Insights from different reproductive gene knockdowns via RNA interference in the lady beetle Eriopis connexa: Establishing a new model for molecular studies on natural enemies.

Insect pest control can be achieved by the application of RNA interference (RNAi), a key molecular tool in functional genomics. Whereas most RNAi research has focused on insect pests, few studies have been performed on natural enemies. Validating the efficacy of RNAi in natural enemies is crucial for assessing its safety and enabling molecular research on these organisms. Here, we assessed the efficacy of RNAi in the ladybird beetle Eriopis connexa Germar (Coleoptera: Coccinellidae), focusing on genes related to reproduction, such as vitellogenin (Vg) and its receptor (VgR). In the transcriptome of E. connexa, we found one VgR (EcVgR) and two Vg genes (EcVg1 and EcVg2). These genes have been validated by in silico analyses of functional domains and evolutionary relationships. Five-day-old females were injected with 500 ng/µL of a specific double-stranded RNA (dsRNA) (dsEcVg1, dsEcVg2, or dsEcVgR) for RNAi tests, while nonspecific dsRNA (dsGFP or dsAgCE8.1) was used as a control. Interestingly, dsEcVg2 was able to knockdown both Vg genes, while dsEcVg1 could silence only EcVg1. Additionally, the viability of the eggs was significantly reduced when both Vg genes were knocked down at the same time (after treatment with dsEcVg2 or "dsEcVg1+dsEcVg2"). Ultimately, malformed, nonviable eggs were produced when EcVgR was silenced. Interestingly, no dsRNA treatment had an impact on the quantity of eggs laid. Therefore, the feasibility of RNAi in E. connexa has been confirmed, suggesting that this coccinellid is an excellent Neotropical model for molecular research on natural enemies and for studying RNAi nontarget effects.

Genome-Wide Identification, Expression and Response to Estrogen of Vitellogenin Gene Family in Sichuan Bream (Sinibrama taeniatus).

To enhance our understanding of teleost reproductive physiology, we identified six Sichuan bream (Sinibrama taeniatus) vitellogenin genes (vtg1-6) and characterized their sequence structures. We categorized them into type Ⅰ (vtg1,4,5 and 6), type Ⅱ (vtg2) and type Ⅲ (vtg3) based on differences in their subdomain structure. The promoter sequence of vtgs has multiple estrogen response elements, and their abundance appears to correlate with the responsiveness of vtg gene expression to estrogen. Gene expression analyses revealed that the vitellogenesis of Sichuan bream involves both heterosynthesis and autosynthesis pathways, with the dominant pathway originating from the liver. The drug treatment experiments revealed that 17ß-estradiol (E2) tightly regulated the level of vtg mRNA in the liver. Feeding fish with a diet containing 100 µg/g E2 for three weeks significantly induced vtg gene expression and ovarian development, leading to an earlier onset of vitellogenesis. Additionally, it was observed that the initiation of vtg transcription required E2 binding to its receptor, a process primarily mediated by estrogen receptor alpha in Sichuan bream. The findings of this study provide novel insights into the molecular information of the vitellogenin gene family in teleosts, thereby contributing to the regulation of gonadal development in farmed fish.

Exploring the endocrine disrupting potential of a complex mixture of PAHs in the estrogen pathway in Oreochromis niloticus hepatocytes.

