Larval Water Research Articles

Abiotic factors shape mosquito microbiomes that enhance host development.

Metazoans rely on interactions with microorganisms through multiple life stages. For example, developmental trajectories of mosquitoes can vary depending on the microorganisms available during their aquatic larval phase. However, the role that the local environment plays in shaping such host-microbe dynamics and the consequences for the host organism remain inadequately understood. Here, we examine the influence of abiotic factors, locally available bacteria, and their interactions on the development and associated microbiota of the mosquito Aedes albopictus. Our findings reveal that leaf detritus infused into the larval habitat water, sourced from native Hawaiian tree "ōhi'a lehua Metrosideros polymorpha, invasive strawberry guava Psidium cattleyanum, or a pure water control, displayed a more substantial influence than either temperature variations or simulated microbial dispersal regimes on bacterial community composition in adult mosquitoes. However, specific bacteria exhibited divergent patterns within mosquitoes across detrital infusions which did not align with their abundance in the larval habitat. Specifically, we observed a higher relative abundance of a Chryseobacterium sp. strain in mosquitoes from the strawberry guava infusion than the pure water control, whereas the opposite trend was observed for a Pseudomonas sp. strain. In a follow-up experiment, we manipulated the presence of these two bacterial strains and found larval developmental success was enhanced by including the Chryseobacterium sp. strain in the strawberry guava infusion and the Pseudomonas sp. strain in the pure water control. Collectively, these data suggest that interactions between abiotic factors and microbes of the larval environment can help shape mosquito populations" success.

A trade-off between desiccation resistance and developmental humidity for pupation height in the North Indian seasonal population of Drosophilid-Zaprionus indianus

Drosophila larvae and pupae are vulnerable to seasonal abiotic stressors such as humidity and temperature. In wild low-humidity habitats, desiccation stress can occur as Drosophila larvae forsake wet food in search of a drier pupation site. Henceforth, the hypothesis that developmental humidity impacts pupation height, affecting larval and pupae water balance and fitness-related traits, was examined. Accordingly, warm-adapted Drosophilid- Zaprionus indianus from two seasons were reared under season-specific simulated conditions, with significantly varying relative humidity (summer RH: 40%; rainy RH: 80%), but nearly identical temperatures. A trade-off between pupation height and developmental humidity was observed. Drier summer conditions lead to pupae wandering farther from drier glass surfaces, resulting in higher pupation height (17.3 cm) while rainy pupae prefer wet food, resulting in lower pupation height (7.12 cm). Additionally, density-dependent pupation height was developmental humidity-specific, with most rainy-season pupae pupated on wetter food, while dry summer pupae pupated on glass surfaces or cotton. Nevertheless, flies from far pupation exhibited greater desiccation resistance, fecundity, and copulation duration than those from near pupation. The cuticular lipid mass of larvae and pupae was higher during far-than-near pupation, indicating decreased water loss rates compared to near-pupation. Finally, pupae eclosion (%) was unaffected by greater humidity (85%) in either season. Still, it considerably decreased at lower humidity (RH: 0% and 38%) for rainy pupae, further supporting the selection of low-humidity desiccation resistance in pupae. In conclusion, low humidity is crucial for survival of pre-adult stages of Zaprionus indianus under desiccation stress and for preference of pupation site.

Pikeperch (Sander lucioperca) larval rearing optimization: utilization of lactic acid bacteria for improving microbiome diversity and digestive enzyme activity

