Hatching Rate Research Articles

Functional analysis of novel cystatins from Haemaphysalis doenitzi and evaluation of their roles in cypermethrin and λ-cyhalothrin resistance

Currently, the primary strategy for tick control relies on chemical agents. Pyrethrins, which are botanically derived compounds, have demonstrated efficacy in controlling ticks without posing a risk to human or animal health. However, research into pyrethrins' metabolic mechanisms remains sparse. Cystatin, as a reversible binding inhibitor of cysteine protease, may be involved in the initiation of pyrethrin detoxification of Haemaphysalis doenitzi. In this study, two novel cystatins were cloned, HDcyst-3 and HDcyst-4, the relative expression of which was highest in the Malpighian tubules compared with the tick midguts, salivary glands, and ovaries. Prokaryotic expression and in vitro studies revealed that cystatins effectively inhibit the enzymatic activities of cathepsins B and S. RNAi results showed that the reduction of cystatins significantly decreased the engorgement weight, egg mass weight, and egg hatching rate of adult female ticks, and prolonged feeding time by two days. The control rate of rHDcyst-3 and rHDcyst-4 protein vaccination against female adults were 55.9% and 63.2%, respectively. In addition, the tick immersion test showed that cypermethrin and λ-cyhalothrin had significant acaricidal effects against adult unfed H. doenitzi. The qPCR result indicated that compared with the control group, the expression of HDcyst-3 and HDcyst-4 was markedly decreased in the sublethal cypermethrin and λ-cyhalothrin group at LC50. Enzyme activity showed that cypermethrin and λ-cyhalothrin could significantly induce the activities of glutathione S-transferase (GST), carboxylesterase (CarE), and acetylcholinesterase (AchE). The aforementioned results provided indirect evidence that cystatin plays an important role in pyrethrin detoxification and provides a theoretical basis for future acaricide experiments and pest management.

One Genome, Multiple Phenotypes: Would Rhodnius milesi Carcavallo, Rocha, Galvão & Jurberg, 2001 (Hemiptera, Triatominae) Be a Valid Species or a Phenotypic Polymorphism of R. neglectus Lent, 1954?

Species of the Rhodnius genus have a complex taxonomy because the events of phenotypic plasticity and cryptic speciation make it difficult to correctly classify these vectors. During the taxonomic history of the genus, five synonymization events occurred. Additionally, some authors suggest that R. milesi possibly represent only phenotypic polymorphisms of R. neglectus. Thus, we analyzed the specific status of R. milesi in relation to R. neglectus using phylogenetic studies with the mitochondrial gene cytochrome B and the study of reproductive barriers. The phylogenetic reconstruction grouped R. milesi together with R. neglectus from different localities, demonstrating that these taxa represent the same species based on the phylogenetic species concept. Experimental crosses demonstrate the absence of pre- and postzygotic barriers under laboratory conditions. Additionally, when the hatch rates of crosses are compared to intraspecific crosses, it can be noted that they are high and very similar. Finally, the mortality rate of the hybrids does not indicate hybrid inviability, the absence of chromosome pairing errors does not indicate hybrid sterility, and the proportion between male and female hybrids demonstrates that Haldane’s rule was not acting. Therefore, we perform the formal synonymization of R. milesi with R. neglectus.

Pectin-nano zero valent iron nanocomposites for efficient heavy metal removal and bactericidal action against waterborne pathogens — Innovative green solution towards environmental sustainability

This study investigated the effectiveness of a pectin-nano zero-valent iron-based nanocomposite in adsorbing heavy metals in bimetallic form (chromium‑lead mixture), along with assessing its antibacterial properties. The nanocomposite was synthesized using a straightforward dispersion method, employing eco-friendly components like biocompatible pectin sourced from banana peels and nano-scale zero-valent iron. Analytical characterization confirmed the formation of stable, nano-crystalline particles with active interactions between the functional groups of pectin and nano iron. Batch adsorption experiments optimized various parameters such as pH, adsorbent dosage, contact time, metal ion concentration, and temperature to enhance bimetal removal from water. The optimal conditions were determined as pH 8.0, a temperature of 40 °C, 1.0 g/L adsorbent dosage, 75 mg/l initial bimetal concentration, and a contact time of 30 min. Further assessments revealed that the nanocomposite did not induce phytotoxic or ecotoxic effects, confirming its non-toxicity and environmental safety. Biocompatibility studies conducted using zebrafish models showed no adverse effects on hatching, survival, or heart rate. These findings underscore the potential of the nanocomposite as a sustainable and efficient solution for heavy metal remediation in water treatment process.

