In vivo and in vitro apoptosis induced by new acaricidal ethyl-carbamates in Rhipicephalus microplus.

The ability of ethyl-4-bromophenylcarbamate (LQM 919) and ethyl-4-chlorophenylcarbamate (LQM 996) to induce in vivo apoptosis of Rhipicephalus microplus ovarian cells and in vitro apoptosis of tick and mammalian cell culture was evaluated. The ovaries of engorged females treated with 1 mg mL-1 LQM 919 or LQM 996 presented more (p < 0.001) peroxidase-TUNEL-positive labeled cells (apoptotic cells) in situ than their respective control groups, and this increase was time-dependent (p < 0.001). The majority of apoptotic cells were observed in the epithelium and ovarian pedicel. HepG2, Vero and Rm-sus cells, as well as cells from primary cultures of R. microplus salivary glands, intestine and ovaries were exposed to different concentrations of the ethyl-carbamates. Both ethyl-carbamates induced a concentration-dependent reduction in the viability of all cell types (p < 0.001). Exposure to the ethyl-carbamates increased caspase 3 activity (p < 0.01) in primary cultures and cell lines, except in HepG2 cells. Fluorescent TUNEL-positive cells were observed in all cell types treated with 600 µM LQM 919 or LQM 996. These results indicate that both ethyl-carbamates induce apoptosis of the ovarian, intestinal and salivary glands cells in R. microplus and strongly suggest that this is their main mechanism of acaricidal action.

Comparative Hemolymph Proteomic and Enzymatic Analyses of Two Strains of Rhipicephalus (Boophilus) microplus Ticks Resistant and Susceptible to Ixodicides.

The cattle tick Rhipicephalus (Boophilus) microplus is one of the most harmful ectoparasites affecting bovines worldwide. It represents a major threat to livestock industry due to the economic losses caused and diseases associated with these ticks. The most important tick control strategy has been the use of ixodicides, resulting in chemically resistant tick populations. It is necessary to understand the mechanisms that result in resistance so as to create new strategies increasing the lifespan of ixodicides or finding alternative targets to produce new acaricides. In this paper, in order to obtain an insight into the mechanisms that govern ixodicides resistance, we will compare the hemolymph proteome of two tick R. microplus strains, one susceptible (MJ) and one resistant (SA) to ixodicides, using HPLC and 2D electrophoresis. Significant differences were found in protein content between strains using HPLC. 2D electrophoresis revealed that 68 hemolymph protein spots were common between strains; however, 26 spots were unique to the susceptible strain MJ and 5 to the resistant strain SA. The most distinctive protein spots on the preparative gels were selected for further analyses. Nine protein spots were identified by mass fingerprinting, revealing proteins that may have a role in the ixodicides resistance or susceptibility. In this paper, we present the tick hemolymph proteome revealing a set of proteins which suggest a possible role in tick detoxification.

Draft Genome Sequence of Enterococcus casseliflavus PAVET15 Obtained from the Oviduct Infection of the Cattle Tick (Rhipicephalus microplus) in Jiutepec, Morelos, Mexico.

Enterococcus spp. are Gram-positive lactic acid-producing bacteria found in the intestinal tracts of animals, like mammals, birds, and arthropods. Enterococcus spp. may cause oportunistic infections in vertebrate and invertebrate hosts. We report here the draft genome sequence of Enterococcus casseliflavus PAVET15 containing 3,722,480 bp, with 80 contigs, an N50 of 179,476 bp, and 41.93% G+C content.

Complete Genome Sequence of Paenibacillus larvae MEX14, Isolated from Honey Bee Larvae from the Xochimilco Quarter in Mexico City.

Paenibacillus larvae strain MEX14 is a facultative anaerobic endospore-forming bacterium that infects Apis mellifera larvae. Strain MEX14 was isolated from domestic bee larvae collected in a backyard in Mexico City. The estimated genome size was determined to be 4.18 Mb, and it harbors 4,806 protein coding genes (CDSs).

Increase of carboxylesterase activity in Fasciola hepatica recovered from triclabendazole treated sheep.

In the present work, we evaluate in vivo the activity of carboxylesterase of Fasciola hepatica exposed to triclabendazole. We observed a statistically significant increase in enzyme activity at 24 and 48 h post treatment (P<0.01 and P<0.001, respectively). The zymogram of cytosolic fractions identified a protein of 170 kDa containing the carboxylesterase activity. The densitograms of the zymograms confirmed the phenomenon of enzyme induction under the experimental conditions of the assay. These results provide not only the understanding of the importance of this metabolic pathway in flukes but carboxylesterase would also be an enzyme that could participate more actively in the development of anthelmintic resistance at TCBZ.

Natural occurrence of lethal aspergillosis in the cattle tick Rhipicephalus (Boophilus) microplus (Acari:Ixodidae).

The purpose of this study was to describe an unreported entomopathogenic fungus that naturally infects the cattle tick Rhipicephalus (Boophilus) microplus (Acari: Ixodidae). Engorged female ticks, showed symptoms of fungal infection after controlled tick infestation of cattle. Infected ticks developed a distinctive dark colour, a pale mould grew over the cuticle and the ticks eventually died covered with fungal conidiophores. The responsible fungus was isolated and cultured on mycological medium and submitted to microscopic morphology, biochemical phenotyping and 18S rRNA ribotyping analyses, which identified it as aflatoxin-producing Aspergillus flavus. Spores from the cultured fungus were experimentally sprayed over healthy engorged female ticks, obtaining an 80% prevalence of experimental infection of healthy ticks and their egg masses, the larval progeny after incubation under laboratory conditions was also infected. These results demonstrate that A. flavus is the causative agent of the natural fungal disease of the cattle tick R. microplus described here.

Megaselia scalaris reared on Rhipicephalus (Boophilus) microplus laboratory cultures.

Different laboratory cultures of the acarine tick Rhipicephalus (Boophilus) microplus (Canestrini, 1888) (Ixodida: Ixodidae) were infested by small Megaselia scalaris (Loew, 1866) (Diptera: Phoridae) flies. Larvae of this species exhibited opportunistic parasitism predominantly on engorged female ticks, causing severe damage to their cuticle through which the flies were able to reach R. microplus internal organs, on which they fed until developing into pupae in the tick's remains. The flies were kept by continuous propagation on fresh ticks over six generations during which the same parasitoid behaviour was observed. Here we report on an ixodid tick laboratory culture used for rearing M. scalaris.

Bovine anaplasmosis prevalence in northern Veracruz state, Mexico.

In order to learn more about the presence of bovine anaplasmosis in northern Veracruz state, México, paired blood and serum samples from 368 cattle were subjected to polymerase chain reaction (PCR) and complement-fixation test (CFt). The overall prevalence of Anaplasma marginale by PCR was 69.2% and seroprevalence by CFt 54.6%. Age-specific prevalence was calculated for each test. Sixty-eight percent of animals from 0 to 3 months of age already were infected (PCR-positive), compared to only 42.4% positive by serology. CFt results suggested that presence of antibody increases with age up to 18 to 36 months, decreasing thereafter. Presence of the rickettsia seems to follow the same early pattern but with a new increase in animals 36 months or older. Serology results provided a biased picture of the true prevalence of anaplasmosis. Calculated specificity and sensitivity (63.5% and 68.2%) for CFt using PCR values as true values, appear very low and unreliable. The data generated by DNA-based surveys seem more appropriate to help design and implement control or eradication programs for bovine anaplasmosis.