Lactic acid bacteria (LAB) probiotics were evaluated for their impact on the microbiota and development of pikeperch (Sander lucioperca) larvae during their initial feeding stage (first 21 days). Pikeperch larvae were exposed to LAB probiotics in two ways: (1) via the live-feed only (Treatment 1, live-feed) or (2) via the live-feed and the larval culture water (Treatment 2, probiotic) in comparison to a control group without LAB supplementation. Total length (TL), myomere height (MH), and survival rate were significantly increased in the probiotic compared the Control group. The administration of probiotics significantly positively influenced the microbiome’s diversity. Specifically, the relative abundance of Cytophagales decreased and that of several other taxa increased in both probiotic treatments although differences between treatments became statistically insignificant by day 21. Furthermore, the different treatments had distinct and significant impacts on digestive enzyme development such as protease and lipase, with the most prominent differences occurring at seven days post-hatching (dph). Taken together, these results indicate that the use of LAB in both the live-feed and rearing water has a positive impact on the larvae microbiota and digestive enzyme development in turn positively impacting their development and viability under intensive rearing conditions.

Environmental dynamics and seasonal wing physiological changes of coastal mosquito vectors (Diptera: Culicidae) in Central Thailand

Mosquitoes (Diptera: Culicidae) are small hematophagous insects that serve as disease vectors by transmitting pathogens to humans and animals. This study was conducted on the coastline of Samut Songkhram Province in Thailand's central region to evaluate the impact of environmental factors on coastal mosquitoes, namely Anopheles epiroticus and Culex sitiens. A zero-inflated negative binomial (ZINB) regression was employed to assess the relationship between the density of larval mosquitoes and water quality factors. Spearman's rank correlation was used to examine the relationship between the number of adult mosquitoes and meteorological factors, and the geometric morphometric (GM) technique was used to study the wing size and shape of adult mosquitoes throughout different seasons. The ZINB analysis revealed a significant negative relationship (p < 0.05) between dissolved oxygen, high-range phosphate, and the number of An. epiroticus larvae. Conversely, in the case of Cx. sitiens, there was a significant positive relationship between nitrate-nitrogen and larval density (p < 0.01), while dissolved oxygen exhibited a significant negative relationship (p < 0.05). Regarding adult mosquitoes, relative humidity was positively correlated with the abundance of adult Cx. sitiens (r = 0.85, p < 0.01), whereas each meteorological parameter—temperature, relative humidity, rainfall, wind speed, and atmospheric pressure—was negatively correlated with adult An. epiroticus abundance (p < 0.05 for each). The GM analysis revealed statistically significant seasonal variations in wing shapes for both species (p < 0.05 for all seasonal comparisons), underscoring their adaptation to seasonal environmental conditions in coastal habitats.

Experimental Infection and Treatment of Vibrio alginolyticus and Vibrio splendidus in Captive Bred Spotted Seahorse, Hippocampus kuda Larvae

Due to their distinctive look, seahorses are famous in the aquarium trade and traded worldwide for their alleged therapeutic benefits. The present study was conducted to identify the causative agents for larval mortality of Hippocampus kuda and to find a treatment to reduce larval mortality at an ornamental fish hatchery in Sri Lanka. Bacteria causing vibriosis in H. kuda larvae were isolated and identified using morphological and biochemical tests. The disc diffusion method was used with 30 μg of chloramphenicol, streptomycin, tetracycline and ampicillin to evaluate antibiotic susceptibility of isolated bacteria. As pathogenic vibrios were more sensitive to tetracycline, the experimentally infected H. kuda larvae were exposed to 20-50 μg/L tetracycline for 30 days. V. alginolyticus, Vibrio splendidus, V. parahaemolyticus, V. mimicus, Aeromonas hydrophila and Plesiomonas shigelloides were found in H. kuda larvae, while V. alginolyticus and V. splendidus were encountered repeatedly. V. alginolyticus, V. mimicus, V. fluvialis, Aeromonas sp., Micrococcus spp., M. leteus and Bacillus circulans were identified from the larval culture water. V. alginolyticus was sensitive to chloramphenicol and tetracycline, while V. splendidus was sensitive to tetracycline. Tetracycline, 30 mg/L as a bath treatment, effectively treats vibriosis in larval rearing tanks of H. kuda.

