Objective To establish a duplex droplet digital polymerase chain reaction (ddPCR) detection method for Japanese encephalitis virus (JEV) and West Nile virus (WNV). Methods Based on the designed primers and probes of JEV and WNV, a duplex ddPCR detection system for JEV and WNV was established. Its sensitivity, specificity, and repeatability were explored. The sensitivity was compared with the number of cycles required for the fluorescent signal to cross the threshold in each reaction tube of dual quantitative PCR. Results The detection sensitivity of the duplex ddPCR detection system could reach 10 ² copies/μl for both JEV and WNV, with good specificity and repeatability. No cross-reactivity was observed with the Dengue virus, Chikungunya virus, Zika virus, Tick-borne encephalitis virus, and human genome. Conclusion The established duplex ddPCR method shows high sensitivity and specificity for JEV and WNV detection, which provides a solution for detection for the two viruses in different scenarios.

Objective To analyze the genetic diversity of nine batches of Lasiopodomys brandtii rodents from three sample plots (New Barag Right banner, East Ujimqin banner, and Xilinhot city) of the Inner Mongolia Autonomous Region (Inner Mongolia), China by using 16 pairs of microsatellite primers. Methods A total of 275 L. brandtii rodents were trapped using snaps or cages in May, July, and September of 2021. DNA was extracted from them with magnetic beads, followed by PCR amplification with 16 pairs of fluorescent-labeled microsatellite primers. The microsatellite data were obtained using GeneMarker. The genetic diversity and structure of L. brandtii populations were analyzed using GenAIEx 6.5, Arlequin (version 30), and structure 2.3.4. Results A total of 472 alleles were detected by using 16 pairs of microsatellite primers. The average polymorphic information content was 0.821 3. The average Shannon’s information index was 1.812. Observed heterozygosity ranged from 0.478 to 0.951. Expected heterozygosity ranged from 0.575 to 0.891. According to cluster analysis and principal component analysis, the nine L. brandtii populations could be divided into four groups by month. The genetic diversity index of the four groups was May > July > September. The analysis of molecular variance showed that the variation among individuals (88.51%) was much greater than that among the groups (5.04%) and that among the populations (6.45%). The mantel test showed no significant correlation between genetic distance and geographical distance of L. brandtii ( R ²=0.001 6). Conclusions The L. brandtii populations sampled in Inner Mongolia had high genetic diversity. The genetic variation of L. brandtii populations was mainly observed among individuals. Seasonal variation could be the main factor affecting the genetic structure of L. brandtii populations.

Objective To investigate the density and seasonal fluctuations of vector mosquitoes in Hangzhou, Zhejiang province, China from 2017 to 2021, and to provide a scientific basis for vector control and mosquito-borne disease prevention. Methods The light trap method was used to investigate 15 districts/counties/cities in Hangzhou from April to November, 2017-2021, in accordance with the "National Vector Surveillance Program" and "Zhejiang Province Vector Surveillance Program". Excel 2016 software was used to summarize the monitoring data, and SPSS 21.0 software was used for statistical analysis. The Chi-square test was used for the difference of mosquito composition ratio in different years, and one-way ANOVA was used for the difference of mosquito density in different habitats. Results From 2017 to 2021, the mosquito density in Hangzhou was 1.99 mosquitoes/light·night, and Culex pipiens pallens/quinquefasciatus (71.90%) was the dominant species. The seasonal fluctuation trend of mosquito density generally showed a bimodal curve, peaking in July and October. There were statistical differences in densities of Cx. pipiens pallens/quinquefasciatus, Cx. tritaeniorhynchus, Aedes albopictus, Anopheles sinensis, and Armigeres subalbatus between different years (all P<0.001). In terms of various habitats, livestock barns/farms had the highest mean mosquito density (4.92 mosquitoes/light·night), and hospitals had the lowest (1.35 mosquitoes/light·night). There were statistical differences in densities of An. sinensis and Ar. subalbatus between various habitats ( F=3.581, P=0.023; F=5.431, P=0.004). Cx. pipiens pallens/ quinquefasciatus was the dominant mosquito species in various habitats, followed by Cx. tritaeniorhynchus, Ar. subalbatus, Ae. albopictus, and An. sinensis. Livestock barns/farms had the highest mosquito densities in terms of any of the above five species. Conclusions The mosquito density in Hangzhou is generally at a low level. Cx. pipiens pallens/ quinquefasciatus is the dominant species. In future mosquito control, livestock barns/farms should be highlighted for special attention. Local districts/cities/counties should take targeted mosquito control measures based on their density changes, distribution, and seasonal fluctuations.