This study aimed to investigate the toxicity and endocrine disrupting potential of a complex mixture of polycyclic aromatic hydrocarbons (PAHs) in the estrogen pathway using hepatocytes of Nile tilapia Oreochromis niloticus, the hepatocytes were exposed to various concentrations of the PAH mixture, and multiple endpoints were evaluated to assess their effects on cell viability, gene expression, oxidative stress markers, and efflux activity. The results revealed that the PAH mixture had limited effects on hepatocyte metabolism and cell adhesion, as indicated by the non-significant changes observed in MTT metabolism, neutral red retention, and crystal violet staining. However, significant alterations were observed in the expression of genes related to the estrogen pathway. Specifically, vitellogenin (vtg) exhibited a substantial increase of approximately 120% compared to the control group. Similarly, estrogen receptor 2 (esr2) showed a significant upregulation of approximately 90%. In contrast, no significant differences were observed in the expression of estrogen receptor 1 (esr1) and the G protein-coupled estrogen receptor 1 (gper1). Furthermore, the PAH mixture elicited complex responses in oxidative stress markers. While reactive oxygen species (ROS) and reactive nitrogen species (RNS) levels remained unchanged, the activity of catalase (Cat) was significantly reduced, whereas superoxide dismutase (Sod) activity, glutathione S-transferase (Gst) activity, and non-protein thiols levels were significantly elevated. In addition, the PAH mixture significantly influenced efflux activity, as evidenced by the increased efflux of rhodamine and calcein, indicating alterations in multixenobiotic resistance (MXR)-associated proteins. Overall, these findings, associated with bioinformatic analysis, highlight the potential of the PAH mixture to modulate the estrogen pathway and induce oxidative stress in O. niloticus hepatocytes. Understanding the mechanisms underlying these effects is crucial for assessing the ecological risks of PAH exposure and developing appropriate strategies to mitigate their adverse impacts on aquatic organisms.

Oxalic Acid Treatment: Short-Term Effects on Enzyme Activities, Vitellogenin Content, and Residual Oxalic Acid Content in House Bees, Apis mellifera L.

Honeybees (Apis mellifera L.) have to face many challenges, including Varroa destructor infestation, associated with viral transmission. Oxalic acid is one of the most common treatments against Varroa. Little is known about the physiological effects of oxalic acid, especially those on honeybees' immune systems. In this study, the short-term effects (0-96 h) of oxalic acid treatment on the immune system components (i.e., glucose oxidase, phenoloxidase, glutathione S-transferase, catalase activities, and vitellogenin contents) of house bees were preliminarily investigated. Oxalic acid contents of bee bodies and haemolymphs were also measured. The results confirm that oxalic acid is constitutively present in bee haemolymphs and its concentration is not affected by treatment. At 6 h after the treatment, a maximum peak of oxalic acid content was detected on bees' bodies, which gradually decreased after that until physiological levels were reached at 48 h. In the immune system, the oxalic acid treatment determined a peak in glucose oxidase activity at 48 h, indicating a potential defence response and an increase in vitellogenin content at 24 h. No significant changes were recorded in phenoloxidase, glutathione S-transferase, and catalase activities. These results suggest a time-dependent response to oxalic acid, with potential immune system activation in treated bees.

The TCTP is essential for ovarian development and oviposition of Rhipicephalus haemaphysaloides.

Tick infestations transmit various infectious agents and result in significant socioeconomic consequences. Currently, the primary focus of tick control efforts is identifying potential targets for immune intervention. In a previous study, we identified a highly conserved protein abundant in tick haemolymph extracellular vesicles (EVs) known as translationally controlled tumour protein (TCTP). We have found that native TCTP is present in various tissues of the Rhipicephalus haemaphysaloides tick, including salivary glands, midgut, ovary, and fat body. Notably, TCTP is particularly abundant in the tick ovary and its levels increase progressively from the blood-feeding stage to engorgement. When the TCTP gene was knocked down by RNAi, there was a noticeable delay in ovarian development, and the reproductive performance, in terms of egg quantity and survival, was also hindered. Our investigations have revealed that the observed effects in ovary and eggs in dsRNA-treated ticks are not attributable to cell death mechanisms like apoptosis and autophagy but rather to the reduction in the expression of vitellogenin (Vg1, Vg2, and Vg3) and ferritin (ferritin 1 and ferritin 2) proteins crucial for ovarian development and embryo survival in ticks. Additionally, phylogenetic analysis and structural comparisons of RhTCTP and its orthologues across various tick species, vertebrate hosts, and humans have shown that TCTP is conserved in ticks but differs significantly between ticks and their hosts, particularly in the TCTP_1 and TCTP_2 domains. Overall, TCTP plays a vital role in tick reproductive development and presents itself as a potential target for tick control in both humans and animals.

Expression and function of the vitellogenin receptor in the hypopharyngeal glands of the honey bee Apis mellifera (Hymenoptera: Apidae) workers.