Assessing Environmental Control on Temporal and Spatial Patterns of Larval Fish Assemblages in a Marine Protected Area

The northern coast of the Iberian Peninsula is an important spawning and nursery area for several marine fish species, some of which are economically exploited by fisheries and under management plans. Larval stages of fish are highly sensitive to environmental change and anthropogenic pressures, and Marine Protected Areas (MPA) can help mitigate the impacts on fish populations. This study investigated the environmental drivers of the temporal and spatial patterns of the larval fish assemblages inhabiting a small coastal MPA along the NW Portuguese Iberian Coast. Seasonal surveys were conducted over two years at nine sampling stations distributed throughout the MPA to collect larval fish samples and water parameters. Results showed that a total of 39 different fish taxa were identified. In terms of abundance, reef-associated species, such as Parablennius gattorugine (54.6%), and marine species that use estuaries as nursery areas, such as Ammodytes tobianus (15.7%) and Clupeidae n.i. (8.8%) dominated the larval fish assemblages. The larval fish assemblages were characterized by a strong temporal pattern that, according to CCA analyses, was related to the temporal variability of water temperature, pH, chlorophyll α, TPM, and also the river flow of an adjacent river. This study showed that 47% of the fish larvae belonged to commercially exploited species, highlighting the importance of this MPA as a nursery area for the early life stages of the fish population. Overall, these new findings emphasize the role of MPAs in ensuring the connectivity of fish species between marine-estuarine habitats and enhancing the conservation of fish populations.

Embryonic and Larval Development of Stinging Catfish, Heteropneustes fossilis, in Relation to Climatic and Water Quality Parameters.

In terms of hatchery-based seed production, one of the most important aquaculture species in Bangladesh is the stinging catfish (Heteropneustes fossilis). Scientific and evidence-based embryonic and larval development research on this fish species in the context of climate change is limited. This experimental study was conducted via induced breeding of stinging catfish using a conventional hatchery system, rearing the larvae in hapas placed in ponds. A series of microscopic observations using a trinocular digital microscope and an analysis of the relationship between larval growth and climate-driven water quality parameters such as temperature, pH, dissolved oxygen, total dissolved solids, alkalinity, and ammonia were performed. During embryonic development, the first cleavage was observed between 30 and 35 min of post-fertilization. Embryonic development (ranging from the 2-cell to the pre-hatching stage) took 21:00 h. Hatching occurred at 22:30 to 23:00 h after fertilization, with an average larvae length of 2.78 ± 0.04 mm. In the post-hatching stage, four pairs of tiny barbels appeared at 36:00 h, and the larvae started feeding exogenously after 72:00 h. These larvae fully absorbed their yolk sacs on the 6th day and attained an average length of 6.44 ± 0.06 mm. Aerial respiration of the larvae was investigated through naked-eye observation on the 10th day of hatching. The average length of the larvae was 32.00 ± 2.0 mm at the end of the 30-day post-hatching observation period. Bivariate correlation analysis showed significant correlations between key climatic variables and water quality parameters under hapa-based larval-rearing conditions. According to canonical correlation analysis, the first canonical function revealed the highest significant correlation between the two sets of variables (r1 = 0.791). The response variable weight of larvae (6.607) was linked to two explanatory variables: pH (0.321) and dissolved oxygen (0.265). For the second canonical correlation function, a positive correlation (0.431) was observed between the two sets of variables. Larval weight (-18.304) was observed to be linked to climatic variables, including air temperature (-0.316) and surface pressure (0.338). Results of this study reveal the subtle correlation between larval growth and water quality driven by climatic variables.