Does duckweed (Lemna minor) feed inclusion play a role on growth, feed conversion ratio and reproductive performance (fertilization, hatchability and survivability rates) in omnivorous fish? Evidence in Nile tilapia (Oreochromis niloticus‐Linnaeus, 1758)

AbstractFish feed production is fraught with high costs due to the inclusion of expensive fishmeal and animal proteins that can be sourced from aquatic macrophytes. Limited attempts have been made on use of the latter for fish feed production. Further, the quality of feed given to fish is known to affect its growth and reproductive performance. Role of feeds containing duckweed (Lemna minor) at 0%‐control feed, 10%, 15%, 20% and 25% inclusion levels on growth and reproductive performance of Oreochromis niloticus of size 18 ± 1 g were evaluated for 12 weeks. The fish were fed twice daily at 10% body weight at 9.00 a.m. and 4.00 p.m. Length–weight measurements were done fortnightly using a measuring board and a weighing balance, respectively. Female mouth‐brooding fish were used to evaluate reproductive performance indicators, namely %: fertilization, hatchability and survivability. Data were subjected to one‐way analysis of variance followed by post hoc and polynomial orthogonal analysis to identify L. minor diets with significant differences (p < 0.05). Fish fed on a diet containing 10% L. minor inclusion showed significantly better growth performance and feed conversion ratio than those fed on the control diet. All L. minor diets gave good fish condition factors above 1.0. Fish fed on a diet containing 10% L. minor and those fed on the control gave reasonably high survival rates of 85.55% and 83.33%, respectively, whereas those fed on 20% L. minor produced the same growth performance as control. Orthogonal polynomial analysis for the final weights across the L. minor diets – 0%–25% – showed a cubic polynomial model (p = 0.000), whereas final lengths portrayed a linear inverse significant relationship (p < 0.05). Inclusion of L. minor in the diets resulted to slightly better fertilization, hatchability and survivability rates at 10%, 20% and 15%, respectively. In conclusion, inclusion of L. minor from 10% to 20% in fish feeds is recommended for enhancing growth and reproductive performance of O. niloticus.

CARP HATCHERY MANAGEMENT (Cyprinus carpio L.) NATURALLY AT THE PUNTEN AQUACULTURE INSTALLATION (IPB), BATU CITY, EAST JAVA

The large market demand for consumption fish makes some farming communities interested in farming, especially consumption fish farming. Fish hatchery business is the first step to the success of fish farming, because for each growing season the aquaculture business requires a supply of seeds from the hatchery business. Goldfish hatcheries are relatively easy because carp can spawn naturally or artificially. Goldfish have a relatively high fecundity or number of eggs, which ranges from 84,000-110,000 eggs per kilogram of brood. The implementation of carp hatchery techniques includes several processes, namely pond preparation, brood selection, kakaban installation, brood release, spawning, hatching eggs, larval care, checking water quality, seeding, pest and disease control. calculation of fecundity, fertilization rate (FR), hatching rate (HR), and survival rate (SR). The data collection method used in this street vendor is primary data. Primary data include active participation, observation, interviews, and documentation. The carp hatchery technique (Cyprinus carpio L.) at IPB Punten, Batu City is carried out naturally, the carp mother used comes from Punten using a ratio of male to female parents of 1: 3. The feeding method, namely Blind Feeding, obtained an F value of 40,000 eggs, FR 84%, HR 84.5% and SR 90.2%.

Influence of Parental Age on Reproductive Potential and Embryogenesis in the Pepper Weevil, Anthonomus eugenii (Cano) (Col.: Curculionidae).

The pepper weevil (Anthonomus eugenii Cano) is a devastating pest that inflicts severe damage to pepper crops, leading to substantial economic losses. This study investigated the impact of aging on the reproductive success of the pepper weevil. Pepper weevil-infested fruit were harvested from pepper fields and subsequently transferred into an insect cage to facilitate the emergence of adults. The emerged adults were housed in separate cages and allowed to mature until they reached specified ages: 10 days old (young), 20 days old (middle-aged), and 30 days old (old) individuals. Eggs laid by each age group were carefully collected and incubated under controlled laboratory conditions (28 ± 1.5 °C). Several reproductive variables including the number of eggs laid, the percentage of hatched eggs, and the egg incubation period were recorded for each age group. Embryonic development was also monitored daily using a VHX digital microscope at a magnification of 200×. Differences in developmental stages such as the blastoderm, germ band, gastrulation, segmentation, and appendage formation were observed, and the time span of every stage was recorded. The results show that the 10-day-old weevils laid the most eggs and had the highest hatching rate and the shortest developmental time. The 30-day-old weevils laid the fewest eggs and had the lowest hatching rate and longest developmental time. Thus, the pepper weevil age significantly influenced the fecundity, length of time for each embryonic development stage, hatching rate, and incubation period, and should be considered when studying the reproductive biology of this pest insect. This first report of the effect of aging on the reproductive potential of the pepper weevil should enable pepper growers to adopt cultural practices aimed at reducing the pepper weevil populations, thereby helping to protect their crop from this important pest.

