Hemocyte Count Research Articles

Ingesting microplastics or nanometals during development harms the tropical pollinator Partamona helleri (Apinae: Meliponini)

The disposal of plastics and metal-derived compounds results in the contamination of the environment with nano/microparticles, leading to the exposure of various organisms to these harmful particles. However, the impacts of these particles on pollinating insects, which provide relevant ecosystem services, are not well understood. The aim of this study was to assess the effects of microscopic particles on the tropical pollinator Partamona helleri (Apinae: Meliponini), specifically evaluating the toxicity of plastic microparticles (polystyrene – PS, and polyethylene terephthalate – PET) and nanoparticles of a metal oxide (titanium dioxide – TiO2) via larval ingestion by bees reared in vitro. The survival rate of P. helleri larvae was not affected by the ingestion of particles of PS (500 ng/bee), PET (500 ng/bee), or TiO2 (10 μg/bee) compared to the non-treated diet (control or diet without the particles). Adults derived from treated larvae had increased body weight compared to the control, and the walking behavior of adults was altered by the ingestion of particles. Adults that ingested PET or TiO2 as larvae tended to rest for a longer time and interact more with other bees than the control. Hemocyte counts also changed, with a shift in the proportion of plasmatocytes and prohemocytes in treated individuals. Our findings suggest that even levels considered low for honey bees of exposure to plastic microparticles or metal nanoparticles can harm the health and behavior of stingless bees.

Immunoregulation and Resistance to Aquatic Pathogens with Dietary Nucleotides in Pacific White Shrimp, Litopenaeus vannamei

Using vegetable protein sources as a replacement for fish meal (FM) in the diet of Pacific white shrimp (PWS) has a negative impact on their health. Acute hepatopancreatic necrosis disease (AHPND), caused by Vibrio parahaemolyticus, affects PWS and causes financial losses. Nucleotides modulate the immune response and could contribute to counteracting these issues. Our objective was to evaluate the effects of nucleotide supplementation on performance, immune response, and survival when challenged with V. parahaemolyticus, in PWS receiving a diet with FM partially replaced with vegetable protein sources. A feeding trial (1000 PWS; 56 days) and a challenge trial (600 PWS; 10 days) were performed using diets with different FM inclusion levels (26%, 23.4%, 22.1%, and 20.8%), with or without 0.1% nucleotides. A non-challenged, non-supplemented group was also used in the challenge trial. Adding nucleotides to diets with reduced FM allowed significantly better results in growth performance parameters and total hemocyte count (THC). In the challenge trial, compared to control, nucleotide supplementation led to significantly higher THC and survival rate 15 h post-challenge. In conclusion, adding nucleotides to PWS diets improves their immune response and resistance to aquatic pathogens, allowing FM to be replaced by vegetable protein sources without negatively affecting performance.

Dietary Astragalus polysaccharides improve the growth and innate immune response of giant freshwater prawn Macrobrachium rosenbergii: Insights from the brain-gut axis

Supplementation with Astragalus polysaccharides (APS) has beneficial effects on aquatic animals. Herein, we aimed to investigate the effects of different doses of APS on the growth, innate immune response, and brain-gut axis of Macrobrachium rosenbergii. The molecular weight and the monosaccharide composition of APS were analyzed. APS were added at concentration of 0 (control), 0.05, 0.10, 0.15, and 0.20 % in practical diets. Growth performance increased significantly under 0.05 to 0.20 % APS, with enhanced lipase and protease activities in intestinal tissues. Prawns receiving APS supplementation had significantly lower amounts of pathogenic intestinal bacteria (Vibrio and Aeromonas) and a markedly different microbial community structure compared with those of the control group. The fecal short chain fatty acid (SCFA) and neurotransmitters γ-aminobutyric acid contents increased in the brains of prawns receiving APS, which was potentially associated with increased Lactobacillus and Bacillus levels. Prawns receiving APS supplementation displayed a significantly enhanced immune function (such as total hemocyte count, total protein concentration, phenoloxidase activity, serum agglutination titer, and lysozyme activity) and improved disease resistance to Vibrio anguillarum compared those in the control group. Thus, dietary APS positively affected the gut-brain axis by altering the microbiota composition, increasing the fecal SCFA content, and enhancing prawn immunity.

