Rhipicephalus bursa Sialotranscriptomic Response to Blood Feeding and Babesia ovis Infection: Identification of Candidate Protective Antigens.

Ticks are among the most prevalent blood-feeding arthropods, and they act as vectors and reservoirs for numerous pathogens. Sialotranscriptomic characterizations of tick responses to blood feeding and pathogen infections can offer new insights into the molecular interplay occurring at the tick-host-pathogen interface. In the present study, we aimed to identify and characterize Rhipicephalus bursa salivary gland (SG) genes that were differentially expressed in response to blood feeding and Babesia ovis infection. Our experimental approach consisted of RNA sequencing of SG from three different tick samples, fed-infected, fed-uninfected, and unfed-uninfected, for characterization and inter-comparison. Overall, 7,272 expressed sequence tags (ESTs) were constructed from unfed-uninfected, 13,819 ESTs from fed-uninfected, and 15,292 ESTs from fed-infected ticks. Two catalogs of transcripts that were differentially expressed in response to blood feeding and B. ovis infection were produced. Four genes coding for a putative vitellogenin-3, lachesin, a glycine rich protein, and a secreted cement protein were selected for RNA interference functional studies. A reduction of 92, 65, and 51% was observed in vitellogenin-3, secreted cement, and lachesin mRNA levels in SG, respectively. The vitellogenin-3 knockdown led to increased tick mortality, with 77% of ticks dying post-infestation. The reduction of the secreted cement protein-mRNA levels resulted in 46% of ticks being incapable of correctly attaching to the host and significantly lower female weights post-feeding in comparison to the control group. The lachesin knockdown resulted in a 70% reduction of the levels associated with B. ovis infection in R. bursa SG and 70% mortality. These results improved our understanding of the role of tick SG genes in Babesia infection/proliferation and tick feeding. Moreover, lachesin, vitellogenin-3, and secreted cement proteins were validated as candidate protective antigens for the development of novel tick and tick-borne disease control measures.

Detection and phylogenetic characterization of Theileria spp. and Anaplasma marginale in Rhipicephalus bursa in Portugal.

Ticks are obligatory blood-sucking arthropod (Acari:Ixodida) ectoparasites of domestic and wild animals as well as humans. The incidence of tick-borne diseases is rising worldwide, challenging our approach toward diagnosis, treatment and control options. Rhipicephalus bursa Canestrini and Fanzago, 1877, a two-host tick widely distributed in the Palearctic Mediterranean region, is considered a multi-host tick that can be commonly found on sheep, goats and cattle, and occasionally on horses, dogs, deer and humans. R. bursa is a species involved in the transmission of several tick-borne pathogens with a known impact on animal health and production. The aim of this study was to estimate R. bursa prevalence in Portugal Mainland and circulating pathogens in order to contribute to a better knowledge of the impact of this tick species. Anaplasma marginale and Theileria spp. were detected and classified using phylogenetic analysis. This is the first report of Theileria annulata and Theileria equi detection in R. bursa ticks feeding on cattle and horses, respectively, in Portugal. This study contributes toward the identification of currently circulating pathogens in this tick species as a prerequisite for developing future effective anti-tick control measures.

Coxiella burnetii DNA detected in domestic ruminants and wildlife from Portugal.

Coxiella burnetii is the etiological agent of Q fever or Coxiellosis, a zoonosis mainly affecting domestic ruminants. Information on the population structure and epidemiology of C. burnetii in animals is scarce in Portugal. Evidence of C. burnetti infection was sought in domestic, wild and captive animals based on the detection of bacterial DNA. Tissue samples from 152 domestic animals (cattle=24, goats=51, sheep=76 and swine=1), 55 wild carnivores (Egyptian mongoose=45, red fox=4, common genet=3, weasel=2 and European badger=1) and 22 zoo animals (antelopes=15, impala=1; rhinoceros=1, deer=2, zebras=2 and giraffe=1) were screened by nested-touchdown PCR. Cloacae swabs from 19 griffon vultures were also analysed. Among the domestic ruminants, goats presented the highest prevalence of infection (23.53%), followed by cattle, (20.83%) and sheep (10.53%). C. burnetii DNA was also detected in five Egyptian mongooses and two antelopes and one giraffe. Using a 6-locus multiple-locus variable-number tandem repeat analysis (MLVA-6) six complete genotypes, T, I and CM and the first reported CN, CO and CP, were identified, respectively, in small ruminants and Egyptian mongooses. Clustering analysis of genotypes exposed four distinct groups, according to detection source, enlightening an apparent association between C. burnetii genotype and host.

Evaluation of the nutritive value of muiumba (Baikiaea plurijuga) seeds: chemical composition, in vitro organic matter digestibility and in vitro gas production.

One of the main constraints hindering the increase of animal production in semi-arid regions of Africa is the inadequate supply of nutrients during the dry season. Incorporation of alternative feed resources in ruminant diets during this period could be a viable approach to overcome these limitations. The objective of this study was to evaluate the nutritive value of muiumba (Baikiaea plurijuga) tree seeds as an alternative nutrient source for ruminants. Muiumba seeds were compared to other eight feedstuffs including two cereal grains (corn and oat), two wheat by-products (wheat bran and distilled wheat) and four protein meals (coconut meal, sunflower meal, soybean meal and rapeseed meal) as to its chemical composition, in vitro organic matter digestibility (IVOMD) and in vitro gas production. The moderate crude protein concentrations (145 g/kg DM) of muiumba seeds indicate that this feedstuff could not be used as a protein supplement, contrarily to the majority of multipurpose tree seeds. Although the starch content was scarce (15 g/kg DM), the low neutral detergent fibre (235 g/kg DM), low molecular weight sugar (76.1 g/kg DM) and non-starch polysaccharide (510.5 g/kg DM) contents indicate that this feedstuff has potential feeding value. This was confirmed by the IVOMD (0.770) and by the data provided by the in vitro gas production showing that muiumba seeds had high (P < 0.05) maximum gas production and fractional fermentation rates, suggesting that these seeds are characterized by a highly fermentable fraction.

PCR-based survey of Anaplasma phagocytophilum in Portuguese ticks (Acari: Ixodidae).

A total of two-thousand and six ticks, collected from 2002 to 2006 in areas belonging to seven districts of Mainland Portugal and also in Madeira Island, were examined by polymerase chain reaction (PCR) for the presence of Anaplasma phagocytophilum. Active infections were detected exclusively in Ixodes species, including six questing I. ricinus nymphs from Madeira Island, one questing I. ventalloi nymph from Setúbal District, and two I. ventalloi adults found parasitizing domestic cats in both Setúbal and Santarém District. These findings confirm prior observations and suggest the persistence of A. phagocytophilum on Madeira Island. Moreover, it adds I. ventalloi and domestic cats to the list of potential elements of the agent's enzootic cycles in Portugal. Molecular analysis of PCR amplicons suggests the existence of two A. phagocytophilum genotypes in Portugal, one of which is identical or very similar to North American strains implicated in human disease.