Antennal Gland Research Articles

The influence of temperature and β-1,3-glucans on the occurrence of white spot syndrome virus and white spot disease in post-larvae of Penaeus vannamei shrimp

Penaeus vannamei is the main species of shrimp farmed worldwide, and for Ecuador it is the first non-oil economic sector. However, various pathogens, including white spot syndrome virus (WSSV), threaten the sustainability of shrimp farming. P. vannamei larvae are susceptible to WSSV infection via vertical or horizontal transmission. To decrease the incidence of WSSV in PLs, a bioassay was performed by exposing P. vannamei shrimp (PL25), which were negative for WSSV and white spot disease (WSD), to two protective factors, water temperature (T = 24 °C and T = 31 °C), and immunestimulation using β-1,3-glucans (BG) (with and without BG added to the food), using a crossed two-factor design, for 20 days. The incidence of WSSV and WSD was modeled using generalized linear model (GLM). The strength of the association between the response and explanatory variables was estimated using the odds ratio [Exp (Beta)] and interpreted as the level of risk for the incidence of WSSV or WSD in one of the categories compared to the baseline category in the GLM. Odds ratios were considered significant if their 95 % confidence interval did not include the value 1. The results showed that BG had a significant negative effect on the number of WSSV-positive animals. In addition, a lower probability of WSSV infection was determined by combining 31 °C and BG. In this treatment, the WSD injuries were almost undetectable. Most larvae shrimps affected by WSD exhibited injuries to the antennal gland and connective tissue. In situ hybridization analysis revealed that 29 % of shrimp, initially negative by histology, were positive for WSSV. The virus was detected mainly in the nervous tissue of the head, epithelium, connective tissue of the head appendages, and the oral region surrounding the integumental glands. Apoptosis analysis showed a weak signal in nerve tissue but was more intense in epithelial cells of the head appendages, in association with cuticular damage. According to the findings, WSSV could disseminate among shrimp populations by infecting them through the TG and epithelium of the head appendage, followed by connective tissue and nervous tissue. However, applying BG at 31 °C may serve as a protective measure to reduce WSD injuries and restrict WSSV dissemination.

North American crayfish harbour diverse members of the Nudiviridae

Three novel crayfish-infecting nudiviruses from crayfish in North America represent the first genomic confirmation of nudiviruses in crayfish: Faxonius propinquus nudivirus (FpNV), Faxonius rusticus nudivirus (FrNV), and Faxonius virilis nudivirus (FvNV). Histopathology and electron microscopy revealed nuclear infections, including nuclear hypertrophy in hepatopancreatic epithelial cells and the presence of membrane-bound bacilliform virions. Metagenomic sequencing resulted in complete circular genome assembly, and phylogenetic analyses (based on nudivirus core genes) placed these viruses within the unofficial Epsilonnudivirus genus. One of the nudiviruses was detected in the antennal gland of its host, and another is correlated with invasive crayfish decline in one infected lake ecosystem - suggesting a potential route for viral transmission through water, and possible population level impact. This study highlights the importance of genomic and ecological data in elucidating the diversity and evolutionary relationships of the Nudiviridae, while expanding their known diversity and range of host species.

Anatomical and molecular insights into the antennal gland of the giant freshwater prawn Macrobrachium rosenbergii

In this study, the complex organization of the AnG in the giant freshwater prawn Macrobrachium rosenbergii was revealed using various techniques, including conventional histology, histochemistry, scanning electron microscopy, and X-ray tomography. The results showed the diversity of cells in the AnG and the detailed organization of the labyrinth’s tubule into four radiated areas from the central to peripheral zones. The study also demonstrated the expression of some vertebrate kidney-associated homolog genes, aquaporin (AQP), solute carrier family 22 (SLC-22), nephrin, and uromodulin, in the AnG by qPCR. The result of in situ hybridization further showed the localization of SLC-22 and AQP transcript in the bladder and labyrinth’s epithelium, specifically in regions 2, 3, and 4. Additionally, the study revealed neuropeptide expressions in the AnG by qPCR and in situ hybridization, i.e., crustacean hyperglycemic hormone (CHH) and molt inhibiting hormone (MIH), implying that the AnG may have a role in hormone production. Moreover, male and female prawns exhibited different levels of AQP, SLC-22, nephrin, and CHH expressions during the premolt and intermolt stages, suggesting a crucial role relevant to the molting stages. In conclusion, this study clarified the complex structure of the AnG in M. rosenbergii and demonstrated for the first time the expression of vertebrate kidney-associated genes and the possible endocrine role of the AnG. Further investigation is needed to clarify the role of these genes, particularly during ecdysis. The implications of these findings could significantly advance our understanding of the AnG in decapod crustaceans.

