Objective To investigate the infection of Babesia bigemina in ticks at Shibing county in Guizhou province of China. Methods Ticks were collected from the body surface of free-range cattle in Shibing from 2021 to 2022. The 16S rDNA and cytochrome c oxidase subunit Ⅰ(COX1) gene detection techniques were used to identify the tick species after preliminary morphological classification. The samples were initially screened with Piroplasma 18S rDNA gene, followed by PCR verification and sequencing with B. bigemina apical membrane antigen-1 (AMA-1) gene. The resulting sequences were aligned using BLAST at the National Center for Biotechnology Information (NCBI) website. Phylogenetic trees were constructed by using the neighbor-joining method to understand the genetic evolution characteristics of tick-borne B. bigemina in Shibing. Results In this study, 615 ticks were collected and assigned to 2 genera and 4 species, including Rhipicephalus microplus, R. haemaphysaloides, Haemaphysalis longicornis, and H. kitaokai. Nucleic acid fragments of B. bigemina were detected in R. microplus. The sequencing result showed that the AMA-1 gene sequence was closely related to the B. bigemina detected from bovine blood in the Philippines and the tick of Hyalomma anatolicum in Sudan, and the 18S rDNA sequence was most closely related to B. bigemina in India. Conclusions The parasitic ticks on the body surface of free-range cattle in Shibing carry the nucleic acid fragments of B. bigemina, suggesting infection of the vector ticks in Shibing with B. bigemina. Our results provide reference data for the prevention and control of tick-borne protozoonosis in this area.

Objective To identify the insulin-like peptide (ILP) genes from the whole genome of Aedes albopictus, and to analyze the gene structure and its expression profiles at various developmental stages and in different tissues. Methods The ILP gene family was identified by homology comparison in the whole genome database of Ae. albopictus on the Vectorbase website. Signal peptide prediction was performed by SignalP 6.0 software. The structural features of ILP genes were analyzed by MEGA 11.0 software. The phylogenetic tree was constructed by the maximum likelihood method. Quantitative real-time PCR (qRT-PCR) was used to detect the changes in ILP gene expression at different mosquito developmental stages (eggs, fourth instar larvae, pupae, and adults) and in different tissues (head, fat body, midgut, thorax, and ovary) of female mosquitoes before and after blood feeding. SPSS 20.0 software was used for statistical analysis. The Tukey’s HSD test was used to analyze the expression of AalbILPs at different developmental stages, and the t-test was used to compare the expression of AalbILPs in different tissues before and after blood feeding. Results Seven AalbILP open reading frame sequences were identified from the whole genome of Ae. albopictus. The seven AalbILP sequences had the conserved characteristics of the insulin superfamily, and the propeptide consisted of continuous signal peptides, B, C, and A chains; AalbILP6 had a truncated C chain and carboxy-terminal extension, similar to the insulin growth factor in vertebrates. The clustering evolutionary tree demonstrated that AalbILP1, 2, 3, and 5 were most conservative among mosquitoes, followed by AalbILP6, and both AalbILP4 and 7 were unique to Aedes. The results of qRT-PCR showed that AalbILP1, 2, 3, and 5 were expressed at different developmental stages. Compared with other developmental stages, AalbILP5 was the highest expressed in male adult mosquitoes (1.358 3±0.576 9; q_egg=6.572, q_larva=5.771, q_pupa=5.409, q_female=3.115, all P<0.05). AalbILP 3 and 4 were specifically expressed in the head of the whole mosquito, and AalbILP6 was principal expressed in the ovaries of female adult mosquitoes. AalbILP7 is a pseudogene and is not transcribed. Compared to the non-blood mosquitoes, the expression level of AalbILP3 and AalbILP4 in the head of female mosquitoes were significantly upregulated by 2.60 and 1.68 times (t_{AalbILP3- head}=9.596, P_{AalbILP3- head}<0.001; t_{AalbILP4-head}=4.524, P_{AalbILP4-head}=0.001); The expression of AalbILP1 in the head, fat body and midgut was up-regulated by 10.33, 6.07, and 3.79 times（t_head=4.255, P_head=0.001; t_{fat body}=4.305, P_{fat body}=0.001; t_migut=10.480, P_migut<0.001）, but the expression of throax and ovary decreased 4.24 and 2.17 times （t_throax=7.922, P_throax<0.001; t_ovary=3.752, P_ovary=0.003）; The expression of AalbILP6 in midgut and ovary was up-regulated 11.91 and 2.16 times (t_migut=5.799, P_migut<0.001; t_ovary=9.074, P_ovary<0.001). Conclusions Six ILP sequences have been identified in the whole genome of Ae. albopictus and the spatiotemporal expression profiles of ILPs have been constructed.