Turbulence affects larval vertical swimming in the cold-water coral Lophelia pertusa

Vertical migration of marine larvae may drastically affect their dispersal, especially if they are spawned in the deep sea. Previous studies have shown that the planktonic larvae of the cold-water coral Lophelia pertusa in still water swim upwards at a speed of ca. 0.5 mm s-1 during a pre-competency period of 3–5 weeks. This behavioral trait is thought to benefit dispersion of larvae as it promotes near surface drift in relatively strong currents. In the ocean however, larvae regularly encounter turbulent water movements potentially impeding their swimming ability. With no apparent stabilizing mechanism, it may be expected that the body orientation of these larvae, and consequently their directed swimming, is sensitive to perturbation by external forces. We investigated the effects of turbulence on vertical swimming of pre-competent L. pertusa larvae by exposing them to relevant turbulence intensities within a grid-stirred tank. Larval movement and water flow were simultaneously recorded, allowing for analysis of individual larval swimming velocities. We showed that the upwards directed swimming speed generally decreased with increasing turbulence, dropping to non-significant in turbulence levels occurring near ocean boundaries. Our results do however suggest that L. pertusa larvae maintain their upwards directed swimming, albeit at reduced speed, in a major part of the water column, thus allowing them to spend part of their planktonic phase in the uppermost ocean layer. This new insight into the behavior of L. pertusa larvae in their natural environment strengthens the notion of the species as one with strong potential for long-distance dispersal. Such information is important for the understanding of L. pertusa population connectivity, and vital when developing tools for modelling of larval transport.

Cup and Pan Behavioral Assays for Assessing Anopheles coluzzii Larval Volatile Responses.

Larval stage Anopheles coluzzii are highly reliant on their olfactory system to locate food sources and to avoid predators and less advantageous microenvironments within their aqueous habitats. The major larval chemosensory appendage, the antenna, is a complex organ with multiple sensory components that is responsible for both gustation and olfaction, thereby facilitating the detection and of both soluble and volatile compounds of biological relevance. Such compounds include food sources, predators, and a range of environmental toxicants. Unlike other mosquitoes, Anopheles coluzzii often position themselves parallel and just under the surface of their aqueous habitats, where they can detect and respond to volatile stimuli. We describe two assays for assessing the behavioral responses of larval anophelines in response to volatile chemicals. The first is a dual-choice, water-surface, inverted-cup assay designed to behaviorally characterize the response valences (attraction, neutral, and repulsion) of anopheline larvae by monitoring and recording the distribution of larvae proximate to chemical volatiles relative to solvent controls. Second, an aqueous-based larval pan behavior assay is designed to assess the responses of mosquito larvae to soluble compounds (as well as potential headspace volatiles) that are released from a point source within larval water. Here, the response valence (attractive, neutral, and repulsive) of mosquito larvae is assessed by quantifying the numbers of larvae in predefined zones proximate to chemical sources.

Phage Therapy for Mosquito Larval Control: a Proof-of-Principle Study.

The mosquito microbiota has a profound impact on multiple biological processes ranging from reproduction to disease transmission. Interestingly, the adult mosquito microbiota is largely derived from the larval microbiota, which in turn is dependent on the microbiota of their water habitat. The larval microbiota not only plays a crucial role in larval development but also has a significant impact on the adult stage of the mosquito. By precisely engineering the larval microbiota, it is feasible to alter larval development and other life history traits of the mosquitoes. Bacteriophages, given their host specificity, can serve as a tool for modulating the microbiota. For this proof-of-principle study, we selected representative strains of five common Anopheles mosquito-associated bacterial genera, namely, Enterobacter, Serratia, Pseudomonas, Elizabethkingia, and Asaia. Our results with monoaxenic cultures showed that Anopheles larvae with Enterobacter and Pseudomonas displayed normal larval development with no significant mortality. However, monoaxenic Anopheles larvae with Elizabethkingia showed delayed larval development and higher mortality. Serratia and Asaia gnotobiotic larvae failed to develop past the first instar. We isolated and characterized three novel bacteriophages (EP1, SP1, and EKP1) targeting Enterobacter, Serratia, and Elizabethkingia, respectively, and utilized a previously characterized bacteriophage (GH1) targeting Pseudomonas to modulate larval water microbiota. Gnotobiotic Anopheles larvae with all five bacterial genera showed reduced survival and larval development with the addition of bacteriophages EP1 and GH1, targeting Enterobacter and Pseudomonas, respectively. The effect was synergistic when both EP1 and GH1 were added together. Our results demonstrate a novel application of bacteriophages for mosquito control. IMPORTANCE Mosquitoes are efficient vectors of multiple human and animal pathogens. The biology of mosquitoes is strongly affected by their associated microbiota. Because of the important role of the larval microbiota in mosquito biology, the microbiota can potentially serve as a target for altering mosquito life-history traits. Our study provides proof of principle that bacteriophages can be used as tools to modulate the mosquito larval habitat microbiota and can, in turn, affect larval development and survival. These results highlight the utility of bacteriophages in mosquito microbiota research and also provide a new potential mosquito control tool.