The role of the antennal glands and gills in acid-base regulation and ammonia excretion of a marine osmoconforming brachyuran

The excretory mechanisms of stenohaline marine osmoconforming crabs are often compared to those of the more extensively characterized euryhaline osmoregulating crabs. These comparisons may have limitations, given that unlike euryhaline brachyurans the gills of stenohaline marine osmoconformers possess ion-leaky paracellular pathways and lack the capacity to undergo ultrastructural changes that can promote ion-transport processes in dilute media. Furthermore, the antennal glands of stenohaline marine osmoconformers are poorly characterized making it difficult to determine what role urinary processes play in excretion. In the presented study, ammonia excretory processes as well as related acid-base equivalent transport rates and mechanisms were investigated in the Dungeness crab, Metacarcinus magister – an economically valuable stenohaline marine osmoconforming crab. Isolated and perfused gills were found to predominantly eliminate ammonia through a microtubule network-dependent active NH4+ transport mechanism that is likely performed by cells lining the arterial pockets of the gill lamella where critical Na+/K+-ATPase detection was observed. The V-type H+-ATPase – a vital component to transbranchial ammonia excretion mechanisms of euryhaline crabs - was not found to contribute significantly to ammonia excretion; however, this may be due to the transporter's unexpected apical localization. Although unconnected to ammonia excretion rates, a membrane-bound isoform of carbonic anhydrase was localized to the apical and basolateral membranes of lamella suited for respiration. Urine was found to contain significantly less ammonia as well as carbonate species than the hemolymph, indicating that unlike those of some euryhaline crabs the antennal glands of the Dungeness crab reabsorb these molecules rather than eliminate them for excretion.

Analysis of adaptive molecular mechanisms in response to low salinity in antennal gland of mud crab, Scylla paramamosain

As an important marine aquaculture species, the mud crab (Scylla paramamosain) is a good candidate for studying the osmoregulatory mechanism of crustaceans. While previous studies have focused on the osmoregulatory function of the gills, this study aims to explore the osmoregulatory function of the antennal glands. By the comparative transcriptomic analysis, we found the pathways of ion regulation including “proximal tubule bicarbonate reclamation” and “mineral absorption” were activated in the antennal glands of the crabs long-term dwelling in low salinity. The enhanced ionic reabsorption was associated with up-regulated ion transport genes such as NKA, CA-c, VPA, and NHE, and with energy metabolism genes such as MDH, SLC25, and PEPCK. The upregulation of NKA and CA-c was also verified by the increased enzyme activity. The lowered osmolality and ion concentration of the hemolymph and the enlarged labyrinth lumen and hemolymph capillary inside the antennal glands indicated the infiltration of external water and the responsively increase of urine excretion, which explained the requirement of enhanced ionic reabsorption. To further confirm these findings, we examined the change of gene expression, enzyme activity, internal ion concentration, and external ion concentration during a 96 h low salinity challenge with seven intervals. The results were basically consistent with the results as shown in the long-term low salinity adaptation. The present study provides valuable information on the osmoregulatory function of the antennal glands of S. paramamosain. The implication of this study in marine aquaculture is that it provides valuable information on the osmoregulatory mechanism of mud crabs, which can be used to improve their culture conditions and enhance their tolerance to salinity stress. The identified genes and pathways involved in osmoregulation can also be potential targets for genetic selection and breeding programs to develop more resilient mud crab strains for aquaculture.

Protection of a defensive symbiont does not constrain the composition of the multifunctional hydrocarbon profile in digger wasps.