Nematicidal Potential of Green Chiretta Extracts Against Meloidogyne enterolobii and M. Incognita: In vitro Assessment for Agricultural Application

Root-knot nematodes pose a persistent threat to various economically important crops in Thailand, with limited available management strategies for farmers. This study aimed to assess the efficacy of aqueous (AE), methanolic (ME), ethanolic (EE), ethyl acetate (EAE), and chloroform (CE) crude extracts derived from green chiretta in controlling Meloidogyne enterolobii and M. incognita under laboratory conditions. The results demonstrated significant inhibitory effects of all crude extracts, except for CE, against both nematode species. Notably, AE (10 and 20 mg/mL), ME (20 mg/mL), and EE (20 mg/mL) treatments exhibited maximum inhibition of hatching, leading to a considerable reduction in nematode hatch rates by 87.5 - 96.8 % for M. enterolobii and 88.5 - 96.5 % for M. incognita, compared with distilled water (DW). In terms of second-stage juveniles (J2s) mortality, AE (10 and 20 mg/mL), ME (20 mg/mL), and EAE (20 mg/mL) demonstrated the highest nematicidal activity, resulting in mortality rates of 94.2 - 100 % for M. enterolobii and 92.2 - 99.2 % for M. incognita. Chemotaxis assays revealed a repellent effect of all extracts on both nematode species, except for the ethyl acetate extract (EAE), which attracted M. incognita J2s without affecting M. enterolobii J2s. The observed efficacy was attributed to the higher concentrations of bioactive compounds, including flavonoids, phenolics, and terpenoids, in the extracts compared to CE. Based on these in vitro findings, it is suggested that AE, ME, EE, and EAE of green chiretta exhibit significant nematicidal activity against M. enterolobii and M. incognita. Further evaluation under greenhouse and field conditions is warranted to ascertain the practical applicability and effectiveness of these extracts in a real-world agricultural setting.

Probiotics Alleviate Microcystin-LR-Induced Developmental Toxicity in Zebrafish Larvae.

Microcystin-LR (MCLR) poses a significant threat to aquatic ecosystems and public health. This study investigated the protective effects of the probiotic Lactobacillus rhamnosus against MCLR-induced developmental toxicity in zebrafish larvae. Zebrafish larvae were exposed to various concentrations of MCLR (0, 0.9, 1.8, and 3.6 mg/L) with or without L. rhamnosus from 72 to 168 h post-fertilization (hpf). Probiotic supplementation significantly improved survival, hatching, and growth rates and reduced malformation rates in MCLR-exposed larvae. L. rhamnosus alleviated MCLR-induced oxidative stress by reducing reactive oxygen species (ROS) levels and enhancing glutathione (GSH) content and catalase (CAT) activity. Probiotics also mitigated MCLR-induced lipid metabolism disorders by regulating key metabolites (triglycerides, cholesterol, bile acids, and free fatty acids) and gene expression (ppara, pparb, srebp1, and nr1h4). Moreover, 16S rRNA sequencing revealed that L. rhamnosus modulated the gut microbiome structure and diversity in MCLR-exposed larvae, promoting beneficial genera like Shewanella and Enterobacter and inhibiting potential pathogens like Vibrio. Significant correlations were found between gut microbiota composition and host antioxidant and lipid metabolism parameters. These findings suggest that L. rhamnosus exerts protective effects against MCLR toxicity in zebrafish larvae by alleviating oxidative stress, regulating lipid metabolism, and modulating the gut microbiome, providing insights into probiotic-based strategies for mitigating MCLR toxicity in aquatic organisms.

Composition, antioxidant and development-promoting activity of fermentation modified crude polysaccharides from stem and leaves of Chenopodium album L.