The vitellogenin receptor (VgR) is essential for the uptake and transport of the yolk precursor, vitellogenin (Vg). Vg is synthesized in the fat body, released in the hemolymph, and absorbed in the ovaries, via receptor-mediated endocytosis. Besides its important role in the reproductive pathway, Vg occurs in nonreproductive worker honey bee, suggesting its participation in other pathways. The objective was to verify if the VgR occurs in the hypopharyngeal glands of Apis mellifera workers and how Vg is internalized by these cells. VgR occurrence in the hypopharyngeal glands was evaluated by qPCR analyses of VgR and immunohistochemistry in workers with different tasks. The VgR gene is expressed in the hypopharyngeal glands of workers with higher transcript levels in nurse honey bees. VgR is more expressed in 11-day-old workers from queenright colonies, compared to orphan ones. Nurse workers with developed hypopharyngeal glands present higher VgR transcripts than those with poorly developed glands. The immunohistochemistry results showed the co-localization of Vg, VgR and clathrin (protein that plays a major role in the formation of coated vesicles in endocytosis) in the hypopharyngeal glands, suggesting receptor-mediated endocytosis. The results demonstrate that VgR performs the transport of Vg to the hypopharyngeal glands, supporting the Ovary Ground Plan Hypothesis and contributing to the understanding of the role of this gland in the social context of honey bees.

Changes in CpG Methylation of the Vitellogenin 1 Promoter in Adult Male Zebrafish after Exposure to 17α-Ethynylestradiol.

Numerous pharmaceutical and industrial chemicals are classified as endocrine-disrupting chemicals (EDCs) that interfere with hormonal homeostasis, leading to developmental disorders and other pathologies. The synthetic estrogen 17α-ethynylestradiol (EE2) is used in oral contraceptives and other hormone therapies. EE2 and other estrogens are inadvertently introduced into aquatic environments through municipal wastewater and agricultural effluents. Exposure of male fish to estrogens increases expression of the egg yolk precursor protein vitellogenin (Vtg), which is used as a molecular marker of exposure to estrogenic EDCs. The mechanisms behind Vtg induction are not fully known, and we hypothesized that it is regulated via DNA methylation. Adult zebrafish were exposed to either dimethyl sulfoxide or 20 ng/L EE2 for 14 days. Messenger RNA (mRNA) expression and DNA methylation were assessed in male zebrafish livers at 0, 0.25, 0.5, 1, 4, 7, and 14 days of exposure; and those of females were assessed at 13 days (n ≥ 4/group/time point). To test the persistence of any changes, we included a recovery group that received EE2 for 7 days and did not receive any for the following 7 days, in the total 14-day study. Methylation of DNA at the vtg1 promoter was assessed with targeted gene bisulfite sequencing in livers of adult male and female zebrafish. A significant increase in vtg1 mRNA was observed in the EE2-exposed male fish as early as 6 h. Interestingly, DNA methylation changes were observed at 4 days. Decreases in the overall methylation of the vtg1 promoter in exposed males resulted in levels comparable to those in female controls, suggesting feminization. Importantly, DNA methylation levels in males remained significantly impacted after 7 days post-EE2 removal, unlike mRNA levels. These data identify an epigenetic mark of feminization that may serve as an indicator of not only estrogenic exposure but also previous exposure to EE2. Environ Toxicol Chem 2024;43:1547-1556. © 2024 SETAC. This article has been contributed to by U.S. Government employees and their work is in the public domain in the USA.

Knockdown of vitellogenin receptor based on minute insect RNA interference methods affects the initial mature egg load in the pest natural enemy Trichogramma dendrolimi.

