Profile of the gut microbiota of Pacific white shrimp under industrial indoor farming system.

The gut microbial communities interact with the host immunity and physiological functions. In this study, we investigated the bacterial composition in Litopenaeus vannamei shrimp's gut and rearing water under different host (developmental stage: juvenile and adult; health status: healthy and diseased) and environmental factors (temperature 25 °C and 28 °C; and light intensity: low and high). The PCoA analysis showed that all water samples were clustered together in a quarter, whereas the gut samples spread among three quarters. In terms of functional bacteria, gut samples of adult shrimp, healthy adult shrimp, adult shrimp raised at 28 °C, and juvenile shrimp under high light intensity exhibited a higher abundance of Vibrionaceae compared to each other opposite group. Gut samples of juvenile shrimp, infected adult shrimp, juvenile shrimp with low light intensity, and adult shrimp with a water temperature of 25 °C showed a higher abundance of Pseudoaltromonadaceae bacteria compared to each other opposite group. Gut samples of juvenile shrimp, healthy adult shrimp, adult shrimp raised at a water temperature of 28 °C, and juvenile shrimp with high light intensity showed the higher abundance of Firmicutes/Bacteroidota ratio compared to each other opposite group. Our results showed that L. vannamei juveniles are more sensitive to bacterial infections; besides, water temperature of 28 °C and high light intensity groups were both important conditions improving the shrimp gut bacterial composition under industrial indoor farming systems. KEY POINTS: • Bacteria diversity was higher among shrimp intestinal microbiota compared to the rearing water. • Shrimp juveniles are more sensitive to bacterial infection compared to adults. • Water temperature of 28 °C and high light intensity are recommended conditions for white shrimp aquaculture.

Long-Term Exposure of the Red Cherry Shrimp Neocaridina davidi to Diclofenac: Impact on Survival, Growth, and Reproductive Potential.

The current study was aimed at studying the long-term effects of diclofenac on the freshwater shrimp Neocaridina davidi, concerning survival, somatic growth, and reproduction. In this study, both ovigerous females and males of this species were exposed for 63 d to 0 (control), 0.1, or 1 mg/L of diclofenac. At the highest concentration, significant mortality was detected, and the somatic growth of females was significantly decreased. The percentage of females with a second spawn, observable from day 45, significantly increased at 1 mg/L, while the time between spawns was significantly reduced at both concentrations assayed. However, the gonadal analysis made at the end of the assay in the surviving females showed a significantly lower proportion of advanced oocytes in females exposed to 1 mg/L, as compared to control. Concerning hatching, the percentage of ovigerous females that could have successful hatching was reduced at 1 mg/L of diclofenac, especially for the first spawn. For the second spawn, the low number of juveniles hatched from females exposed to 1 mg/L also showed a significantly higher incidence of morphological abnormalities, such as hydropsy and underdeveloped appendages. Taken together, these results showed that even when diclofenac was able to produce earlier spawns, the reproductive output of each spawn was reduced.

Imbalances in the male reproductive function of the estuarine crab Neohelice granulata, caused by glyphosate.

The effect of glyphosate, both pure and formulated (Roundup Ultramax®), was evaluated on males of the estuarine crab Neohelice granulata, by means of both in vivo and in vitro assays. The in vivo assays comprised the exposure for 30 d to 1 mg/L of the herbicide, until finally assessing weight gain, levels of energy reserves, sperm number per spermatophore, proportion of abnormal spermatophores, and sperm viability. At the end of this assay, significant (p < 0.05) decrease in weight gain and muscle protein levels was detected by effect of both pure and formulated glyphosate. In spermatophores from the vas deferens, a significant (p < 0.05) decrease of the sperm count was observed by effect of Roundup, while a significant incidence (p < 0.05) of abnormal spermatophores was observed either with glyphosate or with Roundup treatment. No changes were seen in the spermatophore area or in vas deferens secretions. Since no sperm mortality was induced by the formulated herbicide, we propose a probable inhibiting effect on spermatogenesis might explain the observed sperm count decrease. In this sense, an in vitro assay was designed by incubating testes and vasa deferentia with Roundup, in order to corroborate the possible interference of glyphosate with the secretion of the androgenic gland hormone that controls the spermatogenesis, in the presence or absence of the androgenic gland. Although the herbicide per se was able to reduce the sperm count to some extent, the increase in the number of spermatozoa/spermatophore produced by the co-incubation with the androgenic gland was completely reverted by the addition of Roundup (1 mg/L of glyphosate a.e.), suggesting that an inhibition on the secretion and/or transduction of the androgenic gland hormone could be taking place.