Objective: To investigate the density distribution of Aedes mosquitoes in different habitats and seasonal fluctuation in Hangzhou, Zhejiang Province, China from 2017 to 2021, so as to provide a scientific basis for effective prevention and control, risk assessment, and early warning of dengue fever. Methods: According to the National Vector Surveillance Implementation Plan and the Zhejiang Provincial Vector Surveillance Plan, Aedes mosquitoes were monitored in 15 districts (counties and cities) in Hangzhou. Adult mosquitoes were monitored using the double-layered mosquito net method, and larvae were monitored using the Breteau index (BI) method. The surveillance time was set during April to November. The mosquito surveillance data of the 15 districts (counties and cities) were collected. Excel 2019 software was used for data analysis. The net trap index, BI, and container index (CI) were calculated. Categorical data were compared using the Chi-squared test. One-way analysis of variance was used for quantitative data. Results: From 2017 to 2021, the density of Ae. albopictus in Hangzhou by the double-layered mosquito net method was 2.54 mosquitoes/net·h, and no Ae. aegypti mosquitoes were captured. The density of adult mosquitoes was lowest in 2017 and highest in 2020. There was a statistical difference in the net trap index between different years (F=5.117, P=0.017). The seasonal distribution generally presented a bimodal pattern, with the peaks of mosquito densities mainly in July and October. The average BI from 2017 to 2021 was 9.18, and the peak period of the BI was during May to October. The CI showed that Ae. albopictus larvae were distributed in all kinds of water bodies. The CI differed statistically between different water bodies in the same year (all P < 0.001) and between different years in the same water body (all P < 0.001). Conclusions: Ae. albopictus density was high in Hangzhou, indicating a risk of dengue fever outbreaks and regional epidemics. It is recommended that the counties (cities, districts) take timely mosquito control measures according to the density, distribution, and seasonal fluctuation of mosquitoes.

Objective To investigate the level, trend, and epidemiological characteristics of scrub typhus in Changzhou, China, and to provide a basis for prevention and control of the disease. Methods The reported data of scrub typhus in Changzhou from January 1, 2012 to December 31, 2021 were collected from the China Information System for Disease Control and Prevention. We analyzed the incidence of the disease and described the regional, temporal, and population distribution characteristics of scrub typhus using descriptive epidemiology method. Results From 2012 to 2021, a total of 149 cases were reported in Changzhou, with a median annual reported incidence rate of 0.28/100 000, with the highest reported incidence rate of 0.74/100 000 in 2015 and the lowest rate of 0.04/100 000 in 2012. Cases were reported in all districts of Changzhou, with the top three being Wujin district (60, 40.27%), Xinbei district (34, 22.82%), and Liyang city (20, 13.42%). Most cases were reported between October and November in farmers, household workers, and unemployed persons, and in persons aged 45-69 years. Conclusions Scrub typhus surveillance should be strengthened, medical personnel should be trained to improve the diagnosis of scrub typhus, and health education for high-risk populations should be carried out, in order to prevent and control the occurrence and prevalence of scrub typhus.

Mosquito-borne viruses are transmitted mainly by mosquitoes during blood feeding. These viruses include more than 100 species and can cause many infectious diseases and seriously threaten human health. The accurate and rapid detection of mosquito-borne viruses in field is an important part of preventing and controlling mosquito-borne diseases. Field detection allows earlier disease prevention and control and helps control the outbreaks and epidemics of mosquito-borne diseases. However, current detection methods mainly target clinical samples and cannot be used for on-site detection of viruses in mosquito samples. This paper reviews the detection methods for mosquito-borne viruses, including the classical isolation and culture, antigen detection, and nucleic acid detection methods, as well as the new technologies developed rapidly in recent years, such as isothermal amplification and clustered regularly interspaced short palindromic repeats-associatied (CRISPR-Cas). The characteristics of these methods are analyzed, and the directions of developing technologies for monitoring mosquito-borne viruses in field are discussed.

