Impacts of acute hepatopancreatic necrosis disease on commercial shrimp aquaculture.

Acute hepatopancreatic necrosis disease (AHPND) has caused severe losses in farmed populations of marine shrimp Penaeus vannamei and P. monodon. The causative agents are unique strains of the bacteria Vibrio parahaemolyticus and related Vibrio species. The disease emerged in the People's Republic of China (China) and Vietnam in 2010 and spread throughout South-East Asia; it was later reported in countries in both North and South America. The disease has had significant economic impacts on the shrimp aquaculture industry. From 2010 to 2016, combined losses from China, Malaysia, Mexico, Thailand and Vietnam due primarily to outbreaks of AHPND, including losses at the farm gate and those resulting from a drop in feed sales and exports, were estimated at over US$ 44 billion. Other economic losses include those associated with processing facilities, decreased community revenues resulting from increased unemployment, financial investments, and the costs of implementing diagnostic and control measures. The reduced employment opportunities and increases in debt burden and investment risk have had sociological impacts. The responses to the disease have led to a gradual recovery of the shrimp industry in affected countries. These response efforts have included the implementation of changes in farming systems and management, including, among others, enhanced biosecurity and the use of AHPND-free and AHPND-resistant shrimp. This situation of losses and recovery illustrates the importance of having a multi-level response plan in place to prevent, or to reduce the risk of, outbreaks of disease.

La maladie de nécrose hépatopancréatique aiguë (AHPND, selon ses sigles en anglais) a occasionné des pertes importantes dans les élevages des espèces de crevettes marines Penaeus vannamei et P. monodon. La maladie est causée par des souches particulières de la bactérie Vibrio parahaemolyticus et d'autres espèces apparentées de Vibrio. Apparue en 2010 en République populaire de Chine et au Vietnam, la maladie s'est d'abord propagée dans toute l'Asie du SudEst avant d'être notifiée dans plusieurs pays d'Amérique du Nord et du Sud. Ses conséquences économiques sont très lourdes pour le secteur de la pénéiculture. On estime à plus de 44 milliards de dollars US les pertes cumulées enregistrées entre 2010 et 2016 par la Chine, la Malaisie, le Mexique, la Thaïlande et le Vietnam suite aux foyers d'AHPND (il s'agit aussi bien des pertes directes subies par les exploitations que de celles résultant de l'effondrement des ventes et des exportations d'aliments pour les élevages). D'autres pertes économiques sont associées aux établissements de transformation, aux pertes de revenus au sein des communautés locales par suite de l'augmentation du chômage, au déclin des investissements et aux coûts du diagnostic et des mesures de contrôle de la maladie. La réduction des perspectives d'emploi et l'augmentation du poids de la dette et des risques liés aux investissements ont affecté la société dans son ensemble. La mise en place de mesures appropriées a permis un redressement progressif du secteur de l'élevage de crevettes dans les pays atteints. Parmi ces mesures figurent les changements introduits dans les systèmes d'élevage et de gestion, en particulier l'amélioration de la biosécurité et l'utilisation de crevettes résistantes à la maladie ou indemnes. La situation décrite concernant ces pertes et ce redressement illustre l'importance de disposer d'un plan d'action à plusieurs niveaux afin de prévenir ou de réduire le risque de foyers.