Effects of atrazine on growth and sex differentiation, in juveniles of the freshwater crayfish Cherax quadricarinatus.

The effect of the herbicide atrazine was assayed in early juveniles of the redclaw crayfish Cherax quadricarinatus. Four cohorts of juveniles (a total of 280 animals) were exposed for 4 wk to each one of three atrazine concentrations (0.1, 0.5 and 2.5mg/L) or a control (0mg/L), from a commercial formulation having 90% of active principle. At the end of the exposure, no significant (p>0.05) differences in either mortality or molting were noted. However, the weight gain and the protein content of abdominal muscle decreased significantly (p<0.05) in the highest atrazine concentration as compared to control, indicating that atrazine acted as a relevant stressor, although at a concentration higher than those reported in the environment. Besides, the proportion of females increased progressively as the atrazine concentration increases, being significantly (p<0.05) higher than that of controls at the highest concentration assayed. Both macroscopic and histological analysis revealed a normal architecture of gonopores and gonads in both control and exposed animals. The obtained results strongly suggest that atrazine could be causing an endocrine disruption on the hormonal system responsible for the sexual differentiation of the studied species, increasing the proportion of female proportion without disturbing the gonad structure.

Effects of glyphosate on sperm quality of the estuarine crab Neohelice granulata, under the "One Health" perspective.

This study was aimed at evaluating the effects of the commercial formulation Roundup Ultramax® on sperm mortality and viability, in terms of the capacity of spermatozoids (spz) to trigger the acrosome reaction (AR), using the estuarine crab Neohelice granulata as a model. To this, an in vivo assay comprising 100 days duration was carried out, on a control group and two groups exposed to the formulation (0.01 mg/L and 0.2 mg/L of glyphosate) under controlled conditions of photoperiod, feeding, and temperature. At the end of the assay, the right vas deferens (VD, proximal and middle portion) was dissected, and after homogenizing it in calcium-free saline solution, the acrosome reaction was induced in the phase containing the spz. In each treatment, the percentage of spz with total and partial AR was calculated, as well as that of dead spz. Compared to the control, crabs exposed to the herbicide showed a significant decrease in spz with full AR, together with an increase in the percentage of spz with partial AR. Furthermore, spz mortality was significantly higher in both glyphosate concentrations compared to the control, in a concentration-dependent manner. On the other hand, abnormal spermatophores, showing expanded walls and coalescence, were observed in a significant percentage in the left VD of the groups treated with the herbicide. The results obtained are compared with those from other studies on several invertebrate and vertebrate species that found inhibition of the AR and abnormal sperm, together with inhibition of spermatogenesis, endocrine disruption, and reduced sperm motility by effect of pure glyphosate and/or different glyphosate formulations. In summary, the available evidence highlights the possible impact of glyphosate on sperm quality, in a wide variety of species.

Embryology of the spider crabs Leurocyclus tuberculosus (H. Milne-Edwards & Lucas 1842) and Libinia spinosa (H. Milne-Edwards 1834) (Brachyura, Majoidea).

The embryonic development of the spider crabs Leurocyclus tuberculosus and Libinia spinosa was divided into five periods based on the differentiation of: (I) cleavage, (II) embryonic primordium, III) optic lobes, (IV) optic lobes pigmented and (V) chromatophores presence. Different traits such as spines, setae and telson morphology distinguish the two species from period III until hatching. Egg volume was greater in Leurocyclus tuberculosus than in Libinia spinosa. The duration of each period was different during development. Whereas in Leurocyclus tuberculosus period II (morphogenesis) is the longest, in Libinia spinosa the period IV is the longest. Complete embryonic development at 14'C lasted 36.7 +/- 3.1 days in Leurocyclus tuberculosus and 57.4 +/- 4.4 days in Libinia spinosa.