Objective To investigate the species composition, population density, and seasonal fluctuation of mosquitoes in human settlements and surrounding environments in Guizhou province, China, and to provide a basis for risk assessment of mosquito-borne diseases and scientific and effective vector control. Methods Mosquito surveillance data from 78 provincial vector surveillance sites in Guizhou province from 2017 to 2021 were collected to analyze mosquito species composition and compare mosquito population density and seasonal fluctuations in different years, regions, and habitats. SPSS 24.0 software was used to perform analysis of variance on the Breteau index (BI) and density of mosquitoes in different habitats for risk assessment. Results A total of 131 476 female mosquitoes were captured from 2017 to 2021, with a density of 5.82 mosquitoes/light·night. Culex pipiens quinquefasciatus accounted for the highest proportion with 43.34% of the total catches, followed by Anopheles sinensis with 20.68%. Cx. pipiens quinquefasciatus was the dominant mosquito species in most cities/prefectures. Zunyi had the highest composition ratio of Cx. pipiens quinquefasciatus with a total of 17 409 catches (61.32%). There was a statistical difference in mosquito density in different areas ( F=5.276, P<0.001). From 2017 to 2021, the highest densities of mosquitoes were all in livestock sheds, and Cx. pipiens quinquefasciatus was the dominant mosquito species in all habitats. The density of mosquitoes was statistically different in different habitats ( F=114.368, P<0.001). Adult mosquito density by light trapping increased gradually from May, peaked during June to August, and decreased rapidly from September. BI showed two obvious peaks in June and August, respectively. The BI began to rise rapidly in May, and gradually declined in September and October. The BI was greater than 5 for each year. The areas of level-1 risk included Anshun, Guiyang, and Qiandongnan Miao and Dong Autonomous Prefecture. The BI values in Anshun in 2019 and Guiyang in 2021 were both greater than 10, indicating a level-2 risk. Conclusions Cx. pipiens quinquefasciatus and An. sinensis are the dominant mosquito species in human settlements and surrounding environments in Guizhou province. The mosquito density is high in rural habitats, especially in livestock sheds, which are major mosquito breeding sites, and mosquito control should be strengthened in such environments. The peaks of mosquito activity in Guizhou province are in June and August, which may by closely related to the unique climate and rainfall conditions of Guizhou province. Mosquito control should be done before the peak breeding season of mosquitoes to reduce the density of mosquitoes and the risk of mosquito-borne virus transmission.

Objective: To select chemical substances with attractive effects on Aedes albopictus for on-field assessment, and to explore the preference of Ae. albopictus to six human odors. Methods: Under laboratory conditions, six chemical substances (1-octen-3-ol, lactic acid, myristic acid, ammonia, acetone, and ethyl acetate) were separately tested for their individual attracting effects on female Ae. albopictus at concentrations of 0.1, 1.0, and 10.0 mg/ml. The six substances were mixed in pairs (1∶1) at certain concentrations, and then the pairs were tested for attracting effects on female Ae. albopictus. The pair with good attractive effects was combined with lactic acid for testing. The attractant combination selected by laboratory experiments was tested for attractive action for Ae. albopictus in the field. The attracting effects of the substances or combinations on Ae. albopictus were compared using the t test and one-way analysis of variance. Results: Ae. albopictus mosquitoes were attracted to 1-octen-3-ol at 0.1, 1.0, and 10.0 mg/ml, myristic acid at 1.0 and 10.0 mg/ml, and ethyl acetate at 0.1 mg/ml, but not to the other three substances at any concentration. The pairs of 1-octen-3-ol+myristic acid, lactic acid+myristic acid, and lactic acid+1-octen-3-ol statistically attracted Ae. albopictus compared with the control group (t=8.102, P=0.001; t=4.696, P=0.009; t=5.127, P=0.007). The combination of lactic acid+1-octen-3-ol+myristic acid statistically attracted Ae. albopictus compared with the control group (F=86.841, P < 0.001). Field testing showed significant differences in the attracting effect between lactic acid+1-octen-3-ol+myristic acid and the control (day 1: t=7.462, P=0.014; day 2: t=20.500, P < 0.001; day3: t=9.383, P=0.001). Conclusion: The combination of lactic acid, 1-octen-3-ol, and myristic acid has attractive action for Ae. albopictus, which should be further researched in the field.