La enfermedad de la necrosis hepatopancreática aguda (AHPND por sus siglas en inglés) ha infligido enormes pérdidas a las poblaciones de cultivo de camarones marinos de las especies Penaeus vannamei y P. monodon. Sus agentes etiológicos son determinadas cepas de las bacterias Vibrio parahaemolyticus y otras especies emparentadas del género Vibrio. La enfermedad apareció en 2010 en la República Popular de China y Vietnam y desde allí se propagó por todo el Sudeste asiático. Ulteriormente se notificó su presencia en Norteamérica y Sudamérica. La enfermedad ha tenido considerables consecuencias económicas para la industria camaronícola. El total de las pérdidas sufridas entre 2010 y 2016 por China, Malasia, México, Tailandia y Vietnam debidas principalmente a brotes de AHPND, sumando las sufridas en la explotación y las resultantes de la caída de las ventas de piensos y de las exportaciones, ascendieron según los cálculos a más de 44 000 millones de dólares estadounidenses. A estas pérdidas económicas se agregan otras, como las sufridas por las instalaciones de procesamiento, la caída de la renta comunitaria por el aumento del desempleo, la disminución de las inversiones financieras o los costos de aplicar medidas de diagnóstico y control. La reducción de las oportunidades de empleo y el aumento de la carga de la deuda y del riesgo de inversión han tenido también consecuencias sociológicas. Gracias a las medidas de lucha adoptadas, que trajeron consigo una serie de cambios en los sistemas acuícolas y en su gestión, entre ellos la mejora de la seguridad biológica y el uso de camarones no infectados y resistentes a la enfermedad, el sector camaronícola de los países afectados se ha ido recuperando gradualmente. Esta dinámica de pérdida y recuperación pone de manifiesto la importancia de tener instituido un plan de respuesta en múltiples eslabones para prevenir brotes de la enfermedad o reducir el riesgo de que se produzcan.

PirVP genes causing AHPND identified in a new Vibrio species (Vibrio punensis) within the commensal Orientalis clade.

Acute hepatopancreatic necrosis disease (AHPND) has extended rapidly, causing alarming shrimp mortalities. Initially, the only known causative agent was Vibrio parahaemolyticus carrying a plasmid coding for the mortal toxins PirVP. Recently, it has been found that the plasmid and hence the disease, could be transferred among members of the Harveyi clade. The current study performs a genomic characterization of an isolate capable of developing AHPND in shrimp. Mortality studies and molecular and histopathological analyses showed the infection capacity of the strain. Multilocus sequence analysis placed the bacteria as a member of the Orientalis clade, well known for containing commensal and even probiotic bacteria used in the shrimp industry. Further whole genome comparative analyses, including Vibrio species from the Orientalis clade, and phylogenomic metrics (TETRA, ANI and DDH) showed that the isolate belongs to a previously unidentified species, now named Vibrio punensis sp. nov. strain BA55. Our findings show that the gene transfer capacity of Vibrio species goes beyond the clade classification, demonstrating a new pathogenic capacity to a previously known commensal clade. The presence of these genes in a different Vibrio clade may contribute to the knowledge of the Vibrio pathogenesis and has major implications for the spread of emerging diseases.

Crude oil toxicity to fiddler crabs (Uca longisignalis and Uca panacea) from the northern Gulf of Mexico: Impacts on bioturbation, oxidative stress, and histology of the hepatopancreas.

The intensive drilling and extraction of fossil fuels in the Gulf of Mexico result in a considerable risk of oil spills impacting its coastal ecosystems. Impacts are more likely to be far-reaching if the oil affects ecosystem engineers like fiddler crabs, whose activities modify biogeochemical processes in the sediment. The present study investigated effects of oil on the fiddler crabs Uca longisignalis and Uca panacea, which are important as ecosystem engineers and as prey for a wide variety of species. The present study used mesocosms and microcosms to investigate the effects of crude oil on fiddler crab burrowing and to assess cellular and tissue damage by the oil. Fiddler crabs were exposed for periods of 5 or 10 d to oil concentrations up to 55 mg/cm2 on the sediment surface. Their burrowing was delayed, their burrows were smaller, and they transported less sediment in the presence of oil. The hepatopancreas had elevated levels of oxidative stress and a higher abundance of blister cells, which play a role in secretory processes. Interspecific differences were observed; most effects were strongest in U. panacea, though burrowing was more strongly affected in U. longisignalis. The present study demonstrates that crude oil is likely to impact fiddler crabs and many species that depend on them for their diet or for the ecological changes that result from their burrowing. Environ Toxicol Chem 2018;37:491-500. © 2017 SETAC.

Histopathological effects of chlorpyrifos on the gills, hepatopancreas and gonads of the freshwater crab Zilchiopsis collastinensis. Persistent effects after exposure.