Seminal fluid and spermatophore production in a western Atlantic invasive swimming crab, Charybdis hellerii, reveals a different pattern to Portunoidea.

The male reproductive system in Portunoidea is voluminous in order to produce the sperm plug, avoiding sperm competition. The portunid crab, Charybdis hellerii, is a successful invasive species, and this study describes the male reproductive system under light and electron microscopy and evaluates the gonadosomatic index and spermatophore dehiscence compared to other species that produce a sperm plug. The reproductive system is small, and its gonadosomatic index much lower than those of other Portunoidea that produce a sperm plug. The spermatophores are stored in the anterior part of the vas deferens, which is uncommon in Brachyura. Lateral outpocketings are found in the short, middle, and posterior parts of the vasa deferentia, where spermatophores are not found. The seminal fluid composition is different from the species that produce sperm plug. The usual portunid spermatophore wall is absent and the round coenospermic to even cleistospermic spermatophores are enclosed by secretions. This mucous-type spermatophore undergoes dehiscence in seawater or when passing through the gonopod. C. hellerii differ from other Portunoidea described regarding a low amount of seminal fluid production, spermatophore storage, and not show traits of producing sperm plugs. These different male reproductive features can lead to different female sperm storage in Portunoidea.

Brothers are better than nothing: first report of incestuous mating and inbreeding depression in a freshwater decapod crustacean.

This study aimed at evaluating the effect of one generation of full-sibling mating on traits related to the fitness of the gregarious freshwater shrimp Neocaridina davidi, both under optimal and stressful (i.e. starvation) laboratory conditions. Females were maintained either with their brothers (Inbreeding treatment) or non-brothers (Outbreeding treatment), and the first and second broods were used to evaluate egg production and juvenile quality, respectively. The latter was analyzed in a 60-day period following hatching under optimal rearing conditions, and in a 20-day period following hatching under food deprivation conditions. All surviving females from both treatments mated and spawned, indicating that mating with brothers was as likely as mating with non-brothers. With respect to offspring production, inbreeding had no effect on fecundity and fertilization success, but negatively affected the number of hatched juveniles. These results suggest that egg loss was higher in inbred clutches, possibly due to lower embryonic survival. On the other hand, the effect of inbreeding on growth was absent for embryos and for juveniles under optimal rearing conditions, while it was significant for juveniles starved for 10 consecutive days following hatching. Inbreeding depression for survival was only detected in juveniles from stages S8-S9 under optimal rearing conditions. Overall, the present results show that N. davidi fitness decreases after one generation of full-sibling mating. The use of a potentially weak food deprivation protocol and/or the evaluation of only one inbred generation could explain the absence or subtle inbreeding depression for some of the evaluated traits. No evident relationship between life-history traits and the existence and magnitude of inbreeding was found when comparing our results with those previously reported in high-fecundity marine crustaceans and low-fecundity terrestrial crustaceans.

Female energy dynamics in the southernmost fiddler crab: Mixed breeding strategy in Leptuca uruguayensis.

At the south of its geographical distribution, the fiddler crab Leptuca uruguayensis shows two population spawning events, with more than 65 % of the females being ovigerous. The aim of this study was to assess the energy dynamics in L. uruguayensis females to estimate how they mix the capital and the income breeding strategies in the first and second spawns of the short reproductive season. The relationship between the hepatosomatic (HIS) and gonadosomatic (GSI) indices was used to assess the contribution of the hepatopancreas to ovary maturation, as indicative of the capital breeding strategy, whereas the feeding rate and the energy available in the food resource (sediment) were estimated to assess the importance of energy intake in ovary maturation, as indicative of the income breeding strategy. Before the first spawn, a significant negative relationship between the HSI and GSI (p < 0.001) and an increase of 28.2 % in the feeding rate were recorded. This suggests that the first spawn was mainly supported by energy reserves in the hepatopancreas, indicating that females used mostly the capital breeding strategy. After the first spawn, total lipid content in the hepatopancreas decreased by 33 %. On the days before the second spawn, the relationship between the HSI and GSI showed a negative but not significant (p = 0.125) trend, whereas the feeding rate increased even more. This suggests that the intense feeding activity provided the most important energetic source for ovary re-maturation, indicating that females used mostly the income breeding strategy. Also, in the second spawn, the GSI was 24.1% smaller, a fact that affected female fecundity and weight of the egg clutches, which were respectively 12 % and 11 % lower than in the first spawn. However, the energy supply allocated to each embryo was equivalent in both spawns. This study shows how L. uruguayensis females mixed the capital and income breeding strategies to take advantage of all available resources to produce two spawns in a short reproductive season.

