay and Dot-Enzyme immunoassay (Dot-EIA). The sensitivity and specificity were determined as 89 and 92 % respectively, whereas those of Dot-EIA were determined as 94 and 96 %.

A novel assembly pheromone trap for tick control in dog kennels.

A novel ecofriendly sticky tick trap device for the control of dog tick Rhipicephalus sanguineus using gold nanoparticle assembly pheromone complex as a bait was developed. Assembly pheromones comprising of guanine, xanthine and adenine in the ratio of 25:1:1 was encapsulated in gold nanoparticle. The response of the different stages of unfed R. sanguineus ticks was evaluated using petridish bioassay. Statistical analysis was done using chi-square test. Petridish bioassay with unfed stages of R. sanguineus revealed that 100% of the larvae, nymph and adults were attracted to assembly pheromone nanogold complex within 24h. Of the 952 ticks trapped, ticks of different stages trapped in total by the baited sticky trap device, 543 (57%) were engorged and 409 (43%) were unfed ticks. The study revealed that assembly pheromone baited traps has the potential to control tick infestations in dog kennels.

Response of unfed stages of Rhipicephalussanguineus to subtle variations in the composition of assembly pheromone.

Two combinations of assembly pheromone (AP), with and without hematin were utilized as a lure for the unfed larvae, nymph and adults of Rhipicephalussanguineus ticks. In-vitro trials were carried out with the AP encapsulated in calcium alginate beads and the response of different stages of ticks were recorded. Analysis of results revealed that rapid attraction was evident in unfed larvae exposed to beads containing AP without hematin. In case of unfed nymphal and adult stages, the presence or absence of hematin did not have any impact on arrestment.

(Z)-9-Tricosene based Musca domestica lure study on a garbage dump yard using plywood sticky trap baited with fish meal.

A study was undertaken to find out the efficacy of (Z)-9-Tricosene in attracting flies in a garbage dump yard using a plywood sticky glue trap with fish meal as a food bait. (Z)-9-Tricosene was dissolved in acetone or hexane before application on a filter paper strip fixed at the centre of the trap. The traps were left in areas of the garbage dump yard of high fly activity for 6 h and then the trapped flies were counted species wise. Significantly more number of Musca domestica flies were caught in (Z)-9-Tricosene treated fish meal baited traps compared to those traps without (Z)-9-Tricosene. No significant difference was noted in trap catches in (Z)-9-Tricosene treated traps between the solvents acetone and hexane. In addition Sarcophaga sp. and Chrysomyia sp. flies were also caught in the traps.

A six year (2006-2011) retrospective study of hemoprotozoan parasites affecting dogs in Chennai, Tamil Nadu, India.

With the changing global scenario there is effect not only on the human survival but also on the prevalence of existing pathogens and appearance of new infectious agents among animal population. This paper emphasises the need for prevalence studies. A retrospective study for 6 years on prevalence of hemoprotozoans in dogs in Chennai showed Babesia gibsoni the predominant parasite of dogs. Details are discussed in this paper.

A novel approach to control brown dog tick, Rhipicephalus sanguineus using sustained release poly-ɛ-caprolactone-pheromone microspheres.

Control of brown dog tick, Rhipicephalus sanguineus was attempted by utilizing sustained release preparations of synthetic analogues of assembly pheromones. The assembly pheromone, in defined ratio, was encapsulated using poly-ɛ-caprolactone by water-in-oil-in-water double emulsion solvent evaporation technique. In the in vitro bioassay, percent mortality with test microspheres was 95.6, 64 and 44 among the unfed larvae, unfed nymph and unfed adults respectively, 24 hours post-exposure. Field trials were carried out to evaluate the efficacy of microspheres in luring and killing environmental stages of R. sanguineus in dog houses/kennels. Engorged and unfed stages in the environment were found adhered and dead on the specially designed lure.

Control of brown dog tick, Rhipicephalus sanguineus using assembly pheromone encapsulated in natural polymer, chitosan.

In the current study, an attempt was made to encapsulate assembly pheromone using natural polymer, chitosan. Chitosan beads were prepared by incorporating assembly pheromone in conjunction with an acaricide, namely, deltamethrin. In the in vitro bioassay, the test beads attracted and killed 79 % of unfed larvae, 88 % of unfed nymphs and 61 % of unfed adults of the brown dog tick, Rhipicephalus sanguineus, in 24 h of exposure. Field trials were carried out to attract and kill the pre-parasitic environmental stages. The beads were dispersed onto specially designed devices and they were placed in infested kennels. The devices were observed after 10 days.

Use of polymerase chain reaction: Restriction fragment length polymorphism to detect acaricidal resistance to synthetic pyrethroids in Boophilus microplus ticks of South India.

BACKGROUND: Boophilus microplus is an important ectoparasite of livestock. Apart from transmitting diseases, heavy tick burden can decrease production and damage hides. The synthetic pyrethroids which are advantageous over other acaricides for treatment of this infestation are now losing their efficacy due to development of resistant strains of ticks. MATERIALS AND METHODS: Boophilus microplus ticks with a previous history of acaricidal treatment especially synthetic pyrethroids (SP) such as cypermethrin, deltamethrin and flumethrin were randomly collected from different pockets of four Southern States of India namely Tamil Nadu, Kerala, Karnataka, Andhra Pradesh, and Puducherry from cattle. Deoxyribonucleic acid extracted from pooled adult B. microplus tick from each State was subjected to polymerase chain reaction-restriction fragment length polymorphism to detect point mutation in carboxyl esterase gene. RESULTS: A product size of 372 bp was obtained for cattle tick samples collected from all over Southern States of India. CONCLUSIONS: B. microplus ticks found in Southern part of India are not resistant to commonly used SP.