Sublethal effects of the pesticide chlorpyrifos were evaluated in the crab Zilchiopsis collastinensis (Decapoda, Trichodactylidae). Crabs were exposed to high concentrations of chlorpyrifos at the beginning of the experiment and controlled dilution, under natural light and temperature conditions. A control and three concentrations (22.4, 41.25 and 61.4µg chlorpyrifos L-1) were evaluated in triplicate. Nine crabs per concentration and day were used. The gills, hepatopancreas and ovaries were sampled before pesticide exposure (day 0) and 8, 15 and 22 days later, when concentrations were diluted and below the detection limits. The histopathological effects and their variations in time were observed and quantified. In gills, hyperplasias were observed in several cases, mainly in crabs exposed to chlorpyrifos. The number of collapsed lamellae and the number of affected lamellae quickly increased in exposed crabs, as effects were observed on day 8 and remained until day 22. In hepatopancreas there was an increase in the number of F and B -cells and affected tubules, especially after 22 days of exposure (p<0.05). In ovaries, there were no effects on gonadosomatic indexes or oocyte volume, but there was a significant increase in the atretic oocyte proportion related to pesticide exposure (p<0.05). The histopathological effects on the gills, hepatopancreas and ovaries were observed after exposure and persist even after dilution, and might be related to earlier exposures. Thus, these histopathological effects might be used as pesticide biomarkers even after the pesticide is not detected by chemical methods.

Histopathological and biochemical evidence of hepatopancreatic toxicity caused by cadmium in white shrimp, Palaemonetes argentinus.

Cadmium (Cd) is one of the most common pollutants in the environment and induces a range of tissue changes or damages and organ dysfunction. The histopathological effects of Cd and lipid peroxidation (LPO) on hepatopancreas of the freshwater shrimp, Palaemonetes argentinus, were studied. Shrimp were obtained from two lagoons with contrasting environmental quality, De los Padres (LP, impacted site) and Nahuel Rucá (NR, reference site), and were exposed to 3.06 and 12.24µgCdL(-1) for 3, 7, 10 and 15 days. The health status of both populations was also evaluated by histological analysis of control individuals. After exposure, shrimp were transferred to clean water for 28 days to evaluate the recuperation capacity of hepatopancreas. Control shrimp from NR exhibited a normal hepatopancreas structure; unlike control shrimp from LP which showed several alterations. These results were attributed to the different environmental quality of lagoons. The exposure to Cd resulted in several alterations in the histological structure of the hepatopancreas of both populations. The observed alterations included haemocytic and connective infiltrations in the intertubular space, erosioned microvilli, ripple of basal lamina, atrophied epithelium and necrosis, however, the latter was only observed in shrimp from LP. The exposure also caused an increase of LPO levels in both populations. P. argentinus was able to repair the hepatopancreas structure from the damage caused by Cd, evidenced by the histopathological results and LPO levels. Obtained results are indicating that the histological analysis of the hepatopancreas proved to be a highly sensitive method for evaluating water quality, in both environmental and laboratory conditions.

Detection of acute hepatopancreatic necrosis disease (AHPND) in Mexico.

Acute hepatopancreatic necrosis disease (AHPND), which has also been referred to as early mortality syndrome (EMS), initially emerged as a destructive disease of cultured shrimp species in Asia in 2009. The pathogen associated with the disease, Vibrio parahaemolyticus, subsequently spread to the Western Hemisphere and emerged in Mexico in early 2013. The spread to the Western Hemisphere is a major concern to shrimp producers in the region. To date, the only peer-reviewed published method for determining whether mortalities are due to AHPND is through histological examination. A novel PCR detection method was employed to assess samples from Mexico in order to confirm the presence of the pathogen in this country. This manuscript details the detection methods used to confirm the presence of AHPND in Mexico. Both immersion and per os challenge studies were used to expose the Penaeus vannamei to the bacteria in order to induce the disease. Histological analysis confirmed AHPND status following the challenge studies. Also provided are the details of the molecular test by PCR that was used for screening candidate V. parahaemolyticus isolates. A rapid PCR assay for detection of AHPND may help with early detection and help prevent the spread of AHPND to other countries.