Chenopodium album L. (CAL) has many bioactive compounds and pharmacological activities. Fermentation is the preferred modification method of releasing target bioactive. We prepared two crude polysaccharides: SLC (the polysaccharide from the unfermented stem and leaves of CAL) and FSLC (the polysaccharide from the fermented stem and leaves of CAL). In vitro antioxidant and composition of SLC and FSLC were compared, and the effects of FSLC on antioxidant activity in the IPEC-J2 cells model and development-promoting activity in the zebrafish model were evaluated. The results revealed that FSLC possesses stronger DPPH, hydroxyl radical scavenging, and reducing power than SLC. The levels of total polysaccharide, polyphenol, and flavonoid, as well as the molar ratio of glucuronic acid increased in FSLC. Compared with SLC, the relative contents of protocatechuic acid, protocatechualdehyde, gentisic acid, vanillic acid, p-coumaric acid, quercetin, ferulic acid methyl ester, hispidulin, diosmetin, cinnamic acid, isorhamnetin, syringic acid and kaempferol in FSLC increased. In IPEC-J2 cells, antioxidant enzyme activities and GSH levels were significantly increased, while the MDA level was reduced by treatment with 0.25 mg/mL FSLC. In the zebrafish model, treatment with 25 ~ 300 μg/mL of FSLC had no harmful impact on the morphology and viability of embryos at 12 ~72 hpf. At 48 and 60 hpf, treatment with 100 ~ 300 μg/mL FSLC increased the hatching rate of embryos. At 72hpf, treatment with 100 ~ 300 μg/mL FSLC could relieve morphological abnormalities caused by LPS in zebrafish and improve the hatching rate of embryos. Together, these results provide useful information on the potential for applying polysaccharides from the stems and leaves of CAL as natural feed additives to exert its antioxidant and development-promoting functions.Graphical

Year-round natural spawning, early ontogeny, and larval rearing of the painted sweetlips, Diagramma pictum (Thunberg, 1792) in a hatchery: Potential as a new species for mariculture

The painted sweetlips, Diagramma pictum, is an important species for commercial fisheries, game fishing, and the marine aquarium trade. However, limited knowledge is available about the captive breeding and early developmental stages of this species. This study presents the first report on natural spawning, early ontogeny, and appropriate rearing conditions (temperature, salinity, and initial prey) for larviculture of D. pictum. Natural spawning of D. pictum in captivity was observed year-round for two years, from 1 June 2019 to 31 July 2021. During this 792-day period, females produced 127 spawns, with 77.2% ± 27.2% resulting in fertilized eggs. The median fecundity per spawn was 841 eggs with an interquartile range of 1173 eggs, and the average hatch rate was 59.3 ± 22.0%. Fertilized eggs were spherical, buoyant, and had a diameter of 0.78 ± 0.02 mm. Embryonic development lasted 14 h and 30 min at 28.0 °C. Newly hatched larvae measured 1.81 ± 0.20 mm in total length (LT), with 26 myomeres and an oil globule in the ventroanterior area of the yolk sac. Additionally, the ultrastructural observations of the micropyle, accessory openings, neuromast, and head spines of eggs and early larvae were reported for the first time. At 3 days post-hatch (dph), the larvae measured 2.59 ± 0.16 mm LT at full yolk sac and oil globule resorption. First feeding was noted at this stage, as evidenced by the expansion of the digestive tract filled with live feed. The juvenile stage was reached by 25 dph, and juveniles had an average length of 32.54 ± 2.17 mm LT. Based on hatch and survival activity indices (SAI), we propose optimal water quality parameters for D. pictum larval rearing as a temperature of 26–28 °C and a salinity of 32–33 psu. In the first feeding experiment, survival rates were not significantly different between feeding treatments R (Brachionus rotundiformis; 5 rotifers mL−1) and R + C (B. rotundiformis; 4 rotifers mL−1 + Calanopia elliptica; 1 nauplii mL−1), but both were higher compared to treatment C (C. elliptica; 2 nauplii mL−1) at 9 dph. Our study emphasizes the importance of simultaneous consumption of rotifers and copepods as vital initial prey for the first feeding larvae of D. pictum. Additionally, this co-feeding may enhance larval development through an additive effect. These results demonstrate the potential for establishing captive breeding programs for this species.