Hydrocarbons (HCs) fulfil indispensable functions in insects, protecting against desiccation and serving chemical communication. However, the link between composition and function, and the selection pressures shaping HC profiles remain poorly understood. Beewolf digger wasps (Hymenoptera: Crabronidae) use an antennal gland secretion rich in linear unsaturated HCs to form a hydrophobic barrier around their defensive bacterial symbiont, protecting it from brood cell fumigation by toxic egg-produced nitric oxide (NO). Virtually identical HC compositions mediate desiccation protection and prey preservation from moulding in underground beewolf brood cells. It is unknown whether this composition presents an optimized adaptation to all functions, or a compromise due to conflicting selection pressures. Here, we reconstitute the NO barrier with single and binary combinations of synthetic linear saturated and unsaturated HCs, corresponding to HCs found in beewolves. The results show that pure alkanes as well as 3 : 1 mixtures of alkanes and alkenes resembling the composition of beewolf HCs form efficient protective barriers against NO, indicating that protection can be achieved by different mixtures of HCs. Since in vitro assays with symbiont cultures from different beewolf hosts indicate widespread NO sensitivity, HC-mediated protection from NO is likely important across Philanthini wasps. We conclude that HC-mediated protection of the symbiont from NO does not exert a conflicting selection pressure on the multifunctional HC profile of beewolves.

A new freshwater amphipod (Amphipoda, Gammaridae) from the Fakıllı Cave, Düzce Türkiye: Gammarus kunti sp. nov.

Aquatic species (such as fish, amphipods, isopods, hirudineans etc.) adapted to environmental conditions can live in caves connected to groundwater. The species of Niphargus and Gammarus are the most commonly encountered amphipods in caves. Türkiye is very rich in terms of karst areas and is home to more than 2000 known caves. Fakıllı Cave, located in Düzce Province in the Western Anatolian Region, has a length of 1071 m. A new amphipod species belonging to the Gammarus genus has been identified from the cave and named as Gammarus kuntisp. nov. Some of the characteristic features of the newly-identified species can be listed as “Medium-large size; smooth body, well-developed and reniform eyes; non-prolonged extremities; antennal gland cone is straight and long; second antenna with setose peduncular and flagellar segments; medial palmar spine present; posterior margin of pereopod 3 densely setose; anterior margins of pereopods 6 and 7 armed with spines only; epimeral plates not pointed”. Although the mentioned features are generally seen in epigean species, the members of this species were sampled from the dark zone of the Fakıllı Cave. The partial sequences of the COI (573 bp) and 28S (914 bp) genes of the newly-described species, Gammarus kuntisp. nov., were generated. The pairwise genetic distances between the new species, Gammarus kunti sp. nov. and other species ranged from a minimum of 16.23% (G. tumaf) to a maximum of 28.27% (G. roeselii) for the COI gene and a minimum of 0.88% (G. tumaf) to a maximum of 6.81% (G. balcanicus) for the 28S gene. Phylogenies generated by the NJ and ML methods, based on the combined data, assigned the new species as an independent lineage with high support values. In addition, the ASAP method identified the new species as a single MOTU independent of other species. G. tumaf and G. baysali are the sister taxa of G. kuntisp. nov. Detailed descriptions and drawings of the extremities of the male holotype and the female allotype are given and the morphology of the newly-identified species is compared with its relatives.

Host hydrocarbons protect symbiont transmission from a radical host defense

Symbioses with microbes play a pivotal role in the evolutionary success of insects, and can lead to intimate host-symbiont associations. However, how the host maintains a stable symbiosis with its beneficial partners while keeping antagonistic microbes in check remains incompletely understood. Here, we uncover a mechanism by which a host protects its symbiont from the host's own broad-range antimicrobial defense during transmission. Beewolves, a group of solitary digger wasps (Hymenoptera: Crabronidae), provide their brood cells with symbiotic Streptomyces bacteria that are later transferred to the cocoon and protect the offspring from opportunistic pathogens by producing antibiotics. In the brood cell, however, the symbiont-containing secretion is exposed to a toxic burst of nitric oxide (NO) released by the beewolf egg, which effectively kills antagonistic microorganisms. How the symbiont survives this lethal NO burst remained unknown. Here, we report that upon NO exposure in vitro, the symbionts mount a global stress response, but this is insufficient to ensure survival at brood cell-level NO concentrations. Instead, in vivo bioassays demonstrate that the host's antennal gland secretion (AGS) surrounding the symbionts in the brood cell provides an effective diffusion barrier against NO. This physicochemical protection can be reconstituted in vitro by beewolf hydrocarbon extracts and synthetic hydrocarbons, indicating that the host-derived long-chain alkenes and alkanes in the AGS are responsible for shielding the symbionts from NO. Our results reveal how host adaptations can protect a symbiont from host-generated oxidative and nitrosative stress during transmission, thereby efficiently balancing pathogen defense and mutualism maintenance.