Potential toxic effects of titanium oxide (TiO2) nanoparticles on the biological, biochemical, and histological aspects of the land snail Helix aspersa

Nanotechnology has come a long way in our lives. However, it maintains some negative effects on the environment. This study aims to use the land snail Helix aspersa as a bioindicator. Titanium dioxide nanoparticles (TiO2NPs) had been used at 70 and 140 µg/L for two weeks by the spraying method. The oxidative biomarkers, condition index (CI), DNA damage, hemocyte count, and phagocytic activity were estimated. The toxicity of TiO2NPs was determined (LC50 = 544 µg/L). The exposure to TiO2NPs caused a significant reduction of the activities of superoxide dismutase (SOD) and catalase (CAT) in the digestive gland of Helix aspersa (the activity of CAT was 3.4 ± 0.1 (P = 0.001), SOD was 11 ± 1 (P = 0.0002) at concentration 140 µg/L after two weeks). The activity of glutathione peroxidase (GPX) was (1.13 ± 0.01 µ/mg protein at 140 µg/L compared with controls (5.47 ± 0.01 µ/mg protein). The treatment caused DNA damage in the hemocytes (tail DNA % = 8.66 ± 0.02 and tail moment = 52.99 ± 0 at140 µg/L (P = 0.002)). In the digestive gland, both tail DNA % and tail moment increased (tail moment = 78.38 ± 0.08 compared with control = 2.29 ± 0.09 (P = 0.0001)). The total count of hemocytes significantly decreased after two weeks (the average number was 71 ± 1.5 compared with controls 79 ± 1.1 at 140 µg/L). Furthermore, TiO2NPs caused histological alterations in the digestive gland of Helix aspersa. It can be concluded that the Helix aspersa can be used as environmental pollution bioindicator. A comprehensive evaluation of toxic effects induced by TiO2NPs in vivo assays must be investigated.

The effect of white spot syndrome virus (WSSV) envelope protein VP28 on innate immunity and resistance to white spot syndrome virus in Cherax quadricarinatus

VP28 is the most abundant membrane protein of WSSV, and the recombinant protein VP28 (VP26 or VP24) was constructed for the immune protection experiment in this study. Crayfish were immunized by intramuscular injection of recombinant protein V28 (VP26 or VP24) at a dose of 2μg/g. The survival rate of crayfish immunized by VP28 showed a higher value than by VP26 or VP24 after WSSV challenge. Compared with the WSSV-positive control group, the VP28-immunized group could inhibit the replication of WSSV in crayfish, increasing the survival rate of crayfish to 66.67% after WSSV infection. The results of gene expression showed that VP28 treatment could enhance the expression of immune genes, mainly JAK and STAT genes. VP28 treatment also enhanced total hemocyte counts and enzyme activities including PO, SOD, and CAT in crayfish. VP28 treatment reduced the apoptosis of hemocytes in crayfish, as well as after WSSV infection. In conclusion, VP28 treatment can enhance the innate immunity of crayfish and has a significant effect on resistance to WSSV, and can be used as a preventive tool.

Shrimp MANF maintains hemocyte viability via interaction with a tyrosine kinase Abl.

Mesencephalic astrocyte-derived neurotrophic factor (MANF) is a highly conserved cell protective protein. In this study, we explored its functions in shrimp hemocytes. Our results indicated that LvMANF knockdown could cause a decrease in total hemocyte count (THC) and an increase in caspase3/7 activity. To further explore its working mechanism, transcriptomic analyses were performed with wild-type and LvMANF-knockdown hemocytes. Three upregulated genes from transcriptomic data, including FAS-associated factor 2, rho-associated protein kinase 1, and serine/threonine-protein kinase WNK4 were validated with qPCR. Further experiments showed that LvMANF knockdown and tyrosine kinase LvAbl knockdown could decrease tyrosine phosphorylation in shrimp hemocytes. In addition, the interaction between LvMANF and LvAbl was validated with immunoprecipitation. The knockdown of LvMANF would decrease ERK phosphorylation and increase LvAbl expression. Our results suggest intracellular LvMANF may maintain shrimp hemocyte viability by interacting with LvAbl.