Detection and genetic characterization of chicken astrovirus and avian nephritis virus from hatchery to farm

ABSTRACT Astroviruses are an emerging hazard in hatchery management, and they are responsible for great economic losses in poultry farms worldwide. Chicken astrovirus (CAstV) and avian nephritis virus (ANV) are involved in decreased hatching rate, mainly due to embryo deaths, and pale and runted chicks too weak to hatch. Hatchery diseases occur through direct egg infection or vertical transmission. However, the specific role of astroviruses in these diseases is still largely unknown. To elucidate this a high-quality Piedmont hatchery (Italy) was selected based on farm clinical signs and the astrovirus serological positivity in the breeder farm. The viral dynamics were followed from the hatchery to the farm. The presence of both CAstV and ANV was monitored by evaluating the viral load in environmental and bird samples. Interestingly, both viruses were detected in chick yolk sacs at 18 days of incubation, and in the gut contents of 1-day chicks, whereas in the hatchery environmental samples, the eggs' external surfaces and the embryos’ intestinal contents were negative. The CAstV copy number was higher in bird tissues from the hatchery, while a greater ANV load was detected in bird tissues from the farm. Moreover, our data suggest vertical transmission of both viruses. Frequent co-infection was also observed in samples from both the hatchery and the farm. Lastly, the first Italian full-length genome and molecular characterization of ANV was obtained. RESEARCH HIGHLIGHTS Identified the role of the hatchery in astrovirus transmission. Sequenced the avian nephritis virus complete genome. Investigated tissue distribution of astrovirus from egg to chicks. Demonstrated co-infection of ANV/CAstV.

The Improvement Effects of a Nutritional Fortifier on the Reproductive Performance, Sex Steroid Hormone Production, and Health of the Striped Bamboo Shark Chiloscyllium plagiosum.

To explore a method of improving the reproductive performance of the striped bamboo shark, three groups (D0, D1, and D2) of mature individuals were fed squid with (D1 and D2) or without (D0) a nutritional fortifier during the breeding seasons of 2022 and 2023. Compared with the D0 group, the D1 and D2 groups had an increase of 20.90% and 31.34% in total eggs, increases of 32.73% and 41.82% in the proportion of lecithal eggs, and a total 119.07% increase in hatching rate, respectively, in 2022. In 2023, the corresponding increase was 17.12% and 9.91% in total eggs, 19.63% and 12.15% in the proportion of lecithal eggs, 43.37% and 43.94% in fertilization rate, 23.94% and 22.22% in hatchability rate, and 66.70% and 8.70% in the survival rate of fry. Moreover, the levels of serum estradiol, testosterone, progesterone, albumin, and total antioxidant capacity and the levels of ARA, EPA, DHA, n-3 PUFA, and n-6 PUFA in both serum and lecithal eggs significantly increased, while the levels of serum triglyceride and total cholesterol were the opposite (p < 0.05). The results demonstrate that feeding the sharks with a nutritional fortifier can increase spawn production and the quality of eggs, regulate the production of sex steroids, and improve the nutrition of eggs and the health of broodstocks.

Exposure Effects of Environmentally Relevant Concentrations of the Tricyclic Antidepressant Amitriptyline in Early Life Stage Zebrafish.

Antidepressants are one of the most globally prescribed classes of pharmaceuticals, and drug target conservation across phyla means that nontarget organisms may be at risk from the effects of exposure. Here, we address the knowledge gap for the effects of chronic exposure (28 days) to the tricyclic antidepressant amitriptyline (AMI) on fish, including for concentrations with environmental relevance, using zebrafish (Danio rerio) as our experimental model. AMI was found to bioconcentrate in zebrafish, was readily transformed to its major active metabolite nortriptyline, and induced a pharmacological effect (downregulation of the gene encoding the serotonin transporter; slc6a4a) at environmentally relevant concentrations (0.03 μg/L and above). Exposures to AMI at higher concentrations accelerated the hatch rate and reduced locomotor activity, the latter of which was abolished after a 14 day period of depuration. The lack of any response on the features of physiology and behavior we measured at concentrations found in the environment would indicate that AMI poses a relatively low level of risk to fish populations. The pseudopersistence and likely presence of multiple drugs acting via the same mechanism of action, however, together with a global trend for increased prescription rates, mean that this risk may be underestimated using current ecotoxicological assessment paradigms.

Toxicological effect of acetaminophen, metamizole, and nimesulide cocktail on early developmentof zebrafish.