Identifying sex-differential gene expression in the antennal gland of the swimming crab by transcriptomic analysis

The antennal glands (AnGs) are recognized as an important organ that functions in ion regulation and excretion in decapods. Previously, many studies had explored this organ at the biochemical, physiological, and ultrastructural levels but had few molecular resources. In this study, the transcriptomes of the male and female AnGs of Portunus trituberculatus were sequenced using RNA sequencing (RNA-Seq) technology. Genes involved in osmoregulation and organic/inorganic solute transport were identified. This suggests that AnGs might be involved in these physiological functions as versatile organs. A total of 469 differentially expressed genes (DEGs) were further identified between male and female transcriptomes and found to be male-biased. Enrichment analysis showed that females were enriched in amino acid metabolism and males were enriched in nucleic acid metabolism. These results suggested differences in possible metabolic patterns between males and females. Furthermore, two transcription factors related to reproduction, namely AF4/FMR2 family members Lilli (Lilli) and Virilizer (Vir), were identified in DEGs. Lilli was found to be specifically expressed in the male AnGs, whereas Vir showed high expression levels in the female AnGs. The expression of up-regulated metabolism and sexual development-related genes in three males and six females was verified by qRT-PCR and the pattern was found to be consistent with the transcriptome expression pattern. Our results suggest that although the AnG is a unified somatic tissue composed of individual cells, it still demonstrates distinct sex-specific expression patterns. These results provide foundational knowledge of the function and differences between male and female AnGs in P. trituberculatus.

Benzoic Acid Inclusion Effects on Health Status and Growth Performance of Juvenile Pacific White Shrimp Penaeus vannamei

AbstractOrganic acids and their salts are used as additives in aquafeeds, as their participation in some metabolic pathways and energy generation in organisms increases the nutritional value. Benzoic acid (BA; C6H5‐COOH), a white crystalline substance with a chemical structure that contains carboxylic acid attached to a benzene ring, is naturally found in plant and animal tissues. It is one of the most used organic acids due to its antifungal properties, and it acts as a preservative for a variety of human consumption products. Additionally, it demonstrates health and growth benefits when included in feeds by modifying the energy metabolism, microbiota, and intestinal tract morphology. Information on BA inclusion in aquatic food is rare, and such information for Pacific white shrimp Penaeus vannamei is nonexistent. This study evaluated the effects on growth, survival, feed conversion ratio, energy content, and health status (through lipid quantification in the hepatopancreas, histological examination, and salinity stress tests) of Pacific white shrimp juveniles by including BA at different concentrations (0.0, 1.0, 1.5, and 2.0 g/kg) in a commercial feed. Diets with BA inclusion produced significant differences in the hepatosomatic index, hepatopancreas lipid content, and histopathological index value (reflecting damage to the hepatopancreas and antennal gland atrophy) at all BA concentrations. The lowest survival obtained was 94.44% at a BA concentration of 1.0 g/kg. The feed conversion ratio was in accordance with the range established by the Food and Agriculture Organization of the United Nations except for values obtained at 0 g/kg. Calorimetric analysis and BA detection in muscle revealed no significant differences between concentrations. Due to the results obtained in this study, the evaluated inclusions of BA are not recommended for use in juvenile Pacific white shrimp diets.

Characterization of 3 different types of aquaporins in Carcinus maenas and their potential role in osmoregulation

Intertidal crustaceans like Carcinus maenas shift between an osmoconforming and osmoregulating state when inhabiting full-strength seawater and dilute environments, respectively. While the bodily fluids and environment of marine osmoconformers are approximately isosmotic, osmoregulating crabs inhabiting dilute environments maintain their bodily fluid osmolality above that of their environment by actively absorbing and retaining osmolytes (e.g., Na+, Cl−, urea) while eliminating excess water. Few studies have investigated the role of aquaporins (AQPs) in the osmoregulatory organs of crustaceans, especially within brachyuran species. In the current study, three different aquaporins were identified within a transcriptome of C. maenas, including a classical AQP (CmAQP1), an aquaglyceroporin (CmGLP1), and a big-brain protein (CmBIB1), all of which are expressed in the gills and the antennal glands. Functional expression of these aquaporins confirmed water transport capabilities for CmAQP1, CmGLP1, but not for CmBIB1, while CmGLP1 also transported urea. Higher relative CmAQP1 mRNA expression within tissues of osmoconforming crabs suggests the apical/sub-apically localized channel attenuates osmotic gradients created by non-osmoregulatory processes while its downregulation in dilute media reduces the water permeability of tissues to facilitate osmoregulation. Although hemolymph urea concentrations rose upon exposure to brackish water, urea was not detected in the final urine. Due to its urea-transport capabilities, CmGLP1 is hypothesized to be involved in a urea retention mechanism believed to be involved in the production of diluted urine. Overall, these results suggest that AQPs are involved in osmoregulation and provide a basis for future mechanistic studies investigating the role of AQPs in volume regulation in crustaceans.