Systematic and ecological study of mosquitoes (Diptera: Culicidae) in the region of Timgad, Algeria

We conducted an inventory in the wilaya of Batna more particularly the region of Timgad during the period from January 2018 to December of the same year. Sampling is done monthly using the “Dipping” method to collect the mosquito larvae which were then transported to the laboratory for identification. Five mosquito species were collected in the surveyed site presented by 1035 individuals, these were Culex pipiens, Culex theileri, Culex hortensis, Culex deserticola and Culiseta longiareolata which appears to be the most abundant with three significant pullulation peaks. This inventory is supplemented by a physicochemical analysis of water to study the correlation between mosquito larval density and site water quality. J. Biodivers. Conserv. Bioresour. Manag. 2022, 8(1): 49-56

Laboratory studies of oviposition site choice of Toxorhynchites splendens: Role of larval holding-rearing water and predator-prey interactions

Oviposition preference by Toxorhynchites mosquito species is an important aspect of their biological control potential. When ovipositing, the females deposit individual eggs by hovering above the water, indicating that they may be capable of detecting chemical cues from above the water surface. These chemical cues may help the females to determine whether the water is suitable for their larvae. We investigated whether female Tx. splendens has the ability to discriminate against oviposition sites in habitats containing turbid larval/pupal rearing water and predator-prey treatments. In this study, the number of eggs in the test and control cups were compared to assess oviposition preference. According to the results, female Tx. splendens preferred ovipositing in cups with turbid immature water (prey absence) and predator-prey turbid water (prey and predator presences) over ovipositing in filtered water as a control. Results from the third experiment revealed that cups with alive and dead Ae. aegypti larvae proved strongly attractive to Tx. splendens females, while those from conspecific Toxorhynchites larvae caused oviposition deterrence when compared to other treatments in a dual choice test. Our results indicated that both turbid immature water and predator-prey turbid water can efficiently attract Tx. splendens females to deposit eggs. These findings will aid in the development of Toxorhynchites mosquito as a biocontrol agent for the control of Aedes spp. larvae.

Interspecies microbiome transplantation recapitulates microbial acquisition in mosquitoes

BackgroundMosquitoes harbor microbial communities that play important roles in their growth, survival, reproduction, and ability to transmit human pathogens. Microbiome transplantation approaches are often used to study host-microbe interactions and identify microbial taxa and assemblages associated with health or disease. However, no such approaches have been developed to manipulate the microbiota of mosquitoes.ResultsHere, we developed an approach to transfer entire microbial communities between mosquito cohorts. We undertook transfers between (Culex quinquefasciatus to Aedes aegypti) and within (Ae. aegypti to Ae. aegypti) species to validate the approach and determine the number of mosquitoes required to prepare donor microbiota. After the transfer, we monitored mosquito development and microbiota dynamics throughout the life cycle. Typical holometabolous lifestyle-related microbiota structures were observed, with higher dynamics of microbial structures in larval stages, including the larval water, and less diversity in adults. Microbiota diversity in recipient adults was also more similar to the microbiota diversity in donor adults.ConclusionsThis study provides the first evidence for successful microbiome transplantation in mosquitoes. Our results highlight the value of such methods for studying mosquito-microbe interactions and lay the foundation for future studies to elucidate the factors underlying microbiota acquisition, assembly, and function in mosquitoes under controlled conditions.1rc3aWT8PxAqCNbXPTfbB8Video

Drosophila melanogaster as a low-cost and valuable model for studying type 2 diabetes.