Identification and profiling of long non-coding RNAs during molt cycle: An involvement of lnc1182 in the molt of white shrimp, Litopenaeus vannamei

Molt is an essential physiological process for crustacean growth. It is known to be controlled by factors such as hormones and steroids, but the role of long non-coding RNA (lncRNA), a potential regulatory molecule, has not been studied. In this study, we investigated the role of lncRNA in the molt process of white shrimp, Litopenaeus vannamei. The 9437 of 58,142 unigenes of de novo assembled transcriptome were identified as putative lncRNAs, and 768 lncRNAs were differentially expressed during molt cycles. Five differentially expressed lncRNAs were selected to determine their expressions in different tissues and molt stages. Most were expressed more in gills and epidermis than other tissues. Expressions of lnc1182 and lnc336 significantly decreased during premolt stages and increased at postmolt in gills and epidermis, while lnc179 expression increased during premolt stages. RNA interference was used to investigate the role of lnc1182 in the molt process. The molt of lnc1182 knockdown shrimp progressed to early premolt faster than shrimp injected with dsGFP, and also shortened molt duration. In addition, total sugar level in hemolymph was higher in the lnc1182 knockdown shrimp than the control shrimp, whereas gill Na+/K+ ATPase activity, total hemocyte count, and expression of molt-inhibiting hormone (MIH) were not changed. These suggested that lnc1182 is likely involved in the control of sugar metabolism during molting but not in ion exchange activity and MIH expression. Furthermore, lnc1182 and 20-HE probably regulated each other. RNA sequencing was used to investigate the set of genes associated with lnc1182. Fifty-five transcripts were differentially expressed in gills of lnc1182 knockdown shrimp. Expressions of candidate metabolism and immune-related genes were validated for their differential expressions in lnc1182 knockdown shrimp. Our work produced lncRNA profiles and indicated possible functions of lnc1182 in metabolism and immunity during the molt cycle in shrimp.

Susceptibility of Hyphantria cunea larvae to Beauveria bassiana under Cd Stress: An integrated study of innate immunity and energy metabolism

Biological control is widely used for integrated pest management. However, there are many abiotic factors that can affect the biocontrol efficiency. In this study, we investigated the susceptibility of Hyphantria cunea larvae to Beauveria bassiana under Cd stress, and the corresponding mechanism was analyzed around innate immunity and energy metabolism. The results showed that mortality of H. cunea larvae treated with Cd and B. bassiana was significantly higher than those treated with B. bassiana alone, and the combined lethal effect exhibited a synergistic effect. Compared with the single fungal treatment group, the total hemocyte count in the combined Cd and fungal treatment group decreased significantly, accompanied by a decrease in phagocytosis, encapsulation, and melanization activity. The expression levels of three phagocytosis-related genes, one encapsulation-promoting gene, and one melanization-regulating gene were significantly lower in the combined treatment group than those in the single fungal treatment group. Furthermore, pathogen recognition ability, signal transduction level, and immune effector expression level were weaker in the combined treatment group than those in the single fungal treatment group. The expression levels of 14 key metabolites and 7 key regulatory genes in glycolysis and tricarboxylic acid cycle pathways were significantly lower in the combined treatment group than those in the single fungal treatment group. Taken together, the weakness of innate immunity and energy metabolism in response to pathogen infection resulted in an increased susceptibility of H. cunea larvae to B. bassiana under Cd pre-exposure. Microbial insecticide is a preferred strategy for pest control in heavy metal-polluted areas. Availability of data and materialAll the data that support the findings of this study are available in the manuscript.

Immediate and delayed effects of a heatwave and Prorocentrum lima ((Ehrenberg) Stein 1878) bloom on the toxin accumulation, physiology, and survival of the oyster Magallana gigas (Thunberg, 1793)