Objective To investigate the genetic diversity, genetic differentiation, and phylogenetic relationship of Culex pipiens pallens populations in Shangdong, China. Methods Cx. pipiens pallens populations were collected in Heze, Dezhou, Liaocheng, Qingdao, Yantai, and Rizhao, Shandong province from September 2020 to September 2022. Genomic DNA was extracted from a single female adult mosquito. The mitochondrial cytochrome c oxidase subunit Ⅰ (mtDNA-COⅠ) gene sequences were amplified by polymerase chain reaction and sequenced afterwards. The acquired sequences were compared with those in the Basic Local Alignment Search Tool on the GenBank. BioEdit 7.0 was used to compare and analyze the sequencing results. DnaSP v6 was used to analyze the genetic diversity of populations. Arlequin 3.5 was used to calculate the genetic differentiation coefficient and gene flow of populations. PopART 1.7 was used to construct the haplotype network (TCS network). Results A total of 423 sequences of 603 bp were obtained from 6 Cx. pipiens pallens populations in Shandong province. The average content of A+T bases was 69.2%, which was consistent with the AT bias of mitochondrial DNA. There were eight haplotypes in the haplotype analysis, of which haplotype H01 was the dominant haplotype. The results of mtDNA-COⅠ sequence analysis showed that the population had rich genetic diversity. The results of molecular variance analysis indicated that the genetic differentiation of Cx. pipiens pallens mainly came from within the population; there was some genetic differentiation among some populations. The results of neutrality test showed that the populations of Cx. pipiens pallens in Liaocheng and Heze experienced expansion recently. Conclusions The mtDNA-COⅠ gene can be used as a molecular marker to study the genetic diversity of Cx. pipiens pallens populations. The genetic development of Qingdao population is special compared with other geographical populations.

Objective To establish the susceptibility baseline of Musca domestica to pyrethroid insecticides, and to analyze the knockdown resistance gene polymorphism in some wild populations of M. domestica in Henan province, China. Methods The susceptible strain of M. domestica was a laboratory population that had not been exposed to any insecticides. The wild populations were collected in Kaifeng, Anyang, Nanyang, Zhumadian, and Hebi cities in 2019-2021. Topical application method was used to determine the median lethal dose (LD ₅₀) of permethrin, deltamethrin, and beta-cypermethrin in female adult flies. Allele-specific polymerase chain reaction (AS-PCR) was used to determine the genotypes and mutation rates of knockdown resistance genes. Spearman correlation analysis was used to test the correlation between mutation rate and LD ₅₀. DNA sequencing was used to verified the results of AS-PCR. Results The LD ₅₀ of permethrin, deltamethrin, and beta-cypermethrin was 0.151 0, 0.077 2, and 0.166 6 μg/♀ in the susceptible strain, 0.321 6, 0.130 6, and 2.235 4 μg/♀ in Kaifeng population, 0.867 7, 0.459 2, and 1.591 6 μg/♀ in Anyang population, 5.173 7, 1.037 2, and 0.416 1 μg/♀ in Nanyang population, 0.634 1, 0.108 2, and 0.262 4 μg/♀ in Zhumadian population, and 2.745 0, 1.102 9, and 2.556 0 μg/♀ in Hebi population, respectively. Only one genotype, i.e. sensitive homozygote, was detected in the susceptible strain. Five genotypes were detected in the wild populations, including sensitive homozygote (L/L), sensitive/1014F heterozygote (L/F), sensitive/1014H heterozygote (L/H), 1014F/1014H heterozygote (F/H), and 1014H homozygote (H/H). Mutation rates in the wild populations were 14.84%, 17.97%, 13.28%, 10.16%, and 20.31%, respectively. The correlation test showed that the resistance to beta-cypermethrin was related to L1014H mutation. Five genotypes from 130 M. domestica samples were sequenced, and the accuracy of AS-PCR results was 79.23%. Conclusions The susceptibility baseline of M. domestica to pyrethroid insecticides is established in Henan, and this baseline can be used as a reference value for insecticide resistance monitoring. AS-PCR can be used for routine monitoring of resistance genes in M. domestica.