Drosophila melanogaster has been used as the most successful invertebrate model for studying metabolic diseases such as type 2 diabetes (T2D). We induced T2D by feeding Drosophila larvae on a high-sugar diet (HSD). The glucose and trehalose, glycogen, lipid, triglyceride, and protein levels were determined in HSD-fed larvae. Moreover, larval food intake, water content, size, and weight in addition to the development until pupation were observed. Levels of Drosophila insulin-like peptides (DILPs 2, 3, and 5), as well as adipokinetic hormone (AKH), were also determined in HSD-fed larvae by quantitative real-time polymerase chain reaction. The results demonstrated that HSD could induce elevated levels of glucose, trehalose, glycogen, and proteins in larvae. The larvae consumed less food intake and were smaller, lighter, and less developed on HSD than those on the control diet. Moreover, the water content of larvae fed HSD was similar to that fed the control diet.HSD induced higher expression of DILP3 and AKH, confirming hyperglycemia with insulin resistance. In sum, Drosophila offers an appropriate model for quick and inexpensive in vivo experimentation on human metabolic diseases.

Microbial Composition in Larval Water Enhances Aedes aegypti Development but Reduces Transmissibility of Zika Virus.

ABSTRACTArthropod-borne viruses comprise a significant global disease burden. Surveillance and mitigation of arboviruses like Zika virus (ZIKV) require accurate estimates of transmissibility by vector mosquitoes. Although Aedes species mosquitoes are established as competent ZIKV vectors, differences in experimental protocols across studies prevent direct comparisons of relative transmissibility. An understudied factor complicating these comparisons is differential environmental microbiota exposures, where most vector competence studies use mosquitoes reared in laboratory tap water, which does not represent the microbial complexity of environmental water where wild larvae develop. We simulated natural larval development by rearing Californian Aedes aegypti larvae with microbes obtained from cemetery headstone water compared to conventional tap water. A. aegypti larvae reared in environmental cemetery water pupated 3 days faster and at higher rates. Mosquitoes reared in environmental water were less competent vectors of ZIKV than laboratory water-reared A. aegypti, as evidenced by significantly reduced infection and transmission rates. Microbiome comparisons of laboratory water- and environment water-reared mosquitoes and their rearing water showed significantly higher bacterial diversity in environment water. Despite this pattern, corresponding differences in bacterial diversity were not consistently observed between the respective adult mosquitoes. We also observed that the microbial compositions of adult mosquitoes differed more by whether they ingested a bloodmeal than by larval water type. Together, these results highlight the role of transient microbes in the larval environment in modulating A. aegypti vector competence for ZIKV. Laboratory vector competence likely overestimates the true transmissibility of arboviruses like ZIKV when conventional laboratory water is used for rearing.IMPORTANCE We observed that A. aegypti mosquitoes reared in water from cemetery headstones instead of the laboratory tap exhibited a reduced capacity to become infected with and transmit Zika virus. Water from the environment contained more bacterial species than tap water, but these bacteria were not consistently detected in adult mosquitoes. Our results suggest that rearing mosquito larvae in water collected from local environments as opposed to laboratory tap water, as is conventional, could provide a more realistic assessment of ZIKV vector competence since it better recapitulates the natural environment in which larvae develop. Given that laboratory vector competence is used to define the species to target for control, the use of environmental water to rear larvae could better approximate the microbial exposures of wild mosquitoes, lessening the potential for overestimating ZIKV transmission risk. These studies raise the question of whether rearing larvae in natural water sources also reduces vector competence for other mosquito-borne viruses.

