Recombinant cysteine proteinase as anti-tick targeting Hyalomma asiaticum infestation.

Cysteine proteases are involved in the digestion of host blood and the degradation of yolk proteins of arthropod ectoparasites. In this study, a cathepsin L-like cysteine proteinase gene (HasCPL) of Hyalomma asiaticum was cloned, and recombinant (r)HasCPL protein was generated for immunization study. Bioinformatic analysis confirmed HasCPL was a member of the papain family (clan CA) and have high sequence identities with CPLs of other Ixodid ticks. The efficacy of immunization against H. asiaticum infestations in rabbits was assessed. Rabbits (n = 3) were immunized three times with rHasCPL before challenged with 250 larvae per rabbit four weeks post-immunization. A high antibody titer was detected in immunized rabbits in comparison to control. Western blot analysis detected CPLs in midgut, salivary gland, and ovary. Increase of rejection percentage of larvae were noted in ticks fed on immunized animals in comparison to control. Overall, a 55.09% protection against larva ticks was noted.

MaxEnt Modeling of Dermacentor marginatus (Acari: Ixodidae) Distribution in Xinjiang, China.

Dermacentor marginatus Sulkzer is a common tick species found in the Xinjiang Uygur Autonomous Region (XUAR) of China, and is a vector for a variety of pathogens. To determine the potential distribution of this tick species in Xinjiang, a metadata containing 84 D. marginatus presence records combined with four localities from field collection were used for MaxEnt modeling to predict potential distribution of this tick species. Identification of tick samples showed 756 of 988 (76%) were D. marginatus. MaxEnt modeling results indicated that the potential distribution of this tick species was mainly confined to northern XUAR. Highly suitable areas included west side of Altay mountain, west rim of Junggar basin, and Yili River valley in the study area. The model showed an AUC value of 0.838 ± 0.063 (SD), based on 10-fold cross-validation. Although tick presence records used for modeling were limited, this is the first regional tick distribution model for D. marginatus in Xinjiang. The model will be helpful in assessing the risk of tick-borne diseases to human and animals in the region.

Caracterization of glutathione S-transferase of Dermacantor marginatus and effect of the recombinant antigen as a potential anti-tick vaccine.

Dermacentor marginatus is one of the main tick species in northwestern China, and is a vector of various tick-borne pathogens. Tick control method largely depends on chemical agents, but the disadvantages of using such approach would cause environmental damage and the risk of developing tick resistance to acaricides. Vaccination of tick protective antigen is an eco-friendly approach which is an alternative and promising method to mitigate tick infestation in livestock. In the study, a mu-class glutathione S-transferase (GST) sequence of D. marginatus was cloned and the recombinant protein (rDmGST) was expressed. Transcriptional level of the GST was measured together with native GST activity of the tick. Finally, A vaccine trial on rabbits against D. marginatus was proceeded to evaluate the anti-tick effect of rDmGST. Results reveled that the CDs of the D. margiantus glutathione S-transferase mu 1 gene has 669 base pair nucleotide sequence encoding a 223 amino acid. The deduced GST protein sequence had over 95 % similarity with that of D. variabilis. The rDmGST was efficiently expressed soluble and purified by His trap affinity chromatography. Enzyme activity of native GST and transcriptional profiles of the GST showed up-regulation in different stages and organs of D. marginaus during blood feeding. Polyclonal antibody reacted with rDmGST in Western blotting. Tick challenge on rDmGST inoculated rabbits showed reductions in adult female engorgement rate, total egg mass and egg hatching rate with an overall vaccine efficacy of 43.69 %. The results of the experiment indicated the GST has potential value to be an effective protective antigen of D. marginatus.

Preparation of monoclonal antibodies against Bc48 and development of a rapid detection assay for infection with Babesia caballi in China.