Objective To determine the way and level of understanding dengue fever knowledge and the current situation of mosquito control behavior of residents in border areas of Yunnan province,China,to explore the publicity channel factors that affected residents' level of dengue fever knowledge and mosquito control behavior,and to put forward suggestions to improve residents' level of dengue fever knowledge and mosquito control behavior,so as to prevent the spread of dengue fever in China. Methods An offline questionnaire survey was conducted among Chinese residents aged ≥11 years living in Simao district,Ruili,and Jinghong cities. EpiData 3.1 software was used to input the questionnaire results,and SPSS 19.0 software was used for statistical analysis. Results A total of 778 questionnaires were obtained in this offline survey,and 721 respondents were aware of dengue fever,with an awareness rate of 92.67%. There were significant differences in the awareness of dengue fever among residents in Simao district,Ruili,and Jinghong cities ( χ ²=14.807, P=0.001). A further analysis of the questionnaire revealing the awareness of dengue fever showed that there were some differences in the way and level of understanding dengue fever knowledge and mosquito control behavior among residents in Simao district,Ruili,and Jinghong cities ( P<0.05). The basic personal information and publicity channel factors influencing the level of dengue fever knowledge and mosquito control behavior among residents in the three border areas were different. Conclusions The influencing factors of media publicity channels on the level of dengue fever knowledge and mosquito control behavior among residents in Simao district, Ruili, and Jinghong cities are slightly different. According to the differences in different regions,different publicity channels should be adopted to publicize and educate residents.

Objective To investigate the population distribution, virus-carrying status, and virus gene characteristics of host animals for Hantavirus in some areas of Zhoushan Islands, Zhejiang province, China, and to provide evidence for prevention and control of hemorrhagic fever with renal syndrome (HFRS). Methods Small mammals were captured using the night trapping method in Dinghai and Daishan areas of Zhoushan Islands, followed by species identification. The rodent density and species composition were analyzed using the Chi-square test. The small mammal’ lungs were collected to detect the nucleic acid of Hantavirus by quantitative real-time PCR. The M gene was amplified by nested PCR and sequenced for genotyping and homology analysis. Results In Dinghai and Daishan areas of Zhoushan Islands in 2021, the average rodent density was 3.42%, with 1.71% in the wild and 6.16% in residential areas. Six species of small mammals were captured. The dominant species was Apodemus agrarius in the wild and Rattus norvegicus in residential areas. The density of rodents was statistically different between Daishan Island, Yushan Island, and Dinghai Island ( χ ²=108.468, P<0.001), between the wild and residential areas ( χ ²=68.935, P<0.001), and between different species ( χ ²=37.749, P<0.001). Hantavirus RNA was detected in 13 small mammals lung specimens, with a positive rate of 6.70%. The M fragments of three Hantavirus RNA extracts were successfully amplified and sequenced. The homology of the three M fragments was 96.90%-97.10%, belonging to Seoul virus (SEOV, S3 subtype). Conclusions There are HFRS foci with R. norvegicus as the source of infection in Zhoushan Islands, where SEOV S3 subtype is the epidemic strain. Environmental changes may alter natural foci.

