RNAi-Mediated Knockdown of Tssk1 and Tektin1 Genes Impair Male Fertility in Bactrocera dorsalis

Abstract

The genetic-based sterile insect technique (SIT) is an effective and environmentally safe strategy to diminish populations of agricultural and horticultural insect pests. Functional characterization of genes related to male fertility can enhance the genetic-based SIT. Tssk1 has been involved to control male fertility in both mammals and insects. Moreover, Tektin1 has also been revealed to influence male fertility in both human and mammals. These findings suggested that Tssk1 and Tektin1 identified from Bactrocera dorsalis could be required for male fertility in B. dorsalis. In this study, expression profiles of these two genes were studied at different developmental stages and in various tissues of adult males. Remarkably, it was found that Tssk1 and Tektin1 were highly expressed in the testis of mature adult males of B. dorsalis. Furthermore, Tssk1 and Tektin1 genes were downregulated by using the RNA interference (RNAi) method. Fertility assays including egg laying, hatching, and spermatozoa count were also performed to investigate male fertility of B. dorsalis. Results showed that knockdown of Tssk1 and Tektin1 caused male sterility up to 58.99% and 64.49%, respectively. As expected, the total numbers of spermatozoa were also significantly reduced by 65.83% and 73.9%, respectively. These results suggested that male sterility was happened wing to the low number of spermatozoa. In conclusion, we demonstrate that Tssk1 and Tektin1 are the novel agents that could be used to enhance the genetic-based SIT, or their double-stranded RNA (dsRNA) can be used as biopesticides to control the population of B. dorsalis.

lnc94638 is a testis‐specific long non‐coding RNA involved in spermatozoa formation in Zeugodacus cucurbitae (Coquillett)

“…Knockout of Drosophila testis‐specific lncRNA CR42858 −/− also reduced both the number of spermatozoa and male fertility (Wen et al ., 2016). As was demonstrated in B. dorsalis (Sohail et al ., 2019), the impaired male fertility of Z. cucurbitae was probably due to reduced numbers of spermatozoa, which may represent a deficit in spermatogenesis.…”

“…This result suggested that these lncRNAs play essential roles in male Z. cucurbitae as they showed relatively high expression in this developmental stage. Similarly, a testis‐specific expressed gene, Tektin1 , was found to be abundantly distributed in both pupa and adult males of Bactrocera dorsalis (Sohail et al ., 2019). In multiple developmental stages of B. mori , the Voe1 gene is only expressed in pupae and adults, and this was specifically expressed within the testis (Xie et al ., 2020).…”

“…Additionally, the hatchability of the resultant embryos from the lnc94638 suppressed groups was decreased. A similar result was reported in B. dorsalis , where RNAi‐mediated knockdown of two testis‐specific genes, Tssk1 and Tektin1 , reduced the spermatozoa number and male fertility (Sohail et al ., 2019). Knockout of Drosophila testis‐specific lncRNA CR42858 −/− also reduced both the number of spermatozoa and male fertility (Wen et al ., 2016).…”

“…Then, the testes were punctured using forceps, and immediately 2 μl of flowing sperm was collected. Sperm numbers were counted as previously described (Sohail et al ., 2019). In brief, the sperm was diluted in 1 × PBS equably, and then stained with 4′,6′‐diamino‐2‐phenylindole (Sigma, St. Louis, MO, USA) for 15 min.…”

RNAi in fruit flies (Diptera: Tephritidae): Successes and challenges

“…Most studies focussed on the improvement of SIT by silencing genes related to sexual determination or fertility presented positive results for the production of intersex individuals (Salvemini et al., 2009; Sarno et al., 2010), alteration of the reproduction apparatus (Chen et al., 2008; Thongsaiklaing et al., 2018) and reduction in mating rates (Gui et al., 2017, 2018), sperm production and quality (Cruz et al., 2018; Sohail et al., 2019), and number of eggs laid and hatched (Ali et al., 2017; Vannini et al., 2010; Zheng et al., 2015). However, RNAi has been the object of studies in other fields of physiological importance, involving the role of genes related to symbiotic microbiome (Yao et al., 2016), the increase in the susceptibility of fruit flies to insecticides (Huang et al., 2015a; Wang et al., 2017), and the growth and development of larvae and pupae (Li et al., 2017; Xiong et al., 2016; Yang et al., ,2013, 2015).…”