Parasitology Research

Ticks are blood-sucking ectoparasites with significant medical and veterinary importance, capable of transmitting bacteria, protozoa, fungi, and viruses that cause a variety of human and animal diseases worldwide. In the present study, we sequenced the complete mitochondrial (mt) genomes of five hard tick species and analyzed features of their gene contents and genome organizations. The complete mt genomes of Haemaphysalis verticalis, H. flava, H. longicornis, Rhipicephalus sanguineus and Hyalomma asiaticum were 14855 bp, 14689 bp, 14693 bp, 14715 bp and 14722 bp in size, respectively. Their gene contents and arrangements are the same as those of most species of metastriate Ixodida, but distinct from species of genus Ixodes. Phylogenetic analyses using concatenated amino acid sequences of 13 protein-coding genes with two different computational algorithms (Bayesian inference and maximum likelihood) revealed the monophylies of the genera Rhipicephalus, Ixodes and Amblyomma, however, rejected the monophyly of the genus Haemaphysalis. To our knowledge, this is the first report of the complete mt genome of H. verticalis. These datasets provide useful mtDNA markers for further studies of the identification and classification of hard ticks.

The FLOTAC flotation technique has been introduced as a new diagnostic tool to detect parasitic elements from faeces. Samples from naturally infected young deer were used for counting Dictyocaulus larvae and strongylid eggs. The FLOTAC technique, using 11 different flotation solutions with specific gravities (sg) between 1.20 and 1.45, was compared with the Baermann technique and the saturated sodium chloride (sg 1.20)-based McMaster method. In addition, a comparison was made between the FLOTAC technique with magnesium sulphate (sg 1.28) and the Baermann technique for larval recovery from faeces that were examined on the day of collection or after 7 days storage at 4°C. On the whole egg counts between the FLOTAC using different flotation solutions and the McMaster were unremarkable. In contrast, variations of larval counts were detected between different flotation solutions as well as with the Baermann technique. Most flotation solutions with a specific gravity of 1.20 floated significantly fewer lungworm larvae (p < 0.05) compared to flotation solutions with a higher specific gravity. Magnesium sulphate (sg 1.28) consistently produced the highest mean larval counts in all conducted experiments. Larval counts using magnesium sulphate (sg 1.28) were higher than with the Baermann technique both on the day of collection and after 7 days. Overall, the use of magnesium sulphate (sg 1.28) with FLOTAC for larval counts resulted in higher counts than the Baermann recovery technique and was the better choice of those flotation solutions examined. Furthermore, magnesium sulphate (sg 1.28) was also reliable for strongylid egg detection with the FLOTAC apparatus.

To develop immunoprophylactic measures against multi-tick infestation, two glycoproteins of 34 and 29 kDa were isolated from the larvae of Hyalomma anatolicum anatolicum and Boophilus microplus, respectively, and assessed for their efficacy against experimental challenge infestations. The synergistic effect of the antigens in the presence of incomplete Freund’s adjuvant was found to confer protection (DT%) in animals against 56.48% of larvae and 52% of adults of H. a. anatolicum, while the effect was 40% against adults of B. microplus. The efficacy (E%) of the antigens in combination against larvae and adults of H. a. anatolicum was calculated as 70% and 64.3%, respectively, and 63% against adults of B. microplus. A direct correlation between anti-glycoprotein antibody response and protection against infestation was observed. Western blot analysis detected specific antigen in the sera of animals of group A. The antigens in combination with incomplete Freund’s adjuvant could protect animals from H. a. anatolicum and B. microplus infestations for at least 30 weeks. The possibility of employing the vaccination strategy in Indian conditions is discussed.