Vitellogenin receptor (VgR) plays a crucial role in oogenesis by mediating endocytosis of vitellogenin and a portion of the yolk proteins in many insect species. However, the function of VgR in minute parasitoid wasps is largely unknown. Here, we applied Trichogramma dendrolimi, a minute egg parasitoid, as a study model to investigate the function of VgR in parasitoids. We developed RNA interference (RNAi) methods based on microinjection of prepupae in T. dendrolimi. RNAi employs nanomaterial branched amphipathic peptide capsules (BAPC) as a carrier for double-stranded RNA (dsRNA), significantly enhancing delivery efficiency. Also, artificial hosts without medium were used to culture the injected prepupae in vitro. Utilizing these methods, we found that ovarian growth was disrupted after knockdown of TdVgR, as manifested by the suppressed development of the ovariole and the inhibition of nurse cell internalization by oocytes. Also, the initial mature egg load in the ovary was significantly reduced. Notably, the parasitic capacity of the female adult with ovarian dysplasia was significantly decreased, possibly resulting from the low availability of mature eggs. Moreover, ovarian dysplasia in T. dendrolimi caused by VgR deficiency are conserved despite feeding on different hosts. The results confirmed a critical role of TdVgR in the reproductive ability of T. dendrolimi and provided a reference for gene functional studies in minute insects.

Targeting TtVgR via siRNA Knockdown Elicits Ovarian Cell Death in the Tri-spine Horseshoe Crab.

The vitellogenin present in the bloodstream undergoes internalization into developing oocytes through the vitellogenin receptor (VgR), a process mediated by receptor-mediated endocytosis. VgR plays a crucial role in facilitating the accumulation of vitellogenin and the maturation of oocytes. In this study, we characterized a Tachypleus tridentatus vitellogenin receptor (TtVgR) gene from the tri-spine horseshoe crab, revealing a length of 1956 bp and encoding 652 amino acid residues with 12 exons. TtVgR has a molecular weight of 64.26 kDa and an isoelectric point of 5.95. Predictions indicate 85 phosphorylation sites and 7 glycosylation sites within TtVgR. Transcriptional analysis demonstrated specific expression of TtVgR in the ovary and yellow connective tissue. TtVgR was identified and distributed in the plasma membrane of oocytes. The siRNA-mediated TtVgR knockdown significantly reduced the transcriptional activity of TtVgR. This depletion induced excessive ROS production, resulting in DNA damage in ovarian primary cells. TUNEL and flow cytometry analyses confirmed ovarian cell apoptosis following TtVgR knockdown, indicating DNA damage in ovarian primary cells. These findings underscore the importance of TtVgR in ovarian cell development, suggesting its potential involvement in vitellogenesis and oocyte maturation. This knowledge may inform innovative breeding strategies and contribute to the sustainable management and conservation of the tri-spine horseshoe crab.

Lrp13a and Lrp13b serve as vitellogenin receptors in the ovary of zebrafish†.

In oviparous animals, egg yolk is largely derived from vitellogenin, which is taken up from the maternal circulation by the growing oocytes via the vitellogenin receptor. Recently, a novel member of the lipoprotein receptor superfamily termed low-density lipoprotein receptor-related protein 13 was identified and proposed as a candidate of vitellogenin receptor in oviparous animals. However, the roles of low-density lipoprotein receptor-related protein 13 in vitellogenesis are still poorly defined. Here, we investigated the expression, vitellogenin-binding properties, and function of low-density lipoprotein receptor-related protein 13 in zebrafish. Two different lrp13 genes termed lrp13a and lrp13b were found in zebrafish. Reverse transcription polymerase chain reaction and quantitative polymerase chain reaction revealed both lrp13s to be predominantly expressed in zebrafish ovary, and in situ hybridization detected both lrp13s transcripts in the ooplasm of early stage oocytes. Two yeast hybrid studies showed that among eight vitellogenins of zebrafish, Vtg1, 2, and 3 bind to Lrp13a, while Vtg1, 2, and 5 bind to Lrp13b. We created zebrafish lrp13a and lrp13b mutant lines using CRISPR/Cas9. Knockout of lrp13a leads to a male-biased sex ratio and decreased diameter of embryo yolk, while knockout of lrp13b and double knockout of lrp13a and lrp13b leads to the delay of vitellogenesis, followed by follicular atresia. These phenotypes of mutants can be explained by the disruption of vitellogenesis in the absence of Lrp13s. Taken together, our results indicate that both Lrp13a and Lrp13b can serve as vitellogenin receptors in zebrafish among other vitellogenin receptors that are not yet described.