Warming could facilitate the intensification of toxic algal blooms, two important stressors for marine organisms that are predicted to co-occur more frequently in the future. We investigated the immediate and delayed effects of a heatwave and a simulated bloom (3 × 106 cells L-1) of the diarrhetic shellfish toxin (DST)-producing benthic dinoflagellate Prorocentrum lima on the survival, physiology (oxygen consumption rate, condition index, immune parameters), and toxin accumulation in the Pacific rock oyster Magallana (Crassostrea) gigas. Oysters exposed to both stressors contained higher mean DST concentrations (mean ± 1 SE: 173.3 ± 19.78 μg kg−1 soft tissue) than those exposed to P. lima bloom alone (120.4 ± 20.90 μg kg−1) and exceeded the maximum permitted levels for human consumption. Exposure to individual stressors and their combination modified the physiology of M. gigas. Oysters exposed to heatwave alone had significantly higher oxygen consumption rates (0.7 ± 0.06 mg O2 h−1 g−1) than the control (0.3 ± 0.06 mg O2 h−1 g−1). However, this was not observed in oysters exposed to both heatwave and P. lima (0.5 ± 0.06 mg O2 h−1 g−1). This alteration of the metabolic response to warming in the presence of P. lima may affect the ability of rock oysters to adapt to environmental stressors (i.e., a heatwave) to ensure survival. Immunomodulation, through changes in total hemocyte count, was observed in oysters exposed to P. lima alone and in combination with warming. Individual stressors and their combination did not influence the condition index, but one mortality was recorded in oysters exposed to both stressors. The findings of this study highlight the vulnerability of rock oysters to the predicted increased frequency of heatwaves and toxic algal blooms, and the increased likelihood of shellfish containing higher than regulatory levels of DST in warming coasts.

Impact of Pb Toxicity on the Freshwater Pearl Mussel, Lamellidens marginalis: Growth Metrics, Hemocyto-Immunology, and Histological Alterations in Gill, Kidney, and Muscle Tissue

Pb is one of the most extensively used harmful heavy metals in Bangladesh, and its occurrence in waters affects aquatic organisms significantly. The tropical pearl mussel, Lamellidens marginalis, was exposed to different concentrations (T1 21.93 mgL−1, T2 43.86 mgL−1, and T3 87.72 mgL−1) of Pb(NO3)2 and was evaluated against a control C 0 mgL−1 of Pb(NO3)2, followed by a 96 h acute toxicity test. The LC50 value was recorded as 219.32 mgL−1. The physicochemical parameters were documented regularly for each treatment unit. The values of % SGR, shell weight, soft tissue wet weight, and weight gain remained statistically higher for the control group in comparison with the treatment. No mortality was noted for control units, while a gradually decreased survival rate was recorded for the different treatment groups. Fulton’s condition factor was recorded as highest in the control and lowest in the T3 unit, while the condition indices did not vary between the control and treatment groups. The hemocyte was accounted as maximum in the control and T1, while minimum in T2 and T3. The serum lysosomal parameters also followed a similar pattern, and a significantly low level of lysosomal membrane stability, and serum lysosome activity was noted for T3 and T2 units in comparison to the control group. The histology of the gill, kidney, and muscle was well structured in the control group, while distinct pathologies were observed in the gill, kidney, and muscle tissue of different treatment groups. The quantitative comparison revealed that the intensity of pathological alteration increased as the dosage of Pb increased. The current study, therefore, indicated that intrusion of Pb(NO3)2 in the living medium significantly alters growth performance and hemocyte counts, and chronic toxicity induces histomorphological abnormalities in vital organs.

Acidification and hypoxia drive physiological trade-offs in oysters and partial loss of nutrient cycling capacity in oyster holobiont

IntroductionReef building oysters provide vast ecological benefits and ecosystem services. A large part of their role in driving ecological processes is mediated by the microbial communities that are associated with the oysters; together forming the oyster holobiont. While changing environmental conditions are known to alter the physiological performance of oysters, it is unclear how multiple stressors may alter the ability of the oyster holobiont to maintain its functional role.MethodsHere, we exposed oysters to acidification and hypoxia to examine their physiological responses (molecular defense and immune response), changes in community structure of their associated microbial community, and changes in water nutrient concentrations to evaluate how acidification and hypoxia will alter the oyster holobiont’s ecological role.ResultsWe found clear physiological stress in oysters exposed to acidification, hypoxia, and their combination but low mortality. However, there were different physiological trade-offs in oysters exposed to acidification or hypoxia, and the combination of stressors incited greater physiological costs (i.e., >600% increase in protein damage and drastic decrease in haemocyte counts). The microbial communities differed depending on the environment, with microbial community structure partly readjusted based on the environmental conditions. Microbes also seemed to have lost some capacity in nutrient cycling under hypoxia and multi-stressor conditions (~50% less nitrification) but not acidification.DiscussionWe show that the microbiota associated to the oyster can be enriched differently under climate change depending on the type of environmental change that the oyster holobiont is exposed to. In addition, it may be the primary impacts to oyster physiology which then drives changes to the associated microbial community. Therefore, we suggest the oyster holobiont may lose some of its nutrient cycling properties under hypoxia and multi-stressor conditions although the oysters can regulate their physiological processes to maintain homeostasis on the short-term.