Morphology and histopathology of Calyptospora sp. (Apicomplexa: Calyptosporidae) in speckled peacock bass, Cichla temensis Humboldt, 1821 (Perciformes: Cichlidae), from the Marajó-Açu River, Marajó Island, Brazil.

Several species of coccidia are protozoan parasites that cause infection in a wide variety of animal groups. Calyptospora is an important genus of protozoan, which infests both freshwater and marine fish. The hepatopancreases of 150 speckled peacock bass captured on Marajó Island, Brazil were studied macro- and microscopically. Oocysts were found in 84 (56%) of the specimens in both the examination of the fresh material by compression and the analysis of histological sections stained with hematoxylin-eosin. Small, circular, homogeneous forms in negative contrast had a mean diameter of 21.2 µm, frequently with pyriform sporocysts, with a mean length of 9.2 µm and width of 3.1 µm, and a thin-walled capsule, were observed in both the hepatic and the pancreatic parenchyma, but were completely devoid of any inflammatory reaction. Calyptospora infections are documented for the first time in the Marajó-Açu River.

Hypoxia, reoxygenation and cytosolic manganese superoxide dismutase (cMnSOD) silencing in Litopenaeus vannamei: effects on cMnSOD transcripts, superoxide dismutase activity and superoxide anion production capacity.

The effects of silencing the mRNA of cytosolic manganese superoxide dismutase (cMnSOD), an enzyme involved in the antioxidant defense, were analyzed in Whiteleg shrimp, Litopenaeus vannamei adults. Shrimp were intramuscularly injected with long dsRNAs corresponding to the N-terminal portion of the cMnSOD and held under normoxic conditions for 24h. Another group of shrimp was exposed to hypoxia for 6h followed by reoxygenation for 1h. Shrimp injected with long dsRNAs had lower cMnSOD transcripts in gills and hepatopancreas. In the cMnSOD silenced shrimp, superoxide dismutase (SOD) activity decreased in gills but not in hepatopancreas. Shrimp subjected to hypoxia had lower cMnSOD transcripts and SOD activity in gills and hepatopancreas; the production of superoxide anion (O2*-) by hemocytes was also lower in this group. Reoxygenation reverted the effect of hypoxia increasing the levels of cMnSOD transcripts, SOD activity and the production of O2*-. These results suggest that cMnSOD contributes significantly to the SOD activity in gills and hepatopancreas and indicate its importance in the redox system regulation for L. vannamei.

The effect of crude ethanol extract and fractions of Hyptidendron canum (Pohl ex Benth.) Harley on the hepatopancreas of Oreochromis niloticus L.

Hyptidendron canum (Pohl ex Benth.) Harley is a native tree of the Brazilian Savannah. The fish Oreochromis niloticus L. was used as an experimental model to determine the bioactivity of the crude ethanol extract as well as ethyl acetate, hexanic and chloroform fractions obtained from its leaves. The plant ethanol extract and fractions were administered to the fish orally with their feed. Twenty four hours later, the fish were sacrificed and their livers dissected, fixed in neutral formalin, embedded in paraffin and sectioned. Histological analyses were performed using Masson's trichrome and Haematoxylin-Eosin. Histochemical studies were performed using Feulgen, PAS (Periodic Acid Schiff) and PAS + salivary amylase and Sudan IV stain. The qualitative analysis of the material showed that both the crude ethanol extract and the fractions from H. canum induced vasoactive activity, causing vasodilation and vascular congestion, and the hexanic fraction also caused an apparent proliferation of capillaries. Hepatopancreas toxicity was evident through inflammatory processes. Pancreatic (chloroform fraction) and hepatic alterations, hemorrhagic spots and necroses were observed in fish treated with-ethanol extract and fractions. This study is the first description of the biologic action of the crude ethanol extract and the hexane, ethyl acetate and chloroform fractions in fish.

Bioactivity of crude ethanol extract and fractions of Eugenia uniflora (Myrtaceae) in the hepatopancreas of Oreochromis niloticus L.