Objective To analyze the epidemic situation of hemorrhagic fever with renal syndrome (HFRS) and the populations distribution and Hantavirus infection status of the host animals in Zhejiang province, China in 2021, and to provide scientific evidence for establishing preventive measures. Methods Descriptive epidemiological methods were used to analyze HFRS case data and host surveillance data, which were online reported in the Communicable Disease Surveillance Information Reporting and Management System of Zhejiang province in 2021. The Chi-square test was used for rate comparison. Results A total of 180 HFRS cases were reported in Zhejiang province in 2021, with a mean annual incidence rate of 0.29/100 000. Most cases were observed in Taizhou, Ningbo, Shaoxing, and Quzhou. The cases distributed throughout the year, peaking during May to June in summer and December in winter at similar levels. The high-incidence group aged over 40 years old, accounting for 76.67% (138/180) of the total cases. The main occupation was farmers, accounting for 61.67%. A total of 11 921 effective traps were set in the five surveillance sites in Zhejiang province, capturing 660 small mammals, with an average capture rate of 5.54%. A total of 660 serum samples of small mammal were detected, among which 35 (5.30%) were antibody-positive. Hantavirus was positive in 15 of 660 (2.27%) small mammal lung samples. There were statistical differences in capture rate ( χ ²=16.161, P<0.05), the positive rate of HFRS antibody ( χ ²=14.957, P=0.005), and the virus infection rate of small mammals ( χ ²=8.413, P=0.048) among the five surveillance sites. Conclusions HFRS was sporadic at a low level and distributed in extensive areas in Zhejiang province in 2021. The surveillance sites had generally high small mammal densities and fluctuating virus infection rates in small mammals, requiring scientific rodent control-based comprehensive prevention and control measures.

Objective To investigate the microbiota of the dominant tick species Rhipicephalus microplus in the minority autonomous prefectures of Guizhou province, China, and to provide a scientific basis for the prevention and control of tick-borne disease. Methods Rh. microplus ticks were collected from the body surface of cattle and sheep at 18 sampling sites located in Qiandongnan Miao and Dong Autonomous Prefecture, Qiannan Buyi and Miao Autonomous Prefecture, and Qianxinan Buyi and Miao Autonomous Prefecture in April and July of 2019 and 2020. The ticks were grouped by region, growth stage, and sex, with three parallel samples in each group, followed by metagenomic sequencing. The sequencing results were processed with quality control and assembly procedures before homology alignment with the non-redundant protein database of National Center for Biotechnology Information to obtain taxonomic annotation information. R (v. 3.6.3) and GraPhlAn (v. 1.1.3) were used for visual analysis, including species composition analysis, non-metric multidimensional scaling (NMDS) analysis, and analysis of similarities (ANOSIM). Results The dominant bacterial phylum for Rh. microplus was Firmicutes (60.70%), followed by Proteobacteria (33.76%) and Actinobacteria (4.53%). Staphylococcus aureus (38.29%) was the dominant bacterial species, followed by Klebsiella pneumoniae (7.79%) and Streptococcus pneumoniae (5.26%). Several tick-borne pathogens were detected, including various genotypes of spotted fever group Rickettsia and Anaplasma phagocytophilum (1.38%). R. fournieri (2.91%) was an emerging rickettsiae detected from ticks for the first time in China, and R. japonica (1.76%) and R. monacensis (0.04%) were common tick-borne Rickettsia genotypes in China. R. fournieri and R. japonica were mainly carried by male ticks. The NMDS analysis showed that the microbiota composition of Rh. microplus differed by growth stage and sex. The ANOSIM analysis indicated reasonable grouping, with greater intergroup differences than intragroup differences ( R=0.147, P=0.014). Conclusions In the minority autonomous prefectures of Guizhou province, Rh. microplus ticks have a diverse microbiota composition, carrying pathogens that may cause spotted fever and human granulocytic anaplasmosis. Therefore, vector surveillance and pathogen detection should be strengthened, and effective control measures should be taken to reduce the risk of tick-borne diseases.

