The Horizontal Transmission Of Wolbachia In Hylyphantes Graminicola And Its Impact On Host’s Developmental Stages And Sex Ratio

Symbiotic bacteria are widely distributed in arthropods and can affect the host’s reproduction,growth and development.Vertical transmission is the basic mode of endosymbiotic bacteria in the host species.A large number of studies have confirmed the existence of horizontal transmission of endosymbiotic bacteria between different hosts.Hylyphantes graminicola is an important natural enemy of agricultural pests,and it is small,widely distributed in the field.In order to combine the effects of symbiotic bacteria on the reproduction,growth of the host with the use of the spider for the biological control of the pest.In this study,the dominant symbiotic bacteria were detected in Hylyphantes graminicola,and the effects of Wolbachia infection on host’s growth,development and the horizontal transmission of Wolbachia within same host species were also studied.The main results of this paper are as follows:1.The detection of dominant symbiotic bacteria in the Hylyphantes graminicolaA total of 50 adult spiders were randomly selected.DNA was extracted for each individual,then 10 DNA samples were mixed into 1 mixture DNA with equal concentration.5mixed DNA samples were used for high-throughput sequencing of bacteria 16S rRNA genes about the V3 and V4 region.The result of bioinformatics analysis showed that Wolbachia was the dominant symbiotic bacterium in the spider,accounting for 50%of the total symbiotic bacteria group,followed by Cardinium（34%）,and the symbiotic bacteria of Rickettsia accounted for about 4%of the total symbiotic bacteria group.A total of 167 spiders were collected for PCR infection detection.The results showed that the infection rate of Cardinium was 100%,and the infection rate of Wolbachia was 38.9%.2.The effects of Wolbachia on the developmental period and sex-ratio in the Hylyphantes graminicolaThe development periods of 1329 individuals for F₁ generation spider were recorded from 34 Wolbachia uninfected（W^-）female spiders,and 27 Wolbachia infected（W⁺）female spiders.For the female spiders,the second instar,the third instar,the fourth instar and the total duration of W⁺individuals were very significantly lower than those of W^-individuals（P<0.01）,and the first instar of W⁺individuals was significantly lower than that of W^-individuals（P<0.05）.There was no significant difference in the egg stage and fifth instar between W⁺female individuals and W^-female individuals（P>0.05）.For the male spiders,the first instar and the total duration of W⁺individuals were very significantly shorter than those of W^-individuals（P<0.01）,and the second instar,the third instar and the fifth instar of W⁺individuals were significantly shorter than those of W^-individuals（P<0.05）.The egg stage and the fourth instar had no significant difference between W⁺male individuals and W^-male individuals（P>0.05）.A total of 1490 individuals for F₁ generation adult spiders were selected from 30 W^-mother spiders and 34 W⁺mother spiders（not less than 20 adult offspring for each mother spider）,and used to analysis of the effects of symbiotic bacteria infection on the offspring sex-ratio.The results showed that the female-male ratio（_M±SE）of F₁ generation for Wolbachia uninfected spiders was 1.338±0.044（n=789）,and the female-male ratio（_M±SE）of Wolbachia infected spiders was 1.610±0.081（n=701）.The female-male ratio of offspring from Wolbachia infected spiders was very significantly higher than that of offspring from Wolbachia infected spiders（P<0.01）.3.The horizontal transmission of Wolbachia within the host speciesTwo mating group♀⁺×♂^-and♀^-×♂⁺（♀⁺:the female spider infected with Wolbachia;♀^-:non-Wolbachia female spider;♂⁺:the male spider infected with Wolbachia;♂^-:non-Wolbachia male spider）from adult F₁ generation were set up.The transmission situation was detected by fluorescence quantitative PCR after mating at 0h,1d,2d,3d,4d,5d,6d,7d,15d and 30d.The results showed that Wolbachia was detected in Wolbachia uninfected female spider at 1d,3d,5d,6d,7d,15d and 30d after successful mating;Wolbachia was detected in Wolbachia uninfected male spider at 3d,4d,6d and 15d after successful mating.After the mating transmission,the number of copies that Wolbachia target fragment did not have a clear linear relationship over time.The results of this study show that Wolbachia can be transmited through mating behavior.