Ambient salinity affects silver nanoparticles (AgNPs) induced toxicity in the marine bivalve, the rock oyster, Saccostrea cucullata

Silver nanoparticles (AgNPs) penetrate water sources in a variety of ways. Although some studies have reported their toxicity experimentally, little is known on the interactive impacts of nanoparticles and environmental factors on sea organisms. We investigated the interactions of salinity changes and AgNPs exposure in the rock oyster, Saccostrea cucullata off the Oman sea. The exposure test was done by challenging the oysters with environmentally relevant concentrations of AgNPs (Low concentration: 10 µg/l, middle concentration: 20 µg/l, high concentration: 30 µg/l) at different salinities (Low salinity: 12 g/l, middle salinity: 24 g/l, high concentration: 36 g/l) for 30 days, and then the immune components in hemolymph [Total hemocyte concentration (THC), phagocytosis], antioxidant enzyme activities [superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), glutathione S-transferase (GST)] in digestive gland, oxidative stress index (MDA: malondialdehyde) and immune-related gene expressions (pro-Cathepsin L, Rho protein, Ficolin, LPS/−1,3-glucan protein, DOCK 180 protein, Lysozyme, Defensin) in hemocyte were measured. Exposure of oysters to 20–30 µg/l AgNPs at 24 and 36 g/l salinities significantly increased hemocyte counts, phagocytosis, antioxidant enzyme activities and also up-regulated immune-related gene the expressions compared to 10 µg/l AgNPs treatment (P < 0.05). At salinity, 12 g/l, the exposure of the oysters to 10–30 µg/l AgNPs significantly decreased hemocyte counts, phagocytosis, antioxidant enzymes, and down-regulated the immune-related gene expressions (P < 0.05). In same salinity, GST activity showed significant decreases in the oysters exposed to 20–30 µg/l AgNPs (P < 0.05). The oxidative stress index (MDA) significantly elevated at salinity of 12 g/l (for oysters exposed to 10–30 µg/l AgNPs) and 24–36 g/l (for oysters exposed to 20–30 µg/l AgNPs). The results of the present study showed that a decrease in water salinity with can intensify toxicity in the rock oyster.

Application of a nitrifying bacterium Pseudomonas sp. HIB_D to reduce nitrogen waste in the Litopenaeus vannamei cultivation environment.

Pseudomonas sp. HIB_D has been observed to have nitrification activity that can oxidize ammonia/ammonium to nitrite and nitrate. This bacterium was isolated from the aquaculture environment with Ancol Indonesian marine water sources. Pseudomonas sp. HIB_D was applied in the rearing water of Litopenaeus vannamei to decrease nitrogen pollution levels and support a sustainable cultivation environment. This present study was set up using a completely randomized design with four treatments and three replications, i.e., control (without bacterial application) and bacterial application with a cell density of 107, 108, and 109CFUmL-1 in a volume of 100mL for 90 L seawater treatment. Thirty-six individuals of 15-day-old postlarvae L. vannamei (PL 15) (a density of 120 PL m-2) were used in this study for an 8-week rearing period. Based on the water quality analysis, the ammonia level under the bacterial applications was lower than in thecontrol after 8-week cultivation. Nitrate levels increased at week 6 and decreased at week 8, specifically in 109CFUmL-1 treatment. L. vannamei with 109CFUmL-1 bacterial application had the best production performance with a survival rate of 94.33 ± 2.78%, an absolute length gain of 10.59 ± 0.22cm, an absolute weight gain of 8.16 ± 0.33g, a specific growth rate of 14.12% ± 0.01% day-1, and a feed conversion ratio of 1.26 ± 0.03. The blood glucose level and total hemocyte count of shrimp under the bacterial application with a density of 109CFUmL-1 showed the highest level (30.71 ± 13.9mg dL-1) and the lowest level (15 × 106 cells mL-1), respectively. The results showed that all treatments performed better than the control, in which the 109CFUmL-1 bacterial application was the best treatment.