Several countries' most incorrectly discarded medicines are acetaminophen (ACM), metamizole (MTZ), and nimesulide (NMS). These xenobiotics easily reach the aquatic environment; such contamination is very important for the health of humans and other species, yet little explored. To evaluate the cocktail effect of ACM, MTZ, and NMS during zebrafish's initial development. Zebrafish embryos 6-8h post-fertilization (hpf) were exposed to different concentrations of ACM, MTZ, and NMS, separately, to obtain the 50% lethal concentrations (LC50). Next, the embryos were exposed to distinct concentrations of the cocktail (LC50/2, LC50/5, LC50/10, and LC50/20) in a semi-static system. Samples were analyzed 0, 24, 48, and 96h after exposure, and the drugs' concentrations in E3 medium were assessed by high-performance liquid chromatography. For embryotoxicity evaluation, the mortality, hatching, and heart rates; total length; and pericardial and yolk sac areas were determined. In addition, body malformations, edemas, presence of pigmentation, and histopathological assessments were also recorded. The LC50 values obtained for MTZ, ACM, and NMS were 4.69 mgmL-1, 799.98 μgmL-1, and 0.92 μgmL-1, respectively. No difference was observed between the drugs' nominal and observed concentrations at each time point. The cocktail significantly induced mortality and decreased hatching in the LC50/10, LC50/5, and LC50/2 groups. Additionally, body malformations, pigmentation loss, and yolk sac and pericardial edemas were observed in the cocktail groups. The cocktail groups' larvae had decreased total length and slower heart rates compared to the controls (p < 0.05). The histopathological assessment showed that yolk sac edema promoted severe histological changes in the esophageal-intestine junction and intestine in larvae treated with cocktails. Moreover, PAS-positive structures decreased in the esophageal-intestine junction, intestine, and liver in larvae exposed to pharmaceutical cocktails. This study's findings suggest the cocktail of ACM, MTZ, and NMS may be hazardous to aquatic organisms in case of environmental contamination.

Comparative Evaluation of Cytotoxicity of Zinc Oxide Nanoparticles Mixed With Herbal Extract as an Intracanal Medicament: A Zebrafish Model Study.

Objective In this study, zebrafish embryos are used to study the cytotoxic effects of a novel intracanal medication (ICM) based on zinc oxide nanoparticles (ZnO NPs) loaded with polyherbal extracts (Azadirachta indica and Solanum xanthocarpum). Material and methods In the present study, a green and sustainable method was employed for the synthesis of ZnO NPs mixed with bark and seed extracts of Azadirachta indica and Solanum xanthocarpum to be used as a polyherbal ICM. Formulation of ZnO NPs was confirmed with color change in mixture produced upon dissolving zinc acetate dihydrate in distilled water followed by slow addition of sodium hydroxide solution and herbal extracts. The effects of these green synthesized ZnO NPs were evaluated through a zebrafish embryo toxicity test. Embryos were exposed to different concentrations (25, 50, and 100 µg/mL) of synthesized experimental doses of ZnO NP and compared with the control embryos. Toxicological endpoints, such as the zebrafish embryo's survival rate, hatching rate, and heart rate, were noted and described. Results A concentration-dependent increase in mortality rate and hatching delay followed by declined heart rate was observed in green synthesized ZnO NP-treated embryos. The maximum toxicity was observed with an increase in the concentration of 100 µg/mL of the experimental dose, and at a low concentration of 25 µg/mL, it does not effectively show any developmental alteration in zebrafish embryos. Conclusion A novel polyherbal ICM loaded with ZnO NPs exhibited a dose-dependent effect on the heart rate, hatching, and mortality rate of the embryos. At optimal concentrations, the medication demonstrated minimal developmental malformations and cytotoxic effects, indicating its safety for use. However, increasing concentrations of the medication resulted in severe developmental malformations.

Infectivity of Plasmodium parasites to Aedes aegypti and Anopheles stephensi mosquitoes maintained on blood-free meals of SkitoSnack