Structural modeling and gene expression analysis of phosvitinless vitellogenin (vgc) in the Indian freshwater murrel, Channa punctatus (Bloch, 1793).

Vitellogenin (Vg) is a female-specific egg-yolk precursor protein, synthesized in the liver of fish in response to estrogens. In the present study, complete gene of phosvitinless vitellogenin (vgc) was sequenced, its 3D structure was predicted and validated by web-based softwares. The complete nucleotide sequence of vgc was 4126 bp which encodes for 1272 amino acids and showed the presence of three conserved domains viz. LPD_N, DUF1943 and DUF1944. The retrieved amino acid sequence of VgC protein was subjected to in silico analysis for understanding the structural and functional properties of protein. mRNA levels of multiple vg genes have also been quantified during annual reproductive cycle employing qPCR. A correlation has been observed between seasonal changes in gonadosomatic index with estradiol levels and hepatic expression of three types of vg genes (vga, vgb, vgc) during ovarian cycle of murrel. During preparatory phase, when photoperiod and temperature are low; low titre of E2 in blood induces expression of vgc gene. A rapid increase in the levels of E2 favours induction of vgb and vga genes in liver of murrel during early pre-spawning phase when photoperiod is long and temperature is high in nature. These results suggest that among three vitellogenin proteins, VgC is synthesized earlier than VgA and VgB during oogenesis.

Vitellogenin and its upstream gene TOR play essential roles in the reproduction of Dermanyssus gallinae.

In Dermanyssus gallinae, a hematophagous mite, the initiation of vitellogenesis induced by blood feeding is essential for its reproduction. However, the precise gene structures and physiological functions of Vg in D. gallinae and its upstream gene, Target of Rapamycin (TOR), have not been fully understood. This study revealed the presence of four homologous genes within D. gallinae, named Dg-Vg1, Dg-Vg1-like, Dg-Vg2, and Dg-Vg2-like, especially, Dg-Vg2-like was firstly identified in the mites. The expression levels of all these Vg genes were significantly higher in adult females than other stages. Following blood feeding, the expression levels of these genes increased significantly, followed by a subsequent decrease, aligning with egg production. Silencing Dg-Vgs by RNA interference (RNAi) led to decreased fecundity and egg hatching rates, as well as abnormal embryonic development, suggesting a vital role for Dg-Vgs in both egg formation and embryonic development. Furthermore, the knockdown of Dg-TOR significantly reduced the expression of Dg-Vgs and negatively impacted the reproductive capabilities of PRMs, indicating that TOR influences PRM reproduction by regulating the expression of Dg-Vgs. In summary, these findings demonstrated the crucial roles of Dg-Vgs and Dg-TOR in PRM reproduction, highlighting their potential as targets for pest control.

Evaluating the role of social context and environmental factors in mediating overwintering physiology in honey bees (Apis mellifera).

In temperate climates, honey bees show strong phenotypic plasticity associated with seasonal changes. In summer, worker bees typically only survive for about a month and can be further classified as young nurse bees (which feed the developing brood) and older forager bees. In winter, brood production and foraging halt and the worker bees live for several months. These differences in task and longevity are reflected in their physiology, with summer nurses and long-lived winter bees typically having large fat bodies, high expression levels of vitellogenin (a longevity-, nutrition- and immune-related gene), and large provisioning glands in their head. The environmental factors (both within the colony and within the surrounding environment) that trigger this transition to long-lived winter bees are poorly understood. One theory is that winter bees are an extended nurse bee state, brought on by a reduction in nursing duties in autumn (i.e. lower brood area). We examined that theory here by assessing nurse bee physiology in both the summer and autumn, in colonies with varying levels of brood. We found that season is a better predictor of nurse bee physiology than brood area. This suggests that seasonal factors beyond brood area, such as pollen availability and colony demography, may be necessary for inducing the winter bee phenotype. This finding furthers our understanding of winter bee biology, which could have important implications for colony management for winter, a critical period for colony survival.

Genetic adaptations for the oceanic success of fish eggs.