White feces syndrome in Penaeus vannamei is potentially an Enterocytozoon hepatopenaei (EHP) associated pathobiome origin of Vibrio spp

White feces syndrome (WFS) is a commercially important disease in Penaeus vannamei (whiteleg shrimp) farming. The aetiology beyond the white or golden white midgut with mediocre growth performance producing a floating mass of white fecal strings in WFS-affected shrimp farms remains uncharted. To give WFS a perception of pathobiome, healthy P. vannamei shrimps were subjected to an enteric microsporidian Enterocytozoon hepatopenaei (EHP) infection along with Vibrio harveyi and V. alginolyticus in different combinations. Immune responses in haemolymph (total haemocyte count (THC), prophenoloxidase activity (proPO), respiratory burst activity (RBA), superoxide dismutase activity (SOD) and catalase activity (CAT)), plasma biochemical changes (aspartate aminotransferase (AST), alanine aminotransferase (ALT) and alkaline phosphatase (ALP)) and digestive enzymes activity (alpha-amylase (AMY), lipase (LIP) and protease (PRO)) were assessed in the challenged shrimps at 5, 10 and 15 days post-infection (dpi). The microbial interactions between the EHP and Vibrio spp. have led to the formation of WFS in the challenged shrimps. The histological sections of the hepatopancreas revealed the presence of EHP along with colonized bacterial masses, leading to the formation of aggregated transformed microvilli (ATM) structures and increased sloughing of lipid vacuoles into the tubule lumen. A significantly decreased THC and increased proPO levels, dysregulated antioxidant system, prominent hepatic damage, reduced energy metabolism and higher lipid production were the key records supporting that EHP-associated WFS in P. vannamei is due to the pathobiome.

Biofloc system supplemented by Pseudoalteromonas piscicida 1Ub protects the Pacific white shrimp Penaeus vannamei from Vibrio parahaemolyticus infection

This study aimed to evaluate the supplementation of the probiotic Pseudoalteromonas piscicida 1Ub to the biofloc system as an ecofriendly strategy for protecting white shrimp (Penaeus vannamei) from Vibrio parahaemolyticus infection. Shrimp with an average body weight of (0.50 ± 0.09) g were reared in 30 glass jars with a working volume of 2.5 L at a density of 20 ind/L. Shrimp were reared for 5 d for each treatment, which included the biofloc system without and with 106 colony forming unit (CFU) per mL probiotic. The regular clear water system was used as control. All treatment groups were challenged with 103, 105, and 107 CFU/mL V. parahaemolyticus. For the negative control, shrimp were reared without V. parahaemolyticus. The results showed that the density of V. parahaemolyticus cocultured with P. piscicida 1Ub decreased and the density of V. parahaemolyticus in rearing water and shrimp body in the probiotic-treated group was lower than that in the control group (P < 0.05). The survival and immune response (total hemocyte count, phagocytic activity, respiratory burst, phenoloxidase, and superoxide dismutase) of shrimp in the probiotic group was higher than that in the positive control (P < 0.05). Moreover, supplementing the biofloc system with the probiotic could protect shrimp hepatopancreas from damage caused by V. parahaemolyticus, regardless of bacterial density. Thus, the supplementation of the probiotic P. piscicida 1Ub in the biofloc system could significantly protect and increase the resistance of shrimp to V. parahaemolyticus infection.

Differentiation between Bulinus truncatus and Bulinus hexaploidus by morphological characters, chromosomal study and compatibility with Schistosoma haematobium.