Larval Diet Abundance Influences Size and Composition of the Midgut Microbiota of Aedes aegypti Mosquitoes.

The midgut microbiota of the yellow fever mosquito Aedes aegypti impacts pathogen susceptibility and transmission by this important vector species. However, factors influencing the composition and size of the microbiome in mosquitoes are poorly understood. We investigated the impact of larval diet abundance during development on the composition and size of the larval and adult microbiota by rearing Aedes aegypti under four larval food regimens, ranging from nutrient deprivation to nutrient excess. We assessed the persistent impacts of larval diet availability on the microbiota of the larval breeding water, larval mosquitoes, and adult mosquitoes under sugar and blood fed conditions using qPCR and high-throughput 16S amplicon sequencing to determine bacterial load and microbiota composition. Bacterial loads in breeding water increased with increasing larval diet. Larvae reared with the lowest diet abundance had significantly fewer bacteria than larvae from two higher diet treatments, but not from the highest diet abundance. Adults from the lowest diet abundance treatment had significantly fewer bacteria in their midguts compared to all higher diet abundance treatments. Larval diet amount also had a significant impact on microbiota composition, primarily within larval breeding water and larvae. Increasing diet correlated with increased relative levels of Enterobacteriaceae and Flavobacteriaceae and decreased relative levels of Sphingomonadaceae. Multiple individual OTUs were significantly impacted by diet including one mapping to the genus Cedecea, which increased with higher diet amounts. This was consistent across all sample types, including sugar fed and blood fed adults. Taken together, these data suggest that availability of diet during development can cause lasting shifts in the size and composition of the microbiota in the disease vector Aedes aegypti.

Expression of aquaporins in response to distinct dehydration stresses that confer stress tolerance in the Antarctic midge Belgica antarctica

Larvae of the Antarctic midge Belgica antarctica Jacobs (Diptera: Chironomidae) are highly tolerant of diverse environmental stresses, including freezing, severe desiccation, and osmotic extremes. Furthermore, dehydration confers subsequent desiccation and freeze tolerance. While a role for aquaporins—channels for water and other solutes—has been proposed in these dehydration processes, the types of aquaporins involved in dehydration-driven stress tolerance remain unknown. In the present study, we investigated expression of six aquaporins (Drip, Prip, Eglp1, Eglp2, Aqp12L, and Bib) in larvae of B. antarctica subjected to three different dehydration conditions: desiccation, cryoprotective dehydration, and osmotic dehydration. The expression of Drip and Prip was up-regulated under desiccation and cryoprotective dehydration, suggesting a role for these aquaporins in efficient water loss under these dehydration conditions. Conversely, expression of Drip and Prip was down-regulated under osmotic dehydration, suggesting that their expression is suppressed in larvae to combat dehydration. Larval water content was similarly decreased under all three dehydration conditions. Differences in responses of the aquaporins to the three forms of dehydration suggests distinct water management strategies associated with different forms of dehydration stress.

Bio-Control of Anopheles Mosquito Larvae Using Invertebrate Predators to Support Human Health Programs in Ethiopia.

Mosquitoes have been a nuisance and health threat to humans for centuries due to their ability to transmit different infectious diseases. Biological control methods have emerged as an alternative or complementary approach to contain vector populations in light of the current spread of insecticide resistance in mosquitoes. Thus, this study aimed to evaluate the predation efficacy of selected potential predators against Anopheles mosquito larvae. Potential invertebrate predators and Anopheles larvae were collected from natural habitats, mainly (temporary) wetlands and ponds in southwest Ethiopia and experiments were conducted under laboratory conditions. Optimal predation conditions with respect to larval instar, water volume and number of predators were determined for each of the seven studied predators. Data analyses were carried out using the Poisson regression model using one way ANOVA at the 5% significant level. The backswimmer (Notonectidae) was the most aggressive predator on Anopheles mosquito larvae with a daily mean predation of 71.5 larvae (95% CI: [65.04;78.59]). Our study shows that larval instar, water volume and number of predators have a significant effect on each predator, except for dragonflies (Libellulidae), with regard to the preference of the larval instar. A selection of mosquito predators has the potential to control Anopheles mosquito larvae, suggesting that they can be used as complementary approach in an integrated malaria vector control strategy.