Objective To investigate the population changes of plague hosts and vectors, monitor plague epizootics, and assess the risk of plague in Heqing county, Yunnan province, China from 2011 to 2020, and to provide a scientific basis for plague prevention and control. Methods Animal plague surveillance was carried out according to the National Plague Surveillance Program and the Yunnan Provincial Plague Surveillance Program. The surveillance data on plague hosts, vectors, etiology, and immunology in Heqing county from 2011 to 2020 were collected and analyzed. Results A total of 4 146 small mammals were captured in animal plague surveillance in Heqing county from 2011 to 2020, belonging to 10 species, 7 genera, 4 families, and 2 orders. In residential areas, Rattus tanezumi (82.89%) and R. norvegicus (10.10%) were the dominant species, while in farming areas, Eothenomys miletus (71.65%) and Apodemus chevrieri (19.67%) were the dominant species. The average small mammals densities in residential and farming areas were 0.66% and 3.89%, respectively, which were statistically different ( χ ²=1 770.463, P<0.001). A total of 3 229 ectoparasitic fleas were collected from the small mammals, belonging to 7 species, 7 genera, and 4 families. The flea infestation rate was 36.78 %, and the flea index was 0.78. The dominant species were Ctenophthalmus quadratus (85.81%) and Neopsylla specialis specialis (11.12%). Xenopsylla cheopis was not found. A total of 4 078 small animals and 1 587 groups of flea were examined for Yersinia pestis isolation and culture, 1 712 samples were examined for plague F1 antibody with indirect hemagglutination assay, and 354 samples were examined for plague F1 antigen with reverse indirect hemagglutination assay. In 2017, 7 and 3 strains of Y. pestis were detected in small mammals’ organs and fleas, respectively; and 3 samples of living rodents were positive for plague F1 antigen. Conclusions In the plague foci of Heqing county, the dominant species of plague hosts and vectors were prominent, with no obvious change in species composition and quantity. The increase in the density of main hosts was closely related to the occurrence and epidemic of animal plague, with a high risk of plague epizootics in recent years. It is necessary to expand plague surveillance area, improve the monitoring quality, strengthen the early warning and risk assessment, and implement comprehensive joint prevention and control measures.

Objective: To analyze the species, density, distribution, and variation of flies in Shanghai, China, to carry out risk assessment, prediction, and early warning of fly-borne diseases in a timely manner, so as to provide scientific reference for fly control. Methods: Using the cage trap method, fly traps were placed in farmers' markets, residential areas, external environment of restaurants, and large green belts. Fly surveillance data were collected every ten days from March to November in 2016-2021 in Shanghai. The data were processed and analyzed using Excel 2019 and SPSS 20.0. Results: From 2016 to 2021, 18 052 flies were caught with an average density of 1.52 flies/cage. The identified fly species belonged to 5 families, 9 genera, and 14 species. The dominant species were Muscina stabulans, Boettcherisca peregrina, Chrysomya megacephala, Lucilia sericata, and Musca domestica. M. stabulans showed the highest density of 0.28 flies/cage, followed by B. peregrina (0.24 flies/cage). In large green belts, the density of B. peregrina was higher than that of M. stabulans, and in all other habitats, the density of M. stabulans was the highest. There was a significant difference in density between B. peregrina and M. stabulans in the farmers' market (t=-2.674, P=0.023). The proportion of Mu. domestica decreased year by year, and the proportion of M. stabulans increased. The fly density was the highest in 2018 (1.82 flies/cage) and the lowest in 2016 (1.04 flies/cage). Fly density peaked from June to August, with the monthly average densities of 2.65, 2.49, and 2.28 flies/cage, respectively. In 2016-2021, the fly density in different habitats was in the order of farmers' markets > large green belts > residential areas > external environment of restaurants. Conclusions: The fly species are diverse in Shanghai. M. stabulans has the highest density. Fly density peaks from June to August, and farmer's market is a key place for targeted fly control. Fly density rose again after decline in 2019, suggesting that comprehensive control of flies should be continuously strengthened.

Objective To investigate the species, distribution, density, and seasonal fluctuation of sandflies in different areas of Shanxi province, China, and to provide a scientific basis for the risk assessment and control of kala-azar. Methods From May to September in 2021, sandflies were captured with light trapping in different habitats in 14 counties/cities/districts of 11 prefectures of Shanxi province. The non-bloodsucking female sandflies were randomly selected from different areas and identified by morphological and molecular biological methods. Excel 2010 software was used to construct the database, followed by analysis of sandfly catches by habitat or month. The distribution of sandflies was mapped using ArcGIS 10.7. Results A total of 19 006 sandflies were captured, with an average density of 40.87 sandflies/light·night. The seasonal fluctuation of sandflies throughout the year showed a single-peak distribution. Sandflies were first caught in the first half of May, and the density peaked during the second half of June to the first half of July, after which the density decreased rapidly. Sandflies were captured in six environmental types: farmyards, sheep pens, cattle sheds, chicken pens, pig pens, and mule/horse sheds. Sheep pens had the highest density of 121.39 sandflies/light·night, and farmyards had the lowest density of 9.63 sandflies/light·night. Of the 432 sandflies identified by morphology, one was Sergentomyia squamirostris, and the rest were Phlebotomus chinensis. Another 110 sandflies were randomly selected for molecular biological identification: four were S. squamirostris, and the remaining 106 were P. chinensis. P. chinensis was the absolute dominant species. Conclusions Sandflies distributed in all the 14 counties/cities/districts of the 11 cities of Shanxi province. P. chinensis was the dominant species of sandflies in Shanxi province, which was the main vector for kala-azar transmission. They were mainly distributed in rural areas, especially in livestock/poultry-raising places, where environmental management and sandfly control should be strengthened to reduce the risk of kala-azar transmission.