This study evaluates the bioactivity of the crude ethanol extract and ethyl acetate, hexane and chloroform fractions obtained from Eugenia uniflora leaves using the hepatopancreas of Oreochromis niloticus L. as an experimental model. The ethanol extract and fractions were administered to the fish orally with their feed. Twenty-four hours later, the fish were sacrificed and their livers dissected, fixed in neutral formalin, embedded in paraffin and sectioned. Histological analyses were performed using Masson's trichrome and Haematoxylin-Eosin. Histochemical studies were performed using Feulgen, PAS (Periodic Acid Schiff) and PAS + salivary amylase and Sudan IV stain. The qualitative analysis of the material showed that the crude extract and the ethyl, chloroform and hexane fractions induced vasodilation, vascular congestion and toxicity due to the presence of eosinophilic granular cells, rodlet cells, some leukocytic infiltrate and rare focal necroses. The Nile tilapia proved to be a satisfactory model for screening plant products.

The effect of crude ethanol extract and fractions of Hyptidendron canum (Pohl ex Benth) Harley on the hepatopancreas of Oreochromis niloticus L

Hyptidendron canum (Pohl ex Benth.) Harley is a native tree of the Brazilian Savannah. The fish Oreochromis niloticus L. was used as an experimental model to determine the bioactivity of the crude ethanol extract as well as ethyl acetate, hexanic and chloroform fractions obtained from its leaves. The plant ethanol extract and fractions were administered to the fish orally with their feed. Twenty four hours later, the fish were sacrificed and their livers dissected, fixed in neutral formalin, embedded in paraffin and sectioned. Histological analyses were performed using Masson's trichrome and Haematoxylin-Eosin. Histochemical studies were performed using Feulgen, PAS (Periodic Acid Schiff) and PAS + salivary amylase and Sudan IV stain. The qualitative analysis of the material showed that both the crude ethanol extract and the fractions from H. canum induced vasoactive activity, causing vasodilation and vascular congestion, and the hexanic fraction also caused an apparent proliferation of capillaries. Hepatopancreas toxicity was evident through inflammatory processes. Pancreatic (chloroform fraction) and hepatic alterations, hemorrhagic spots and necroses were observed in fish treated with-ethanol extract and fractions. This study is the first description of the biologic action of the crude ethanol extract and the hexane, ethyl acetate and chloroform fractions in fish.

Bioactivity of crude ethanol extract and fractions of Eugenia uniflora (Myrtaceae) in the hepatopancreas of Oreochromis niloticus L

This study evaluates the bioactivity of the crude ethanol extract and ethyl acetate, hexane and chloroform fractions obtained from Eugenia uniflora leaves using the hepatopancreas of Oreochromis niloticus L. as an experimental model. The ethanol extract and fractions were administered to the fish orally with their feed. Twenty-four hours later, the fish were sacrificed and their livers dissected, fixed in neutral formalin, embedded in paraffin and sectioned. Histological analyses were performed using Masson's trichrome and Haematoxylin-Eosin. Histochemical studies were performed using Feulgen, PAS (Periodic Acid Schiff) and PAS + salivary amylase and Sudan IV stain. The qualitative analysis of the material showed that the crude extract and the ethyl, chloroform and hexane fractions induced vasodilation, vascular congestion and toxicity due to the presence of eosinophilic granular cells, rodlet cells, some leukocytic infiltrate and rare focal necroses. The Nile tilapia proved to be a satisfactory model for screening plant products.

Hepatopancreas alteration of the blue crab Callinectes sapidus by the rhizocephalan barnacle Loxothylacus texanus.

A histological study of the hepatopancreas of the blue crab Callinectes sapidus parasitized by the rhizocephalan barnacle Loxothylacus texanus was conducted to explore if the degree of development of the parasite's rootlet system was correlated to its maturation process as seen by external characters of its reproductive body or externa. Four types of crabs were examined: control, with virgin and mature externa, and scarred. A clear progression with an increase in number and size of the parasite's rootlets in the hosts' hepatopancreas can be seen. Although the hepatopancreatic tubules remain functional, the hepatopancreas appears as a loose structure, completely infiltrated with L. texanus rootlets, in advanced stages of the parasitism.