Physiological responses to cryoprotectant treatment in an early larval stage of the malaria mosquito, Anopheles gambiae

The development of cryopreservation protocols for Anopheles gambiae could significantly improve research and control efforts. Cryopreservation of any An. gambiae life stage has yet to be successful. The unique properties of embryos have proven to be resistant to any practical cryoprotectant loading. Therefore, we have chosen to investigate early non-feeding first instar larvae as a potential life stage for cryopreservation. In order to determine an appropriate cryoprotective compound, larvae were treated with progressively better glass-forming cryoprotective mixtures. Toxicity evaluation in combination with calorimetry-based water content and supercooling point depression assessments were used to determine the cryoprotectants that could be used for cryostorage of viable larvae. Approximately 35–75% of the larvae were viable after reasonably high osmotic and biochemical challenge. This study provides ample evidence for an active osmoregulatory response in the Anopheles larvae to counter the permeation of cryoprotectants from the surrounding medium. The data show a strong correlation between the larval mortality and water content, indicating an osmoregulatory crisis in the larva due to certain cryoprotectants such as the higher concentrations of ethane diol (ED). The observations also indicate that the ability of the larvae to regulate permeation and water balance ceases at or within 20 min of cryoprotectant exposure, but this is strongly influenced by the treatment temperature. Among the compound cryoprotectants tested, 25% ED + 10% dimethyl sulfoxide (DMSO) and 40% ED + 0.5 M trehalose seem to present a compromise between viability, larval water content, supercooling point depression, and glass forming abilities.

Comparing larval microbiomes of the eastern oyster (Crassostrea virginica) raised in different hatcheries

An ongoing goal of the aquaculture industry is the improvement of oyster larval growth and production. One component of oyster hatcheries that may contribute to the health of oyster larvae is the early establishment and subsequent development of the oyster larval microbiome. The main objectives of this study were to investigate the effects of environmental conditions (hatchery types and spawning seasons) and oyster phenotypes (larval developmental stages) on the composition and diversity of the larval microbiome. In addition, the members of core larval microbiomes were identified and quantified. Microbiome composition and diversity of larval and corresponding hatchery water samples were examined using high throughput sequencing of 16S rRNA gene amplicons and represented three development stages from two different spawning events at four different hatcheries located in Chesapeake Bay tributaries. Larval and water samples from each spawning cohort were taken at 24-hours ‘D’ shape (D-stage), 1-week veliger (V-stage), and 2-weeks pediveliger (PV-stage). Larval microbiomes were significantly different from water microbiomes and were significantly influenced by hatchery and spawning, with hatchery having the greatest effect on microbiome composition. While development stage did not show a significant effect on the larval microbiomes, there was a decrease in species richness from the initial D-stage larvae through the final PV-stage and nMDS clustering patterns showed some separation between early (D- and V-) and late stages (PV-) of development, suggesting a shift towards a more selective microbiome as the larvae developed. A total of 25 members (OTU level) were identified as the core larval microbiome (core OTUs), comprising approximately one quarter of the total relative abundance of the larval microbiome. Core OTUs belonging to genera Alteronomonas and Roseobacter have been shown to offer bivalve larvae some protection against pathogens, while those belonging to family Cryomorphaceae are commonly isolated from microalgal species and most likely indicate an association with oyster larval feeding. These findings underscore the importance of environmental conditions on microbiome development of oyster larvae associated with hatchery success of larval cultivation. Further studies are needed to determine the contribution of the core larval microbiome to oyster health and disease resistance.