A Multi-Species Comparison and Evolutionary Perspectives on Ion Regulation in the Antennal Gland of Brachyurans.

Brachyurans inhabit a variety of habitats and have evolved diverse osmoregulatory patterns. Gills, antennal glands and a lung-like structure are important organs of crabs that maintain their homeostasis in different habitats. Species use different processes to regulate ions in the antennal gland, especially those with high terrestriality such as Grapsoidea and Ocypodoidea. Our phylogenetic generalized least square (PGLS) result also suggested that there is a correlation between antennal gland NKA activity and urine-hemolymph ratio for Na+ concentration in hypo-osmotic environments among crabs. Species with higher antennal gland NKA activity showed a lower urine-hemolymph ratio for Na+ concentration under hypo-osmotic stress. These phenomenon may correlate to the structural and functional differences in gills and lung-like structure among crabs. However, a limited number of studies have focused on the structural and functional differences in the antennal gland among brachyurans. Integrative and systemic methods like next generation sequencing and proteomics method can be useful for investigating the differences in multi-gene expression and sequences among species. These perspectives can be combined to further elucidate the phylogenetic history of crab antennal glands.

Comparative Morphology of the Symbiont Cultivation Glands in the Antennae of Female Digger Wasps of the Genus Philanthus (Hymenoptera: Crabronidae).

Females of the solitary digger wasp tribe Philanthini, called the beewolves (Hymenoptera, Crabronidae), cultivate strains of symbiotic bacteria that belong to the genus Streptomyces in unique and highly specialized glands in their antennae. The glands consist of large reservoirs that are surrounded by numerous gland cell complexes (class III). The symbionts are cultivated inside the reservoirs and are probably provisioned with nutrients secreted from the surrounding glands and/or sequestered from the hemolymph. The wasp female delivers the bacteria into the subterranean brood cell prior to oviposition. Fully grown larvae take up the bacteria and apply them to their cocoon. There the bacteria produce several antibiotics that protect the wasp offspring against fungus infestation. Hitherto Streptomyces bacteria were detected in the antennae of 38 species of the Philanthini. However, a detailed morphological analysis of the antennal glands is only available for a few species. In order to shed light on the evolutionary history of the association between beewolf wasps and bacteria, we investigated the morphology of the antennal glands of another 14 Philanthus species from the Palearctic, Paleotropic, and Nearctic. We generated 3D-models of the glands based on serial semithin sections and/or micro-CT (μCT). Despite broad similarities in number and structure of antennal glands, the results revealed interspecific differences with regard to overall shape, complexity, and relative size of the reservoirs as well as the number of the surrounding gland cell units. Mapping the morphology of all species studied so far on the phylogeny (that parallels geographical distribution) revealed that related species share similarities in gland morphology, but there are notable differences between lineages. In particular, compared to the North American species the European and African species possess more complex gland structures with a higher number of gland cells. We discuss morphological, ecological, and physiological aspects and provide scenarios for the evolution of the antennal glands of the Philanthini as symbiont cultivation organs.

Metabolic changes in antennal glands of Caribbean spiny lobsters Panulirus argus infected by Panulirus argus virus 1 (PaV1).

Panulirus argus virus 1 (PaV1) (Family Mininucleoviridae) causes chronic and systemic infection in wild juvenile spiny lobsters Panulirus argus (Latreille, 1804), ending in death by starvation and metabolic wasting. In marine decapods, the antennal gland is involved in osmoregulation and excretion. In this compact organ, fluid is filtered from the hemolymph, and ions are reabsorbed to produce a hypotonic urine. Although PaV1 is released with the urine in infected individuals, little is known regarding the metabolic effect of PaV1 in the antennal gland. The objective of this study was to perform a comparative evaluation of the metabolic profile of the antennal gland of clinically PaV1-infected lobsters versus those with no clinical signs of infection, using proton nuclear magnetic resonance analysis. Overall, 48 compounds were identified, and the most represented metabolites were those involved in carbohydrate, amino acid, energy, and nucleotide metabolism. Most of the metabolites that were down-regulated in the infected group were essential and non-essential amino acids. Some metabolites involved in the urea cycle and carbohydrate metabolism were also altered. This study represents a first approach to the metabolic evaluation of the antennal gland. We broadly discuss alterations in the content of several proteinogenic and non-proteinogenic amino acids and other key metabolites involved in energetic and nucleotide metabolism.