Functional morphology of the hepatopancreas of Palaemonetes argentinus (Crustacea: Decapoda): influence of environmental pollution

We analyzed the morphological and functional state of hepatopancreas in Palaemonetes argentinusfrom two environments with different pesticide concentrations. Los Padres lagoon (Argentina) is an area subjectedto contamination due to the slow exchange of water, the shallow depth and the input of contaminatedwater. Prawns living in this lagoon accumulate high amounts of organochlorine pesticides in their tissues. Hepatopancreas of prawns from Canal 5, an adjacent shallow stream where the amount of pesticides is belowtoxic levels, and from Los Padres lagoon were processed by standard histological techniques with light microscopyand transmission electronic microscopy. At Los Padres lagoon, we found important tissular alterations,such as intertubular infiltration of haemocytes and connective tissue, epithelial retraction in some tubules, anda folded basal lamina. Important necrotic desquamation, with cariolysis, cariorrexis and lack of cellular detailswere also observed. Numerous tubules presented an enlarged and irregular lumen with the epithelium atrophiedor completely absent. In general, the lesions were particularly located in the medullar region of the organ. Atthe ultrastructural level, R and F cells were the most damaged. Both cell types had nuclear retraction, chromatincondensation and cytoplasmic lysis. Some R cells also had dilated mitochondria and numerous lysosomes, and the basal cytoplasm was nearly completely lysed. The hepatopancreas of prawns from Canal 5 did not evidenceany alterations. The histopathological study of the hepatopancreas is a highly sensitive tool to evaluate thephysiological condition of prawns and water quality. Other environmental conditions were similar, so it can beassumed that pollutants were the main cause of organ deterioration.

Analizamos el estado morfológico y funcional del hepatopáncreas de Palaemonetes argentinus de dosambientes con diferentes concentraciones de plaguicidas. La laguna Los Padres (Argentina) es un área sujeta a contaminación debido al lento intercambio del agua, la escasa profundidad y el influjo de agua contaminada. Las gambas acumulan aquí grandes cantidades de plaguicidasorganoclorados en sus tejidos. Los hepatopáncreas de gambas del Canal 5 y de la laguna Los Padres fueronprocesados mediante técnicas histológicas estándar para microscopía óptica y electrónica de transmisión. Loshepatopáncreas de los individuos recolectados en Los Padres tenían alteraciones tisulares importantes, comoinfiltración intertubular de hemocitos y tejido conectivo, retracción epitelial en algunos túbulos y láminas basalesplegadas. También se observó descamación necrótica importante, con cariolisis, cariorrexis y falta de detallescelulares. Vimos muchos túbulos con un lúmen irregular y agrandado, con el epitelio atrofiado o completamenteausente. En general las lesiones se localizaron en la región medular del órgano. Ambos tipos celulares mostraronretracción del núcleo, condensación de la cromatina y ruptura del citoplasma. Algunas células R también teníanmitocondrias y numerosos lisosomas dilatados, y el citoplasma basal casi completamente desintegrado. Nohallamos alteraciones en los hepatopáncreas de gambas deCanal 5. El estudio histopatológico del hepatopáncreas es una herramienta muy apropiada para evaluar la condiciónfisiológica de las gambas y la calidad del agua. Por la semejanza de otras condiciones ambientales, suponemosque los contaminantes fueron la causa principal del deterioro de los órganos.

Effects of methyl parathion on Chasmagnathus granulatus hepatopancreas: protective role of sesamol.

The protective role of sesamol, an inhibitor of the mixed function oxygenase (MFO) system, against histopathological effects of methyl parathion in the hepatopancreas of the estuarine crab, Chasmagnathus granulatus, was studied. Exposure (72 h) to a sublethal dose (0.05 mg/kg/day; 10% of 72 h-LD50) of injected methyl parathion increased the percentage of damaged hepatopancreatic tubules. Presence of melanin-like deposits in the connective tissue between hepatopancreatic tubules was also observed. Antioxidant enzyme activities (catalase and glutathione S-transferase) and lipid peroxidation (LPO) levels were also increased in hepatopancreas of crabs injected with methyl parathion. Pretreatment with sesamol (0.85 mg/kg/day) significantly protected against all these effects. These findings suggest that the hepatopancreatic damages induced by methyl parathion are due to LPO of hepatopancreatocytes membranes, as a consequence of the oxidative stress generated after methyl parathion oxidative biotransformation mediated by the MFO system.