BackgroundAedes and Anopheles mosquitoes are responsible for tremendous global health burdens from their transmission of pathogens causing malaria, lymphatic filariasis, dengue, and yellow fever. Innovative vector control strategies will help to reduce the prevalence of these diseases. Mass rearing of mosquitoes for research and support of these strategies presently depends on meals of vertebrate blood, which is subject to acquisition, handling, and storage issues. Various blood-free replacements have been formulated for these mosquitoes, but none of these replacements are in wide use, and little is known about their potential impact on competence of the mosquitoes for Plasmodium infection.MethodsColonies of Aedes aegypti and Anopheles stephensi were continuously maintained on a blood-free replacement (SkitoSnack; SS) or bovine blood (BB) and monitored for engorgement and hatch rates. Infections of Ae. aegypti and An. stephensi were assessed with Plasmodium gallinaceum and P. falciparum, respectively.ResultsReplicate colonies of mosquitoes were maintained on BB or SS for 10 generations of Ae. aegypti and more than 63 generations of An. stephensi. The odds of engorgement by SS- relative to BB-maintained mosquitoes were higher for both Ae. aegypti (OR = 2.6, 95% CI 1.3–5.2) and An. stephensi (OR 2.7, 95% CI 1.4–5.5), while lower odds of hatching were found for eggs from the SS-maintained mosquitoes of both species (Ae. aegypti OR = 0.40, 95% CI 0.26–0.62; An. stephensi OR = 0.59, 95% CI 0.36–0.96). Oocyst counts were similar for P. gallinaceum infections of Ae. aegypti mosquitoes maintained on SS or BB (mean ratio = [mean on SS]/[mean on BB] = 1.11, 95% CI 0.85–1.49). Similar oocyst counts were also observed from the P. falciparum infections of SS- or BB-maintained An. stephensi (mean ratio = 0.76, 95% CI 0.44–1.37). The average counts of sporozoites/mosquito showed no evidence of reductions in the SS-maintained relative to BB-maintained mosquitoes of both species.ConclusionsAedes aegypti and An. stephensi can be reliably maintained on SS over multiple generations and are as competent for Plasmodium infection as mosquitoes maintained on BB. Use of SS alleviates the need to acquire and preserve blood for mosquito husbandry and may support new initiatives in fundamental and applied research, including novel manipulations of midgut microbiota and factors important to the mosquito life cycle and pathogen susceptibility.Graphical

Asymmetrical reproductive interference between Aedes aegypti and Aedes albopictus: Implications for coexistence.

Aedes aegypti and Aedes albopictus are two sympatric mosquito species that compete with each other for resources when their breeding habitats overlap. This study examines what happens when sympatric Ae. aegypti and Ae. albopictus mosquitoes' mate with each other and other species by looking at insemination rates, fecundity, and hatchability rate. We performed controlled mating experiments in laboratory setting, assessing both conspecific and interspecific crosses. We measured insemination rates, egg numbers, and hatching success to examine the reproductive interference dynamics between these two distinct mosquito species. In the context of conspecific mating, it was observed that both female Ae. aegypti and Ae. albopictus exhibited high insemination rates, with percentages of 98% and 94% respectively. However, interspecific mating exhibited interesting asymmetries: Ae. albopictus males achieved a notable insemination success rate of 28% when mating with Ae. aegypti females, while Ae. aegypti males achieved only an 8% insemination success with Ae. albopictus females. Additionally, females that mated with interspecific males had reduced production of viable eggs compared to conspecific mating. Most notably, interspecific mating resulted in the production of infertile eggs, while conspecific mating led to successful hatching. The study reveals that Ae. aegypti and Ae. albopictus can asymmetrically interfere with each other's reproduction, causing a 'satyr' effect. This understanding of interspecific competition and reproductive interference in these mosquito species could impact their coexistence in shared breeding habitats.

P-151 Investigation of pregnancy outcomes for blastocysts with microcellular leakage of trophectoderm or inner cell mass from ICSI-induced zona pellucida microslits