Babesia caballi (Nuttal, 1910) is one of the causative agents of equine piroplasmosis which causes economic losses to horse industry in China. There is an urgent need for rapid detection method for B. caballi infection in Xinjiang Province, China. To prepare monoclonal antibodies (mAbs) against Bc48 gene of B. caballi (Xinjiang local strains) and establish colloidal gold-immunochromatographic (ICT) assay for diagnosis of the disease, recombinant Bc48 was expressed and purified from Escherichia coli. With purified Bc48 as immunogen in mice, three hybridoma cells named 11F4, 1H2 and 7F4 secreting mAbs against Bc48 of B. caballi were obtained, which showed strong reaction with recombinant Bc48 and Bc48 gene transfected cells. Furthermore, colloidal gold labelled ICT assay based on purified Bc48 recombinant antigen and its mAb was developed. The ICT assay showed high sensitivity and specificity and no cross-reaction with Theileria equi (Laveran, 1901). Total of 56 horse serum samples collected from Xinjiang were tested by ICT and compared with the detection by commercial ELISA kit. The results showed that 32 out of 56 serum samples were positive by ICT and 33 were positive by ELISA. ICT assay had high coincidence (98%) to the reference ELISA kit. mAbs and ICT developed in this study could be provided as an efficient diagnosis tool for infection with B. caballi in horse in Xinjiang area.

Photooxidative damage in retinal pigment epithelial cells via GRP78 and the protective role of grape skin polyphenols.

Blue light induced oxidative damage and ER stress are related to the pathogenesis of age-related macular degeneration (AMD). However, the mechanism of blue light-induced damage remained obscure. The objective of this work is to assess the photooxidative damage to retinal pigment epithelial cells (RPE) and oxidation-induced changes in expression of ER stress associated apoptotic proteins, and investigate the mechanism underlying the protective effects of grape skin extracts. To mimic lipofuscin-mediated photooxidation in vivo, ARPE-19 cells that accumulated A2E, one of lipofuscin fluorophores, were used as a model system to investigate the mechanism of photooxidative damage and the protective effects of grape skin polyphenols. Exposure of A2E containing ARPE-19 cells to blue light resulted in significant apoptosis and increases in levels of GRP78, CHOP, p-JNK, Bax, cleaved caspase-9, and cleaved caspase-3, indicating that photooxidative damage to RPE cells is mediated by the ER-stress-induced intrinsic apoptotic pathway. Cells in which GRP78 had been knocked down with shRNA were more vulnerable to photooxidative damage. Pre-treatment of blue-light-exposed A2E containing ARPE-19 cells, with grape skin extracts, inhibited apoptosis, in a dose dependent manner. Knockdown GRP78 blocked the protective effect of grape skin extracts.

Anti-yeast activity of a food-grade dilution-stable microemulsion.

The anti-yeast activities of a food-grade dilution-stable microemulsion against Candida albicans and Saccharomyces cerevisiae have been studied. The weight ratio of the formulated microemulsion is glycerol monolaurate (GML)/propionic acid/Tween 80/sodium benzoate (SB)/water = 3:9:14:14:24. Results of anti-yeast activity on solid medium by agar diffusion method showed that the anti-yeast activity of the microemulsion at 4.8 mg/ml was comparable to that of natamycin at 0.1 mg/ml as positive control. Results of anti-yeast activity in liquid medium by broth dilution method showed that the growth of both C. albicans and S. cerevisiae was completely inhibited when the liquid medium containing 10(6) cfu/ml was treated with 1.2 mg/ml microemulsion, which was determined as minimum fungicidal concentration. The kinetics of killing results showed that the microemulsion killed over 90% yeast cells rapidly within 15 min and caused a complete loss of viability in 120 min. Among the components, SB and GML had a similar anti-yeast activity, followed by propionic acid, while Tween 80 exhibited no activity and could not enhance the anti-yeast activities of these components, and it was revealed that the anti-yeast activity of the microemulsion was attributed to a combination of propionic acid, GML, and SB. The anti-yeast activity of the microemulsion was in good agreement with the leakage of 260-nm absorbing materials and the observation of transmission electron microscopy, indicating that the microemulsion induced the disruption and dysfunction of the cell membrane.

Constructing biological networks through combined literature mining and microarray analysis: a LMMA approach.

MOTIVATION: Network reconstruction of biological entities is very important for understanding biological processes and the organizational principles of biological systems. This work focuses on integrating both the literatures and microarray gene-expression data, and a combined literature mining and microarray analysis (LMMA) approach is developed to construct gene networks of a specific biological system. RESULTS: In the LMMA approach, a global network is first constructed using the literature-based co-occurrence method. It is then refined using microarray data through a multivariate selection procedure. An application of LMMA to the angiogenesis is presented. Our result shows that the LMMA-based network is more reliable than the co-occurrence-based network in dealing with multiple levels of KEGG gene, KEGG Orthology and pathway. AVAILABILITY: The LMMA program is available upon request.