From copula to fertilization: The reproduction of primitive crab Hypoconcha parasitica (Linnaeus, 1763) (Podotremata: Dromiidae).

This study describes the mating behavior of Hypoconcha parasitica under laboratory conditions highlighting the spermathecal morphology and focusing on the seminal fluid storage and release of spermatozoa. The pairs were kept in aquaria where the mating behavior was recorded and described. The spermathecae of the female were analyzed using scanning electron microscopy, X-ray micro-CT, histology, and histochemistry. No pre- or post-copulatory mate guarding was observed in H. parasitica. The sperm transfer occurred with each pair maintaining the protection shield (bivalve shell) on their dorsum. The pair of spermathecae is covered exclusively by the cuticle, following the Podotremata pattern. Many muscle fiber bundles are attached to the cuticular wall facing toward the cephalothorax cavity, especially covering the lateral and slightly dorso-anterior region toward the apodeme of Sternite 7. The spermathecal organization indicates that the process of sperm release during fertilization occurs through muscular action exerted by the female on the wall of the chamber. Thus, the musculature distribution in Hypoconchinae distinguishes them from described for other Podotremata such as Homolidae, which shows the musculature associated with the spermathecae aperture. Like the Homolidae, the first pleopod in H. parasitica seems to take part in the transfer/ movement of spermatozoa and oocytes between the sternum and abdomen, which forms a temporary chamber where fertilization occurs. In conclusion, the spermathecal morphology and associated structures bring new insights to the mechanisms involved in the sperm storage and fertilization of primitive crabs and how the Dromiidae spermatheca perform a new pattern among the poorly studied Podotremata.

Effects of androgenic gland ablation on growth and reproductive parameters of Cherax quadricarinatus males (Parastacidae, Decapoda).

This work investigates the effects of androgenic gland (AG) ablation on the structure of the reproductive system, development of secondary sexual characters and somatic growth in Cherax quadricarinatus males. The AG ablation, which was performed at an early developmental stage (initial weight: 1.85±0.03 g), had no effect on the somatic growth parameters (specific growth rate and growth increment), but it prevented the re-formation of male gonopores and appendices masculinae. However, the red patch differentiation and chelae size were similar to those in control males. All the ablated animals developed a male reproductive system. Testis structure was macroscopically and histologically normal. The distal portion of the vas deferens (DVD) was enlarged in some animals, with histological alterations of the epithelium and the structure of the spermatophore. Results suggest that the higher growth in males than in females may be due to an indirect effect of the AG on energy investment in reproduction rather than to a direct effect of an androgen. This is the first report of a potential action of the AG on the secretory activity of the distal VD and the structural organization of the spermatophore. Although the AG may play a role in the development of male copulatory organs, its association with the red patch development deserves further research. The results obtained in the present study support and complement those from intersexes of the same species.

The influence of seasonal temperature variation on embryonic development in two species of spider crabs (Majoidea).