Genetic adaptations of organisms living in extreme environments are fundamental to our understanding of where life can evolve. Water is the single limiting parameter in this regard, yet when released in the oceans, the single-celled eggs of marine bony fishes (teleosts) have no means of acquiring it. They are strongly hyposmotic to seawater and lack osmoregulatory systems. Paradoxically, modern teleosts successfully release vast quantities of eggs in the extreme saline environment and recorded the most explosive radiation in vertebrate history. Here, we highlight key genetic adaptations that evolved to solve this paradox by filling the pre-ovulated eggs with water. The degree of water acquisition is uniquely prevalent to marine teleosts, permitting the survival and oceanic dispersal of their eggs.

Intraperitoneal administration of kisspeptin-10 modulates follicle maturation, gonadal steroids, calcium and metabolites in Sterlet sturgeon, Acipenser ruthenus.

Kisspeptin is a multifunctional neurohormone, primarily involved in the regulation of reproduction. We tested whether peripheral administration of kisspeptin10 (KP-10) via intraperitoneal injection or slow release affects reproductive hormones and metabolites in Sterlet sturgeon (Acipenser ruthenus). Plasma and mucus 17ß-estradiol (E2), and testosterone (T), plasma and follicular vitellogenin (VTG) and calcium (Ca) as well as glucose and lipids were determined. Mature Sterlet sturgeon were grouped into six groups: saline i.p injection (control), human kisspeptin (hKP-10) i.p injection; acipenser kisspeptin (aKP-10) i.p injection; hKP-10 (slow release); aKP-10 (slow-release) and no treatment control. No effect for KP-10 on sturgeon body weight was found after 4 weeks of treatment. Multivariate analysis revealed a significant disparity in plasma E2 levels. It was significantly different between groups (time, P = 0.0022). E2 in epithelia mucosa showed significant difference between and within groups in the acute group (time, P = 0.0252; treatment, P = 0.0423; time × treatment, P = 0.0429). T levels were unaffected by treatments (P > 0.05). The presence of synthetic aKP-10 led to an elevation in oocyte and plasma VTG levels (P < 0.05). Prolonged exposure to this peptide resulted in an increase in plasma calcium levels. Simultaneously, there was an augmentation in the number of mature follicles. Regardless of the duration of exposure, aKP-10 significantly elevated plasma glucose levels in Sterlet (P < 0.0). Additionally, KP-10 led to an increase in plasma lipids and cholesterol in Sterlet. Overall, our data support an involvement for KP-10 in the regulation of gonadal steroid hormones, oocyte maturation and metabolite levels in sturgeon, suggesting a positive role for this peptide in the reproductive physiology of this species.

Comparative Study of the Effect of Pollen Substitute Diets on Honey Bees during Early Spring.

The nutritional quality of a colony significantly affects its health and strength, particularly because it is required for population growth in the early spring. We investigated the impact of various artificial pollen substitute diets on colony performance in the Republic of Korea during early spring, a critical period for colony health and growth. The colonies were provided with different diets, including the commercial product Megabee (positive control), our developed diet Test A, and four upgraded versions (Diet 1, Diet 2, Diet 3, and Diet 4) of Test A. The negative control group received no supplementary feed. Over 63 days, we observed 24 experimental colonies and assessed various parameters at the colony and individual levels. The results revealed that Diet 2 had the highest consumption and had the most positive impact on population growth, the capped brood area, colony weight, honey bees' weight, and vitellogenin levels. These findings suggested that Diet 2 is most attractive to honey bees and thus holds great promise for improving colony maintenance and development during the crucial early spring period.

Effect of Guanylate Cyclase-22-like on Ovarian Development of Orius nagaii (Hemiptera: Anthocoridae).

