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IntroductionFemales of the Northern house mosquito,Culex pipiens, enter an overwintering dormancy, or diapause, in response to short day lengths and low environmental temperatures that is characterized by small egg follicles and high starvation resistance. During diapause,Culex pipiensMajor Royal Jelly Protein 1 ortholog (CpMRJP1) is upregulated in females ofCx. pipiens. This protein is highly abundant in royal jelly, a substance produced by honey bees (Apis mellifera), that is fed to future queens throughout larval development and induces the queen phenotype (e.g., high reproductive activity and longer lifespan). However, the role of CpMRJP1 inCx. pipiensis unknown. MethodsWe first conducted a phylogenetic analysis to determine how the sequence of CpMRJP1 compares with other species. We then investigated how supplementing the diets of both diapausing and nondiapausing females ofCx. pipienswith royal jelly affects egg follicle length, fat content, protein content, starvation resistance, and metabolic profile. ResultsWe found that feeding royal jelly to females reared in long-day, diapause-averting conditions significantly reduced the egg follicle lengths and switched their metabolic profiles to be similar to diapausing females. In contrast, feeding royal jelly to females reared in short-day, diapause-inducing conditions significantly reduced lifespan and switched their metabolic profile to be similar nondiapausing mosquitoes. Moreover, RNAi directed againstCpMRJPIsignificantly increased egg follicle length of short-day reared females, suggesting that these females averted diapause. DiscussionTaken together, our data show that consuming royal jelly reverses several key seasonal phenotypes ofCx. pipiensand that these responses are likely mediated in part by CpMRJP1. 

Sex is determined by multiple factors derived from somatic and germ cells in vertebrates. We have identifiedamhy,dmrt1,gsdfas male andfoxl2,foxl3,cyp19a1aas female sex determination pathway genes in Nile tilapia. However, the relationship among these genes is largely unclear. Here, we found that the gonads ofdmrt1;cyp19a1adouble mutants developed as ovaries or underdeveloped testes with no germ cells irrespective of their genetic sex. In addition, the gonads ofdmrt1;cyp19a1a;cyp19a1btriple mutants still developed as ovaries. The gonads offoxl3;cyp19a1adouble mutants developed as testes, while the gonads ofdmrt1;cyp19a1a;foxl3triple mutants eventually developed as ovaries. In contrast, the gonads ofamhy;cyp19a1a,gsdf;cyp19a1a,amhy;foxl2,gsdf;foxl2double andamhy;cyp19a1a;cyp19a1b,gsdf;cyp19a1a;cyp19a1btriple mutants developed as testes with spermatogenesis via up-regulation ofdmrt1in both somatic and germ cells. The gonads ofamhy;foxl3andgsdf;foxl3double mutants developed as ovaries but with germ cells in spermatogenesis due to up-regulation ofdmrt1. Taking the respective ovary and underdeveloped testis ofdmrt1;foxl3anddmrt1;foxl2double mutants reported previously into consideration, we demonstrated that oncedmrt1mutated, the gonad could not be rescued to functional testis by mutating any female pathway gene. The sex reversal caused by mutation of male pathway genes other thandmrt1, including its upstreamamhyand downstreamgsdf, could be rescued by mutating female pathway gene. Overall, our data suggested thatdmrt1is the only male pathway gene tested indispensable for sex determination and functional testis development in tilapia. 

Abstract Culex pipiens, the northern house mosquito, is a major vector of West Nile virus. To survive the severe winter, adult mosquitoes enter a diapause programme. Extended lifespan and an increase in lipid storage are key indicators of diapause. Post‐translational modifications to histone proteins impact the expression of genes and have been linked to the lifespan and energy utilisation of numerous insects. Here, we investigated the potential contribution of epigenetic alterations in initiating diapause in this mosquito species. Multiple sequence alignment of H3 sequences from other insect species demonstrates a high conservation of the H3 histone inCx. pipiensthroughout evolution. We then compared the levels of histone methylation in the ovaries and fat body tissues of diapausing and non‐diapausingCx. pipiensusing western blots. Our data indicate that histone methylation levels in the ovaries ofCx. pipiensdo not change during diapause. In contrast, H3K27me2 levels decrease more than twofold in the fat body of diapausing mosquitoes relative to non‐diapausing counterparts. H3K27 methylation plays a crucial role in chromosome activation and inactivation during development in many insect species. This is predominantly governed by polycomb repressor complex 2. Intriguingly, a previous ChIP‐seq study demonstrated that the transcription factor FOXO (Forkhead box O) targets the genes that comprise this complex. In addition, H3K27me2 exhibits dynamic abundance throughout the diapause programme inCx. pipiens, suggesting its potential role in the initial activation of the diapause programme. This study expands our understanding of the relationship between alterations in epigenetic regulation and diapause.