The aim of this study was to analyze the ecological influence of seasonal temperature variations on embryonic development in two species of Majoidea. Leurocyclus tuberculosus and Libinia spinosa. These species inhabit stable populations in the San José Gulf (42°25'41″S, 64°08'27″O, Patagonia-Argentina) in a wide depth range and seasonal temperature variations from 8 °C to 18 °C. The influence of seasonal temperatures on embryonic development was examined in ovigerous females of both species, through a bioassay in laboratory conditions at 8 °C, 14 °C, and 18 °C from gastrula to hatching. Additionally, monthly female samplings were conducted for one year, characterizing their spatial depth distribution associated with the seawater temperature in the study area and their ovigerous/non ovigerous condition. Laboratory experimental results showed that duration of incubation increased 67 % in L. tuberculosus and 55% in L. spinosa by a decline in temperature from 18 °C to 8 °C with one diapause period. The embryonic development for the 3 assayed temperatures was divided in five periods. The assayed temperatures did not affect the morphology of the embryo, and its growth during each period did not present significant differences among the different temperatures. Field analysis showed significant differences in the spatial distribution of ovigerous females which were associated to the assessed temperatures along the year. This distribution might modulate the development of embryos leading to two spawnings synchronized with the phyto- and zooplankton peaks in the San José gulf. Finally, ecological and potential impact of ocean warming in these species was also addressed.

Male reproductive strategies in two species of spider crabs, Leurocyclus tuberculosus and Libinia spinosa.

Reproductive strategies vary according to season length, individual reproductive traits, and factors associated with the social context such as density and sex ratio of the individuals. The social context predicts the intensity of sexual selection due to its effect on mate choice and intra-sexual competition. Moreover, it exacerbates the costs derived from sexual conflict and allows for the existence of alternative reproductive tactics to counteract such costs. However, these mechanisms are still underexplored in many animal taxa. Majoidea "spider crabs" show a diversity of life history, and behavioral and morphological adaptations for reproduction, which have evolved as an outcome of competition to maximize reproductive success. We aimed to analyze in this study, male reproductive strategies through mating behavior under different social contexts, density of individuals, and males size differences in two species of "spider crabs", Leurocyclus tuberculosus and Libinia spinosa. The experiments were as follows: one female and one male per aquarium (Experiment 1); a female and three males of similar sizes per aquarium (Experiment 2); and a female and three males of distinct sizes per aquarium (Experiment 3). The experiments showed that mating behavior and duration of mate guarding differ between species and among different social contexts. The presence of males with similar or different sizes showed a relevant different behavior, the female copulated with more than one male when males were similar in size (E2), and copulated only with the largest male when males were different in size (E3). In mate guarding, the shape and the use of the chelipeds had an important role: the male of Leurocyclus tuberculosus grasped the female with the chelipeds and did not fight with other males, but the male of Libinia spinosa surrounded the female with the chelipeds and fought with other males. The longest duration of mate guarding was during precopulatory guarding. Precopulatory guarding was present in all the experiments and its duration did not present significant differences among the experiments. However, in both species, the experiments differ in the duration of postcopulatory guarding until spawning. Gonopod morphology revealed important differences between species and are probably associated with sperm competition. The experiments under distinct social contexts linked to the use and shape of the chelipeds and the morphology of the gonopods, could allow to understand the functional significance of the different reproductive strategies.

Fluoxetine exposure disrupts food intake and energy storage in the cichlid fish Cichlasoma dimerus (Teleostei, Cichliformes).

Human pharmaceuticals are pollutants of special concern due to their widespread consumption over the last decades, their high persistence in the environment, and the reported alterations produced on non-target organism. The antidepressant fluoxetine (FLX) exerts its effect by inhibiting serotonin (5-HT) reuptake at the presynaptic membrane, thus increasing brain serotonergic activity. In vertebrates, there is a clear inverse relationship between hypothalamic 5-HT levels and food intake, therefore we hypothesized that FLX would inhibit food intake, and in consequence alter energy metabolism in freshwater fish. The aim of this study was to analyze the effect of FLX on feeding behavior and energy storage of the cichlid fish Cichlasoma dimerus. Adult fish were intraperitoneally injected daily with 2 or 20 µg.g-1 FLX or saline for a 5-day period, during which the 20 µg.g-1 FLX-injected fish exhibited a marked reduction in food intake, consistent with a decrease in total body weight and total hepatocyte area observed at the end of the experiment. Although not statistically significant, a marked 50% decrease in glycogen and lipid content and an increase in protein levels in liver was observed for the 20 µg.g-1 FLX dose. This was evidenced histochemically by a weak PAS positive reaction and an intense Coomasie Blue stain. Taken together, these results suggest that the SSRI antidepressant FLX produces an anorectic effect in adults of C. dimerus, which could alter normal physiological function and, in consequence, have a negative impact on fish growth, reproduction, and population success.