Toxicity of hexachlorobenzene and its transference from microalgae (Chlorella kessleri) to crabs (Chasmagnathus granulatus).

The aim of this work was to study the transference of hexachlorobenzene from a green alga (Chlorella kessleri) to an estuary crab (Chasmagnathus granulatus), and to analyze the toxic effects that the xenobiotic has on the latter. The effect of hexachlorobenzene uptake was evaluated measuring oxidative stress, Uroporphyrinogen decarboxylase activity and morphometric parameter alteration, and also performing a histological analysis of crab hepatopancreas. Results demonstrated that hexachlorobenzene enters the alga, is accumulated in it, and then transferred into the crab, causing a decrease in Uroporphyrinogen decarboxylase activity in both organisms. The high malondialdehyde levels detected in crab hepatopancreas after the toxic treatment suggested the existence of hexachlorobenzene-induced lipid peroxidation. Antioxidant defenses such as superoxide dismutase activity and reduced glutathione content fell below normal values on the fourth week of treatment. At the same time, the hepatosomatic index, used as a morphometric parameter, reduced 20% with respect to the control. The histological analysis revealed epithelium disorganization in hepatopancreas tubules, confirming the existence of structural damage caused by hexachlorobenzene.

Toxic effects of microcystins in the hepatopancreas of the estuarine crab Chasmagnathus granulatus (Decapoda, Grapsidae).

Microcystins are toxins produced by cyanobacteria, being toxic to aquatic fauna. It was evaluated alternative mechanisms of microcystins toxicity, including oxidative stress and histopathology in the hepatopancreas of the estuarine crab Chasmagnathus granulatus (Decapoda, Grapsidae). Microcystins was administered to crabs (MIC group) over 1 week, whereas the control (CTR group) received the saline from cyanobacteria culture medium. At day 7, catalase activity was higher in the MIC than in the CTR group, although a decrease of activity was verified in both groups with respect to time 0. Glutathione-S-transferase activity augmented in MIC with respect to CTR, suggesting a higher conjugation rate of the toxins with glutathione. No differences were detected in the superoxide dismutase activity. Lipid peroxidation remained stable in both groups. Histopathological analyses showed that the number of B cells decreased significantly in the CTR as a possible effect of starvation, while no significant change was observed in the MIC group. The hepatopancreas from the MIC group exhibited some necrotic tubules and melanin-like deposits. Overall, results showed that some enzymes of the antioxidant defense system were activated after microcystins exposure, this response being able to maintain lipid peroxidation levels, but insufficient to completely prevent histological damage.

Fecal samples as DNA source for the diagnosis of Necrotizing Hepatopancreatitis (NHP) in Penaeus vannamei broodstock.

Necrotizing Hepatopancreatitis (NHP) is a severe disease of cultivated penaeid shrimp caused by a pleomorphic, gram-negative, intracellular rickettsia-like bacterium. Current diagnostic methods for this disease are invasive, requiring dissection of the animal to perform histopathological analysis. In Colombia, NHP affects mainly broodstock, being a major cause of mortalities in maturation laboratories. In order to identify the presence of NHP without having to dissect the animal, we developed a PCR-based method using fecal samples as the DNA source. The DNA was extracted using a quick isolation protocol followed by amplification with primers specific for 16S ribosomal RNA gene sequences. To verify the sensitivity and specificity we analyzed samples from the same animal by PCR and in situ hybridization, and found 100% agreement. In addition, we amplified DNA extracted form paraffin blocks to confirm NHP diagnosis. PCR amplification from fecal samples and paraffin blocks yielded the expected 440 bp fragment. We conclude that PCR amplification from fecal samples is a valuable tool for the diagnosis of NHP in broodstock organisms, and that paraffin-fixed tissues can be used as a source of DNA for PCR amplification of NHP.