The Osmoregulatory Function of Antennal Glands of the Green Mud Crab (Scylla Paramamosain)

The Osmoregulatory Function of Antennal Glands of the Green Mud Crab (Scylla Paramamosain)

Microanatomy of the antennal glands and their Na+/K+-ATPase activity in three true crab species (Brachyura), Portunus pelagicus (Linnaeus, 1758) (Portunidae), Macrophthalmus dentipes Lucas in Guérin, 1836 (Macrophthalmidae) and Eriocheir hepuensis Dai, 1991 (Varunidae)

Abstract There are differences between various crab species in the function and structure of organs involved in ionic and osmotic regulation processes. The antennal glands together constitute one of the most important organs involved in the osmoregulation in crabs. The present investigation aimed to study the tissue structure of the antennal glands and their Na+/K+-ATPase (NKA) pump activity in three true crab species from three different habitats, including the marine (Portunus pelagicus (Linnaeus, 1758)), the estuarine (Macrophthalmus dentipes Lucas in Guérin, 1836) and the freshwater habitat (Eriocheir hepuensis Dai, 1991). In this regard, the tissue structure of the antennal glands and the activity of the Na+/K+-ATPase (NKA) pump were assessed in these three selected species. The results showed that the antennal glands in all studied species consisted of two anterior parts and a posterior part. The anterior parts are composed of the proximal tubular region (PT) and the distal tubular region (DT). The PT and DT parts comprised the coelomosac and labyrinths in the anterior portion, and the bladder located in the posterior portion. However, despite the similarity in the general tissue structure of the antennal gland in marine, estuarine and freshwater crab species, some structural differences were observed between those species. Labyrinth cells, coelomosac podocytes, and bladder cells in the estuarine crab M. dentipes contained large vacuoles especially on the top (i.e., near the lumen) of the cells. The highest amount of NKA pump activity was measured in the antennal glands of M. dentipes (). The NKA pump plays a more important role in the estuary and seawater adaptation of crabs, but freshwater species are not highly dependent on the NKA enzyme for osmoregulation.

QUANTIFYING BILATERAL INFECTION PATTERNS IN THE TREMATODE ALLOGLOSSIDIUM RENALE.

Within-host distributions of parasites can have relevance to parasite competition, parasite mating, transmission, and host health. We examined the within-host distribution of the adult trematode Alloglossidium renale infecting the paired antennal glands of grass shrimp. There are 4 possible parasite distributions for infections of paired organs: random, uniform, biased aggregation to 1 particular organ (e.g., left vs. right), or inconsistently biased (aggregated, but does not favor 1 side). Previous work has shown that morphological asymmetries in hosts can lead to biased infections of paired organs. Apparent symmetry between the antennal glands of grass shrimp leads to the prediction that there would be no bias for 1 particular organ. However, an alternative prediction stems from the fact that A. renale is hermaphroditic: aggregation between glands would increase outcrossing opportunities and thus, avoid inbreeding via self-mating. Existing methods to test for an overall pattern did not apply to the A. renale system because of low-intensity infections as well as many 0 values for abundance per unit of the antennal gland. Hence, we used Monte Carlo simulations to determine if the observed overall patterns differed from those expected by randomly allocating parasites into groups of 2. We found that in 3 of 4 data sets, A. renale infections did not deviate from random distributions. The fourth data set had a more uniform pattern than expected by chance. As there was no aggregation between glands and the proportion of worms in single gland infections did not differ from that expected by chance alone, we found no evidence of inbreeding avoidance as might be manifested via a within-host distribution. Given the large proportion of worms in single infections, we predict as a major evolutionary outcome that populations of A. renale will be largely inbred.