Objective To investigate the species composition and spatial distribution of mosquitoes in the human habitats of the Yellow River basin in Qinghai province, China. Methods Ten counties/cities/districts in the Yellow River basin in Qinghai province were set as the study space. From July to September of 2018 to 2021, blood-sucking mosquitoes active at night were collected using mosquito trap lamps (Gongfu Xiaoshuai) around rural livestock pens and residential houses, followed by identification and classification. Excel 2016 was used to establish the database to calculate population dominance and niche breadth. SPSS 22.0 was used to perform hierarchical cluster analysis on the correlation coefficients between mosquito species. Results A total of 2 684 mosquitoes belonging to 12 species of 4 genera were captured,among which Aedes vexans was the dominant species with species abundance of 30.030. The species abundance of Aedes vexans, Ae. falvidorsalis, Culex pipiens pallens, and Cx. modestus is more than 10.000; The ecological breadth of Ae. detrituss, Ae. dorsalis, Ae. vexans, and Cx. pipiens pallens is more than 6; Geographic niche width of Ae. dorsalis, Cx. modestus, Anopheles sinensis, and Ae. flavescens is greater than or equal to 1.000. There were niche overlaps among 90% of mosquito species, with 13 pairs >0.50 (including one pair with a complete overlap). Ae. sergievi was found in Guide county for the first time. Conclusions Ae. vexans, Ae. dorsalis, and Cx. pipiens pallens have relatively high numbers of species, species abundance, ecological amplitude, niche breadth, and niche overlap in human habitats in the Yellow River basin of Qinghai province. These mosquito species are widely distributed and highly competitive in the plateau. It is the first time to discover Ae. sergievi, a new record in Qinghai province, China.

Objective To investigate the knockdown resistance gene mutations of Aedes albopictus in Yuzhou, Henan province, China, and to provide a basis for the control of Ae. albopictus. Methods The double-layered mosquito net method was used to capture mosquitoes in Yuzhou in mid-September of 2020 and 2022. Mosquito species was identified based on morphological characteristics. The DNA of Ae. albopictus mosquitoes was extracted for knockdown resistance gene detection by sequencing. Results A total of 80 Ae. albopictus mosquitoes (40 in each year) were captured. The sequencing results showed that three knockdown resistance gene loci were found mutant in Ae. albopictus. The 1016 locus mutated from valine (Val, V) encoded by the codon GTA to glycine (Gly, G) encoded by GGA. The 1532 locus mutated from ATC-encoded isoleucine (Ile, I) to ACC-encoded threonine (Thr, T). Mutations at 1534 were most frequent, from TTC-encoded phenylalanine (Phe, F) to TCC-encoded serine (Ser, S) or Leu (Leu, L) encoded by TTA/CTC. There were 10 genotypes in total for the three mutant sites. For all the three mutant sites, wild-type homozygotes were predominant, followed by wild/mutant heterozygotes and then mutant homozygotes. There were a total of 11 combination genotypes: the frequency of wild/mutant heterozygous combination genotypes at single loci was 55.00%, which was highest; the frequency of wild homozygous combination genotypes at three loci was 28.75%; the frequency of wild/mutant heterozygous genotypes at two loci was 8.75%; and the frequency of mutant homozygous genotypes at single loci was 7.50%, which was lowest. Conclusions It was the first time to carry out knockdown resistance gene mutation detection in Ae. albopictus in Yuzhou, discovering that the mutations were frequent and complex, which requires further research in this area.