Structural alterations in the male reproductive system of the freshwater crayfish, Cherax quadricarinatus (Decapoda, Parastacidae).

No diseases affecting reproductive performance have been previously reported in freshwater crayfishes. This study aims to characterise one reproductive system abnormality found in males of Cherax quadricarinatus reared in captivity. Fifteen adult males of C. quadricarinatus (70-110 g) were purchased from San Mateo S.A. farm (Entre Ríos, Argentina) each season during 2007. Macroscopic analysis showed that 26.6% of the animals sacrificed in winter presented brownish distal vasa deferentia. Histological analysis showed different levels of structural abnormality in the epithelium of the vasa deferentia and spermatophore. Granular and hyaline haemocytes were identified within the vasa deferentia but no significant differences were found in the sperm count between normal and brownish vas deferens. Histological analysis of the crayfishes sacrificed in autumn also showed these modifications in 22% of the animals, however, they did not show the brownish colour under macroscopic analysis. The similarities between the male reproductive system syndrome in shrimps and the abnormalities found in C. quadricarinatus are notable. An unspecific response to thermic stress is a possible explanation of these structural alterations.

Seminal fluid production and sperm packaging in dromiid crabs (Brachyura, Podotremata).

Reproductive anatomy, including sperm storage structures and sperm transfer, is an important feature used to analyze phylogenetic relationships among taxa. We describe the male reproductive anatomy, seminal fluid production and packaging of spermatozoa in the vas deferens of primitive crabs. In all species of Dromiidae, the testes were tubular type and the vas deferens is a tube with a simple epithelium. The spermatozoa are in a central mass immersed in type I secretion, forming a large spermatic cord. In Moreiradromia antillensis and Dromia erythropus the spermatic cord is surrounded by a more complex secretion layer composed by protein granules and polysaccharides that seem to be responsible to form the sperm plaque which was absent in Hypoconcha. The sperm plaque was found in all Dromiinae species, which may result from the mating and probably act as a barrier to subsequent copulations. These patterns of elongated coenospermic spermatophore found here are unique and different from the spermatophores of all true crabs and may be considered a plesiomorphic state to spermatophoric structure in Brachyura. Our results show a novel type of spermatozoa packaging and new insights into how sperm might be transferred in Podotremata.

Sexual system, reproductive cycle and embryonic development of the red-striped shrimp Lysmata vittata, an invader in the western Atlantic Ocean.

Several decapod crustaceans are invaders, but little is known about the biological characteristics that potentiate the success of these decapods in invaded ecosystems. Here, we evaluate and describe some aspects of the reproductive biology and development of Lysmata vittata, an invasive shrimp species in the Atlantic Ocean. In addition, we intend to provide important insights into the biology of invasion by comparing the reproductive traits of this shrimp with some of the predictions about aquatic invasive species. We used experimental and laboratory observations to evaluate the functionality of protandric simultaneous hermaphroditism (PSH), the macro and microscopic development of the ovarian portion of the ovotestes, the reproductive cycle, and the embryonic development of L. vittata. We confirm the functionality of PSH in L. vittata. This shrimp has a rapid reproductive cycle; the ovarian portion of the ovotestes develops (mean ± SD) 6.28 ± 1.61 days after spawning. Embryonic development also occurs over a short time, with a mean (± SD) of 8.37 ± 0.85 days. The larvae hatch without macroscopically visible yolk reserves. Our study provides evidence that the invasive shrimp L. vittata has reproductive and embryonic developmental characteristics (i.e., short generation time and high reproductive capacity) that may be favorable to the establishment of populations during invasive processes.

Dynamics of energy reserves and the cost of reproduction in female and male fiddler crabs.