Abstract Study question How does ICSI-induced leakage of trophectoderm (TE) or inner cell mass (ICM) from microslits in the zona pellucida (ZP) affect pregnancy outcomes following blastocyst transfer? Summary answer Microcellular leakage of TE and ICM did not affect pregnancy outcome, but blastocysts with ICM leakage above 53.6% had a lower pregnancy rate. What is known already The leakage of TE cells through the ZP (trophectoderm vesicles; TV) has been reported to be more common in ICSI than in conventional IVF, caused by the opening of the ZP with microslits for ICSI. This phenomenon may be improved through the implementation of assisted hatching during transplantation, however, there are few reports investigating its impact on pregnancy.Furthermore, there are no reports on the impact of inner cell mass leakage (inner cell mass vesicles; IV) on pregnancy outcomes, and it is unclear what level of ICM leakage is acceptable for transplantation. Study design, size, duration In this retrospective study, we analyzed clinical medical reports at the Takahashi Women’s Clinic in Japan. A total of 551 patients (1093 blastocysts) who underwent freeze-thaw single blastocyst transfer with laser assisted hatching between January 2019 and October 2023 were included. Blastocysts used for transfer were 2PN embryos obtained by PIEZO-ICSI and cultured in a time-lapse incubator (EmbryoScope+). Participants/materials, setting, methods The blastocysts were classified into control (N = 682), TV (N = 377), and IV groups (N = 34). Pregnancy outcomes were compared using logistic regression analysis, considering patient age, blastocyst grade (Gardner criteria), culture days, BMI, and basal AMH. Bonferroni correction was used for multiplicity correction, considering P &amp;lt; 0.0167 as significant. The ICM leakage rate (%) was calculated as: (leaked ICM area/ICM area before leakage) × 100. Pregnancy cutoff was determined by ROC analysis. Main results and the role of chance The hatching rates for the control, TV, and IV groups were 69.6% (475/682), 78.8% (297/377), and 82.4% (28/34), respectively. Pregnancy rates (fetal heartbeat) were 44.6% (304/682), 57.0% (215/377), and 50.0% (17/34), respectively, and miscarriage rates were 26.3% (80/304), 26.0% (56/215), and 23.5% (4/17), respectively. Logistic regression analysis considering patient background showed no significant differences in hatching, pregnancy, and miscarriage rates (P &amp;gt; 0.05). Furthermore, mean weight, sex ratio, and congenital anomaly rates for the 107 and 7 children born in the TV and IV groups, respectively, were not significantly different than those for the 175 children born in the control group. In examining ICM leakage, the mean area before leakage was 3162.6 µm2 and mean area of leaked ICM was 1292.1 µm2 in the IV group, with an average of 40.9% of ICM leaking outside the clear zone. ROC analysis showed that the cutoff for ICM leakage rate for pregnancy was 53.6%, with significantly lower pregnancy rates when the ICM leakage rate was ≥53.6% (18.2% [2/11] vs 65.2% [15/23], p = 0.0255, Fisher’s exact test). Limitations, reasons for caution This study was conducted at a single in vitro fertilization center. Pregnancy, miscarriage, and fetal anomaly rates at birth related to embryo transplantation are based on ongoing pregnancy dates, and complete live birth data for all pregnancies are not yet available. Wider implications of the findings No differences in pregnancy outcomes or fetal anomalies were found among groups. However, pregnancy rates decreased when ICM leakage rates were over 53.6%. Blastocysts with IV may successfully establish a pregnancy, but considering the leaked ICM proportion is crucial. If considered for transplantation, obtaining sufficient informed consent is essential. Trial registration number not applicable

The impact of non-lethal doses of pyriproxyfen on male and female Aedes albopictus reproductive fitness.

Control of the mosquito Aedes albopictus is confounded by its behavior due to females preferring to oviposition in small natural and artificial containers that are often difficult to remove or treat with insecticides. Autodissemination strategies utilizing highly potent insect growth regulators (IGRs) have emerged as promising tools for the control of this container-inhabiting species. The intended goal of autodissemination approaches is to use mosquitoes to self-deliver an IGR to these cryptic oviposition locations. Previous studies have focused on the efficacy of these approaches to impact natural populations, but little focus has been placed on the impacts on mosquitoes when exposed to non-lethal doses of IGRs similar to the levels they would be exposed to with autodissemination approaches. In this study, the impact of non-lethal doses of pyriproxyfen (PPF) on the reproductive fitness of Ae. albopictus was investigated. Female and male Ae. albopictus mosquitoes were exposed to non-lethal doses of PPF and their fecundity and fertility were measured. To examine the impact of non-lethal doses of PPF, the expression of the ecdysone-regulated genes USP, HR3, and Vg, which are involved in vitellogenesis, was determined. Our results demonstrated a significant reduction in female fecundity and in the blood feeding and egg hatching rates upon exposure to non-lethal doses of PPF. Oocyte development was also delayed in PPF-treated females. Furthermore, exposure to non-lethal doses of PPF altered the expression of the genes involved in vitellogenesis, indicating disruption of hormonal regulation. Interestingly, PPF exposure also reduced the sperm production in males, suggesting a potential semi-sterilization effect. These findings suggest that non-lethal doses of PPF could enhance the efficacy of autodissemination approaches by impacting the reproductive fitness of both males and females. However, further research is needed to validate these laboratory findings in field settings and to assess their practical implications for vector control strategies.

Influence of temperature on hatching success of skipjack tuna (Katsuwonus pelamis): Implications for spawning availability of warm habitats.

Exploring the effects of water temperature on egg development in skipjack tuna (Katsuwonus pelamis) has substantial implications for evaluations of wild spawning habitats. In the present study, we examined the hatching success and duration as a function of temperature from 21 to 33°C under captive environments. A high hatching rate of over 50% between 23 and 31°C was observed, with the shortest hatching duration at 31°C. Because the egg period is vulnerable to predators, a shortened hatching duration with warming water would be ecologically advantageous for K. pelamis, as its main spawning grounds are located in tropical areas.