Studies have demonstrated the potential of Copaifera sp. oleoresin to control Aedes aegypti proliferation. However, the low water solubility is a factor that limits its applicability. Thus, the micro- or nanoencapsulation could be an alternative to allow its use in larval breeding places. The purpose of this study was to evaluate if achievable lethal concentrations could be obtained from Copaifera sp. oleoresin incorporated into polymers (synthetic or natural) and, mainly, if it can be sustained in the residual activity compared to the pure oil when tested against the A. aegypti larvae. Microcapsules were prepared by the process of emulsification/precipitation using the polymers of cellulose acetate (CA) and poly(ethylene-co-methyl acrylate) (PEMA), yielding four types of microcapsules: MicPEMA1 and MicPEMA2, and MicCA1 and MicCA2. When using only Copaifera sp. oleoresin, the larvicidal activity was observed at concentrations of LC50 = 48 mg/L and LC99 = 149 mg/L. For MicPEMA1, the LC50 and LC99 were 78 and 389 mg/L, respectively. Using MicPEMA2, the LC50 was 120 mg/L and LC99 >500 mg/L. For microcapsules MicCA1 and MicCA2, the LC50 and LC99 were 42, 164, 140, and 398 mg/L, respectively. For a dose of 150 mg/L of pure oleoresin, the residual activity remained above 20% for 10 days, while the dose of 400 mg/L remained above 40% for 21 days. The MicPEMA1 microcapsules showed a loss in residual activity up to the first day; however, it remained in activity above 40% for 17 days. The microcapsules of MicCA1 showed similar LC50 of pure oil with 150 mg/L.

The usefulness of the polymerase chain reaction with restriction fragment length polymorphism (PCR-RFLP) and partial sequencing of the cytochrome oxidase I (COI) gene was tested regarding the utility of these techniques in unraveling philometrid life cycles and, in particular, to determine putative paratenic host species. Our focus was to study three species of philometrids commonly found in the estuaries of South Carolina: Philometroides paralichthydis and Philometra overstreeti from the southern flounder, Paralichthys lethostigma and Philometra carolinensis from the spotted seatrout, Cynoscion nebulosus. A total of 473 fish in 19 species known to be potential prey of the spotted seatrout and the southern flounder were dissected. Of all nematode larvae found in the mesenteries of 53 fish of 10 species, 21 specimens were determined to be philometrids using PCR. The use of PCR-RFLP allowed the identification of larvae of P. carolinensis and P. overstreeti in the freshwater goby, Ctenogobius shufeldti, and P. overstreeti in mummichog, Fundulus heteroclitus. However, 12 RFLP profiles could not be matched to control species, thus demonstrating the limitation of this technique in areas where diversity of philometrids is not well known and higher than anticipated. Similarly, COI procedures provided unknown sequences that did not match those of nine philometrid species used as controls. We concluded that although both techniques showed some usefulness and promise, at this point, however, they demonstrate the need of increasing our knowledge of marine and estuarine philometrid biodiversity.

The embryogenesis of the tapewormCittotaenia variabilis from the oocyte to the oncosphere is followed using scanning electron microscopy (SEM) in conjunction with ethanol cryofracture. Transmission electron micrographs and histochemistry are used to corroborate the inferences drawn from the SEM studies. Evidence is given for oocyte granules, the uterine epithelium, and the outer envelope as sources of components for the outer capsule. The contributions and senescence of the inner envelope and its relationship to the embryophore are given.

Strongyloidiasis is an important neglected disease, which is life threatening in immunocompromised patients. This study aimed to evaluate the frequency of Strongyloides stercoralis infection among immunosuppressed subjects living in endemic communities by conventional PCR of the 18S rRNA and Cox1 genes to detect cell-free DNA in the patients’ serum samples. Fresh stool and serum samples were obtained from participants. The stool samples were examined using parasitological methods. Total DNA was extracted from the serum samples and S. stercoralis larvae isolated from patient fecal samples. Conventional PCR to amplify a 101 bp fragment of the 18S rRNA gene was carried out for all extracted DNA, and then positive samples were further evaluated for a 121 bp fragment of the Cox1 gene. The PCR products of selected samples were sequenced and BLAST analysis was performed. Out of 120 patients, 57 and 63 cases had autoimmune disorders and cancer, respectively. The 101 bp fragments of the 18S rRNA were successfully amplified in 36 out of 120 (30%) serum samples. The PCR products of five samples were sequenced and compared with reference sequences in GenBank, which showed 97% identity and 90% coverage. In conclusion, to the best of our knowledge, this is the first molecular study for the detection of S. stercoralis cell-free DNA in human serum samples. These results provide useful insights for future studies and show that serum is an alternative specimen and may be useful in molecular diagnosis of diseases, particularly in immunosuppressive patients.