Changes in calcium content, histopathology and calreticulin expression in the juvenile Chinese mitten crab Eriocheir sinensis under different salinity conditions

The growth, development and reproduction of Eriocheir sinensis are influenced by salinity changes. In order to study the response of juvenile Chinese mitten crab to salinity, the crabs were exposed to 5 salinity conditions (0, 3, 6, 12 and 18‰) for 48 h. The calcium content was detected in the hepatopancreas, posterior gill, intestine, haemolymph, antennal gland and carapace. The hepatopancreas, posterior gills and intestines were selected for subsequent experiments, and their histopathological changes and calreticulin (CRT) gene expression levels were evaluated. The calcium content of hepatopancreas and intestine was highest at 6‰ salinity and that of gill at 3‰ salinity. With the increase in salinity, the degree of damage was more obvious in tissues. Compared with the control group (0‰), CRT expression in the gills decreased significantly. However, in the hepatopancreas and intestine, CRT expression increased at 3‰ salinity and decreased at 3–6‰ salinity. Overall, our results showed that juvenile E. sinensis could survive short-term exposure to saline water within a certain salinity range (<18‰). However, a high concentration of saline water exposed the juvenile crab to excessive stress, resulting in serious tissue damage, which was not conducive to the long-term survival of juvenile crabs.

Effect of oxytetracycline on the biosafety, gut microbial diversity, immune gene expression and withdrawal period in Pacific whiteleg shrimp, Penaeus vannamei

Aquaculture disease management has been one of the primary focuses of improved production and profitability. Oxytetracycline (OTC) is a broad-spectrum antibiotic approved for therapeutic applications in aquaculture. Though maximum residue levels, safety and withdrawal period is established for finfish and tiger shrimp (Penaeus monodon), similar information is scanty on Pacific whiteleg shrimp, Penaeus vannamei, farmed under the tropical climates of South Asia. In this study, P. vannamei was administered with OTC for 14 days at different doses, 2.25, 4.5, 9.0, 18.0 g kg−1. OTC administration at 4.5 g kg−1 feed dose or less caused no significant impact on the growth or survival of the shrimp during the study period. The residual concentration in shrimp muscle declined below 0.1 ppm in 96 h post-treatment and was undetected at 120 h post-treatment. There were no significant histological changes in hepatopancreas, antennal gland and hematopoietic tissue at the therapeutic dose (4.5 g kg−1 feed), while dose-dependent toxic changes were observed in the lymphoid organ. Therapeutic dose of OTC administration did not influence the relative expression of crustin, penaeidin, lysozyme, pro-phenol oxidase and superoxide dismutase. Dose-dependent reduction in gut microbial diversity was observed as revealed by denaturing gradient gel electrophoresis (DGGE). In all the treatments, however, the microbial diversity was restored 14 days post-cessation of OTC administration. The observed 50% inhibition of phytoplankton Arthrospira platensis and Chlorella vulgaris (68.17 and 78.18 mg L−1 OTC, respectively) and 50% immobilisation of copepods and rotifers (173.89 and 257.16 mg L−1 OTC, respectively) was significantly higher than the therapeutic dose. Oxygen consumption, ammonia excretion, feeding rate and ingestion rate of copepod was significantly reduced at and above 187.5 mg L−1 OTC, while rotifers showed no impact of OTC up to 750 mg L−1, which is higher than the recommended therapeutic dose. The study has indicated that 96 h of withdrawal period may be required for safe use of OTC at 4.5 g kg−1 feed for 14 days in Indian P. vannamei farming.

Courtship with two spoons-Anatomy and presumed function of the bizarre antennae of Cardiocondyla zoserka ant males.

Mating in ants often occurs on the wing during nuptial flights or on the ground when scattered female sexuals attract males by pheromones. In both scenarios, there is little opportunity for males to engage in prolonged aggressive competition or elaborate courtship displays. Male morphology is therefore adapted to locating female sexuals and mating, and it lacks specific weapons or other traits associated with courtship. In contrast, sexuals of the ant genus Cardiocondyla typically mate in their natal nests. As a consequence, in many species winged males have been replaced by wingless fighter or territorial males, which kill or expel rival males with their strong mandibles and show complex mating behavior. However, no wingless males are known from Cardiocondyla zoserka from West Africa, and instead, winged males have evolved a bizarre secondary sexual trait: uniquely shaped antennae with spoon‐like tips that show heavily sculptured ventral surfaces with numerous invaginations. We here report on the courtship behavior of C. zoserka males and describe antennal glands with class 3 gland cells, which presumably secrete a close range sex pheromone. Antennal glands have not yet been found in males of other ant species, including a close relative of C. zoserka, suggesting that in ants with intranidal mating sexual selection can rapidly lead to highly divergent adaptations and the evolution of novel structures.