This study identified and characterized the gene encoding recep tor-type guanylate cyclase-22-like (GCY-22; OnGCY) from the pirate bug Orius nagaii, an important biological control agent. The full-length cDNA of the GCY of O. nagaii was obtained by rapid amplification of cDNA ends (RACE); it had a total length of 4888 base pairs (bp), of which the open reading frame (ORF) was 3750 bp, encoding a polypeptide of 1249 amino acid residues. The physicochemical properties of OnGCY were predicted and analyzed by using relevant ExPASy software, revealing a molecular formula of C6502H10122N1698O1869S57, molecular weight of ~143,811.57 kDa, isoelectric point of 6.55, and fat index of 90.04. The resulting protein was also shown to have a signal peptide, two transmembrane regions, and a conserved tyrosine kinase (tyrkc). Silencing OnGCY by RNA interference significantly inhibited ovarian development and decreased fertility in female O. nagaii in the treated versus the control group. Additionally, OnGCY silencing significantly decreased the expression levels of other GCY and Vg genes. Thus, these results clarify the structure and biological function of OnGCY, which has an important role in insect fecundity. The results also provide a reference for agricultural pest control and future large-scale breeding of biological control agents.

Gonad pathology, sex hormone modulation and vitellogenin expression in Chrysichthys nigrodigitatus from Lagos and Epe lagoons within the southern-lagoon system, Nigeria.

Introduction: Estrogenic chemicals in aquatic environments impact fish reproductive health, with vitellogenin protein levels serving as a crucial biomarker for xenoestrogen exposure. Limited knowledge exists on estrogenic effects in tropical environments, prompting an investigation into the influence of environmental estrogens on Chrysichthys nigrodigitatus in Lagos and Epe lagoons. Methods: A total of 195 fish samples underwent analysis for vitellogenin protein, sex hormones (testosterone and 17 ß-estradiol), and gonad pathology in effluent-receiving areas of the specified lagoons. Results: Gonadal alterations were observed in male and female fish, including empty seminiferous tubules and distorted ovaries. Intersex occurred in 3.81% of Lagos and 3.33% of Epe. Testosterone levels were generally higher in females and males from both lagoons, while E2 levels were higher in females from both lagoons, with Lagos showing higher levels than Epe. Vtg levels were higher in males than females in Lagos samples but showed no significant difference in Epe samples. Discussion: Contaminant analysis revealed similar trends in metals (Hg, As, Cr) and phthalates (DEHP, DBP, DEP) in both sexes in the Epe population. Multivariate depictions from the PCA showed sex-specific patterns of metal uptake (Cd) in male fishes at the Lagos Lagoon. The positive association between higher pH loadings and metal and DBP levels in sediment at the Lagos lagoon suggests the influence of higher alkalinity in lower bioavailability of contaminants. Conclusion: Endocrine disrupting effects were observed in male and female Chrysichthys nigrodigitatus in Lagos and Epe lagoons populations, with notable differences in hormone and contaminant concentrations between the two lagoon systems. Identification of specific contaminants and their spatial and temporal trends can inform targeted management and remediation efforts to protect and restore these valuable aquatic ecosystems.

Molecular characterization of Vitellogenin-like1 gene in Sogatella furcifera (Hemiptera: Delphacidae), and its function on reproduction.

In this study, a vitellogenin like1 gene (SfVg-like1) in Sogatella furcifera was identified. The open reading frame (ORF) encoded 1,321 amino acid sequence. Structure analysis reveals that the amino acid sequence of SfVg-like1 has 3 conserved LPD_N, DUF1943 and VWFD domains. Phylogenetic analyses showed that SfVg-like1 was clustered in the same branch with the Vg-like1 of Nilaparvata lugens (100% bootstrap value) compared with other Hemiptera insects Vgs associated with vitellogenesis. Temporo-spatial expression analyses showed that SfVg-like1 expressed during all stages, and in both genders. The relative expression levels of SfVg-like1 mRNA were higher in adults than in nymph developmental stages. The knockdown of SfVg-like1 gene resulted in the inhibition of the ovarian development in female adults, whereas the morphology of the testis in male adults was not been affected. The silence of SfVg-like1 could decrease the relative expression levels of target of rapamycin (SfTOR, GenBank MW193765) and vitellogenin (SfVg, GenBank MH271114) genes significantly in female adults. However, the knockdown of SfTOR or SfVg genes in female adults did not affect the transcript level of SfVg-like1. Therefore, it demonstrated that SfVg-like1 might locate on the upstream signaling pathways of SfTOR and SfVg. These results demonstrate that SfVg-like1 is essential for S. furcifera reproduction, and it could be the potential target for the control of this pest.