Role and trans infection efficacy of the Wolbachia in the leafhopper vectors of sugarcane white leaf disease

Wolbachia-based strategies are currently accepted as a novel alternative method for controlling vector-borne diseases, and has been developed to control insect pests of agricultural economic crops. Wolbachia prevalence was detected in the Yamatotettix flavovittatus (Matsumura), one of the leafhopper vectors of phytoplasma that causes sugarcane white leaf disease (SCWL). The main purpose of this study was to develop Wolbachia-based strategies for control the leafhopper vectors of SCWL disease. The important steps involved in this approach include describing the exactly role of Wolbachia or related mechanisms that affect the host. Additionally, the microinjection technique is also the important method to delivery Wolbachia to create novel Wolbachia-positive host that do not occur in nature. In terms of Wolbachia prevalence on biological traits and cytoplasmic incompatibility (CI), the results revealed that Wolbachia-positive leafhoppers had reduced the developmental period of immature stages and prolonged the female longevity. In the cross experiments, the results revealed that Wolbachia exhibited significant CI in crosses between males and females with different Wolbachia statuses, resulting in 83.34% unhatched eggs, while when both parents had the same Wolbachia status, the unhatched egg rate was 13.15%. The localization of Wolbachia was widely distributed in the gut, which may be related to nutritional processing, and the gene annotation was found the bfmBAB and pabA genes, which involved in vitamin synthesis. Furthermore, Wolbachia was localized in the testis, confirming CI induction by Wolbachia-positive males. Gene annotation revealed that the histone H2A genes, which involved in sperm cell division.Regarding the effect of Wolbachia prevalence on SCWL acquisition and retention in the leafhopper Y. flavovittatus, a significant difference in SCWL phytoplasma infection frequency was observed during the acquisition (6 and 12 h). The SCWL phytoplasma titers were reduced in Wolbachia-positive leafhoppers from all acquisition time points (<7.00 x 104 copies), whereas it increased for Wolbachia-negative leafhoppers (ranged from 5.37 to 26.86 x 104 copies). During the retention, the proportion of SCWL phytoplasma infection ranged from 47.06-100% and 83.33-100% for Wolbachia-positive and negative leafhoppers, respectively. The Wolbachia-positive leafhoppers retained a low SCWL phytoplasma titer (<5.00 x 104 copies) throughout the retention. By contrast, Wolbachia-negative leafhoppers exhibited increased SCWL phytoplasma levels, ranging from 6.34 to 30.38 x 104 copies. The co-occurrence of Wolbachia and SCWL phytoplasma was observed in the salivary glands, gut, and ovaries, suggesting potential competition for space and resources within these tissues. Additionally, gene annotation identified the def gene, which involved in the production of antimicrobial peptides that are expected to inhibit the abundance of SCWL phytoplasma.The results of microinjection technique development for the leafhopper Matsumuratettix hiroglyphicus (Matsumura) found that the PBS buffer is a suitable solution, as it had normal survival rates, biological traits, and reproductive traits. For Wolbachia transinfection, the injected M. hiroglyphicus exhibited a 61.33% positivity rate for Wolbachia. Wolbachia density observed in injected M. hiroglyphicus ranged from 3.75 to 4.16 x 104 copies. Wolbachia was successfully established and transmitted in transinfected M. hiroglyphicus across four generations, with a prevalence rate ranging from 64-80%.In conclusion, the role of Wolbachia on the leafhopper Y. flavovittatus, including providing essential nutrients, inducing reproductive abnormalities, and affecting the acquisition and retention of SCWL phytoplasma. Gene annotation analysis of Wolbachia reveals potential genetic factors that may play a role in these processes. Additionally, the successful transfer of Wolbachia to M. hiroglyphicus highlights the importance of further studying its effects on growth, reproduction, and phytoplasma transmission. These findings provide useful information that may be utilized in developing a new control strategy for vectors and phytoplasma diseases.