Objective To investigate the Rickettsia infection and species of pet-dog-parasitizing ticks by the molecular biological technique. Methods Tick samples were collected from the body surfaces of pet dogs in pet centers in Wudang district, Guiyang, China from March to May 2021. Tick species identification and the polymerase chain reaction detection and genotyping of citrate synthase gene (gltA) and outer membrane protein A gene (ompA) of tick-borne Rickettsia were performed. Results A total of 133 ticks were collected from the body surfaces of pet dogs. The morphological characteristics and tick 16S rRNA amplification confirmed that all collected ticks were Rhipicephalus sanguineus. Rickettsia-specific ompA gene amplification, sequencing, and phylogenetic analysis confirmed that R. sanguineus were infected with R. raoultii. Conclusion R. sanguineus is found to parasitize the body surfaces of pet dogs and be infected with R. raoultii, indicating a risk of Rickettsia infecting humans through pet-dog-parasitizing ticks. It is recommended for the local health authorities to make further assessment of the pathogenicity and public health risks of pet-dog-parasitized tick-borne Rickettsia.

Objective To investigate the effects of different diets on the density of Wolbachia in the ovary, fat body, and the other tissues in female Aedes aegypti of the WB strain. Methods After eclosion, Ae. aegypti mosquitoes of the WB strain were divided into glucose group (fed with glucose), white sugar group (fed with white sugar), and blood group (fed with blood for 2 hours on the 3rd day). Mosquitoes in each group were collected on days 4, 4.5, 5, and 6 to harvest the ovary, fat body, and the other tissues. DNA was extracted to determine the density of Wolbachia in all tissues using quantitative real-time PCR. SPSS 20.0 software was used for data analysis. The Kruskal-Wallis H test was used to compare the density of Wolbachia under different diets and at different days of age.Results On day 4.5, Wolbachia densities in the ovary in the glucose, white sugar, and blood groups were 2.149, 2.773, and 0.761, respectively, with a statistically lower density in the blood group (H=40.754, P<0.001); Wolbachia densities in the fat body in the three groups were 0.859, 1.189, 1.298, respectively, and the differences were not statistically significant (H=1.631, P=0.442). Wolbachia densities in the other tissues were 0.505, 0.405, 1.012, respectively, and the differences were not statistically significant (H=6.306, P=0.043). Under different diets but at the same days of age, Wolbachia densities in the ovary were lowest in the blood group on days 4, 5, and 6, and highest in the white sugar group on days 4 and 6 (H=14.335, P=0.001; H=22.049, P<0.001; H=4.266, P=0.084); for the fat body on days 4 and 6 and the other tissues on days 4, 5, and 6, Wolbachia densities were highest in the blood group, followed by the white sugar group, and lowest in the glucose group (H=7.186, P=0.028; H=10.504, P=0.005; H=16.338, P<0.001; H=14.083, P=0.001; H=4.266, P=0.118). At different days of age but under the same diets, Wolbachia densities in the ovary, fat body, and the other tissues in the glucose group had no statistical differences between different days of age (H=4.683, P=0.096; H=2.451, P=0.294; H=0.293, P=0.864); under the white sugar diet, Wolbachia densities in the ovary and fat body were no statistically increased with days of age (H=4.731, P=0.094; H=0.390, P=0.823); under the blood diet, Wolbachia densities in the ovary and fat body were first decreased and then increased with days of age (H=20.572, P<0.001; H=9.675, P=0.008). Conclusion Feeding blood reduces the density of Wolbachia in the reproductive tissue but increases its density in the non-reproductive tissues of Ae. aegypti mosquitoes of the WB strain. Under laboratory conditions, the mosquitoes can be fed with white sugar to increase the density of Wolbachia in the body.