The physiological costs of reproduction can be measured as the energy allocated to reproductive activities. In fiddler crabs, females allocate energy to vitellogenesis and brooding, whereas males perform expensive courtship behaviors. We evaluated in a large-scale study the reproduction cost of females and males of Leptuca uruguayensis in a temperate estuary, where their reproductive efforts are synchronized in a short reproductive season. The reproductive investments (vitellogenesis, spermatophore production, and male reproductive behaviors) were measured and related to the dynamics of storage and expenditure of energy reserves (glycogen, total lipids, and total protein) in the hepatopancreas, ovary, and muscle of the enlarged cheliped, throughout one annual cycle. Maximum energy storage occurred in winter, a period of low activity, whereas minimum energy storage occurred during the reproductive period, characterized by expensive activities. The glycogen reserves of the hepatopancreas decreased about 66% in females and 61% in males, suggesting high and similar physiological costs of reproduction between sexes, despite their different reproductive strategies to maximize their fitness.

Losing Reproduction: Effect of High Temperature on Female Biochemical Composition and Egg Quality in a Freshwater Crustacean with Direct Development, the Red Cherry Shrimp, Neocaridina davidi (Decapoda, Atyidae).

Maternal provisioning is particularly important in invertebrates with abbreviated development because large energy reserves must be provided for the developing embryo. In this context, the objective of the present study was to analyze in an aquatic invertebrate with direct development the effect of temperature on female biochemical composition and reserve allocation to maturing ovaries, which determine egg quality. A decapod crustacean, the freshwater shrimp Neocaridina davidi, was used as experimental model. Newly hatched juveniles were exposed to 28 °C or 33 °C. Females showed mature ovaries and spawned at 28 °C (control ovigerous females), but no ovigerous female was found at 33 °C. After a 200-day period, half of the females at 33 °C were transferred to 28 °C, where they rapidly showed mature ovaries and spawned (transferred ovigerous females). Ovigerous females and females that did not spawn at 28 °C (control non-ovigerous females) and at 33 °C (high-temperature non-ovigerous females) were sacrificed to determine their biochemical composition. The number, volume, weight, and biochemical composition of the eggs from transferred and control ovigerous females were also analyzed as indicators of their quality. Female biochemical composition was not influenced by temperature, because control and high-temperature non-ovigerous females had similar lipid, protein, and glycogen contents. However, ovarian maturation and spawning were inhibited at 33 °C, which indicates a negative effect of this temperature on nutrient transfer to the oocytes. This effect was rapidly reversed after females were moved to 28 °C; the eggs from control and transferred ovigerous females were of similar quality, except for a lower protein content in the latter. The present results provide valuable information on reserve allocation to reproduction under thermal stress.

Getting ready for mating: The importance of male touching as an accelerator of ovarian growth in a caridean shrimp.

The present study was aimed at evaluating the effect of male presence on ovarian maturation in juvenile females and the role of potential chemical, visual and tactile cues emitted by males in that physiological process. A highly gregarious caridean shrimp with sexual dimorphism, Neocaridina davidi, was used as experimental model. We tested the hypothesis that male presence accelerates ovarian maturation, mainly through chemical cues. Two experiments were performed. In Experiment 1, juvenile females were reared with adult males, adult females or alone, allowing full contact among shrimps. In Experiment 2, these treatments were evaluated allowing chemical and visual communication, only visual communication, or only chemical communication among shrimps. In both experiments juvenile females were observed once a week under a stereomicroscope to determine ovarian growth rate. Although male presence was not necessary for ovarian maturation, it clearly accelerated the rate of ovarian growth, particularly in the last maturation phase. This lead to relatively longer mature ovaries with higher lipid content. On the contrary, the presence of adult females delayed ovarian maturation in juvenile females, while females reared alone showed an intermediate ovarian growth. All these results suggest that adult males release certain cues that stimulate ovarian maturation, while adult females release cues that delay this physiological process. Neither visual cues nor chemical cues released at a distance from females were responsible, either alone or in combination, for the observed effects. Ovarian growth was only influenced when shrimps were allowed to interact freely, probably because of the "mounting" behavior of males towards females. Tactile cues and/or potential chemical cues released by males during this behavior may mediate male stimulatory effect on ovarian growth. Altogether, present results partially support our initial hypothesis and contribute to increase the limited amount of information available on the role of intraspecific multimodal communication in non-behavioral reproductive processes in invertebrate species.