Schistosomiasis is a snail-born, neglected tropical disease (NTD) caused by blood flukes (trematode worms) of the genusSchistosoma. It is the second most socioeconomically devastating parasitic disease after malaria. Urogenital schistosomiasis is caused by Schistosoma haematobium which is transmitted by snail intermediate host of the genus Bulinus. This genus is a model system for the study of polyploidy in animals. This study aims to investigate ploidy levels existing among the Bulinus species and their compatibility with S. haematobium. The specimens were collected from two governorates in Egypt. Chromosomal preparation was made from gonad tissue (ovotestis). This study found two ploidy levels (tetraploid, n=36 and hexaploid, n=54) of B. truncatus/tropicus complex in Egypt. Tetraploid B. truncatus was found in El-Beheira governorate while-unexpectedly and for the first time in Egypt, the hexaploid population was found in Giza governorate. This identification focused on shell morphology, chromosomal count, and spermatozoa of each species. Afterward, all species were exposed to S. haematobium miracidia where B. hexaploidus snails were the only refractory species. The histopathological study showed early destruction and abnormal development of S. haematobium in B. hexaploidus tissues. In addition, the hematological investigation showed increasing in the total hemocyte count, the formation of vacuoles, several pseudopodia, and more dense granules in the hemocytes of infected B. hexaploidus snails. In conclusion, there were two types of snails one was refractory and the other was susceptible.

MORPHOLOGICAL AND QUANTITATIVE STUDIES OF THE HEMOCYTES IN NYMPH AND IMAGO OF BLAPTICA DUBIA (SERVILLE, 1839)

The purpose of the research is to analyze how the hemocyte counts and their morphological characteristics change during development of Blaptica dubia, especially during molting. And to suggest a possible scheme of the transformation of spherulocytes into crescent-cells.&#x0D; Materials and methods. Quantitative and morphological features of hemocytes of imago and nymphs of 1-4 stages of Blaptica dubia using light microscopy and AFM were researched. Cell, nucleus and granule sizes were determined using NIS-Elements and NOVA software. Cytochemical tests with neutral red and alcyan blue allowed us to identify different cell types.&#x0D; Results. This study presents a classification of hemocytes in Blaptica dubia, that based on morphological and cytochemical characteristics of cells. We analyzed the dynamics of the hemocytes counts in 6 nymphs (1-4 stages) and 6 imago. Special attention is given to the moment of molting. Seven hemocyte types were described and identified as: prohemocytes, plasmocytes, granulocytes, vermicytes, spherulocytes, coagulocytes and crescent-cells. The stem cells (prohemocytes) and spherulocytes were the predominant hemocytes. Prohemocytes counts decrease as a result of insect development, instead the relative number of plasmocytes and granulocytes increases. Spherulocytes counts could increase to 35% at the moment of leaving the ootheca and at the molting period. Due to the study of hemocytes diversity in different nymph stages, interim cells and possible ways of their transformations were identified. The unusual crescent-cells, that were unique for Gromphadorhina portentosa, had been found in Blaptica dubia. For the first time, the cytochemical test allowed to determine the origin of crescent-cells from spherulocytes.

Physiological and transcriptomic analyses reveal critical immune responses to hypoxia and sulfide in the haemolymph of clam Tegillarca granosa

Hypoxia and sulfide are inducing potential damage to aquatic organisms. However, the effects of hypoxia and sulfide on their immune systems and molecular mechanisms are not fully understood. In the present study, the clam Tegillarca granosa was exposed to hypoxia alone or in combination with sulfide (0.1, 0.5 mM) to investigate the physiological and transcriptomic responses in haemolymph. The IBR analysis revealed that moderate sulfide stimulated immune responses via increasing the total hemocyte counts, phagocytic activity, antibacterial activity, and antioxidant activity. The transcriptomic analysis revealed many critical signaling pathways (Toll and Imd, FoxO, NLR) and biological processes (antimicrobial/antibacterial peptide, interferon, interleukin, leukocyte, lymphocyte, mitophagy) involved in the immunostimulation. Our results would offer insights into the sulfide-tolerant molecular mechanisms in this species and provide a useful tool for assessing the integrated biological impacts of hypoxia and sulfide on shellfish.

Hemato-immunological parameters can be influenced by microalgae addition and fish feed supplementation in the integrated rearing of Pacific white shrimp and juvenile Nile tilapia using biofloc technology

This study evaluated the effects of microalgae (Scenedesmus obliquus) addition and fish feed supplementation on hemato-immunological parameters of Pacific white shrimp (Penaeus vannamei) and juvenile Nile tilapia (Oreochromis niloticus) cultured in integration using biofloc technology for 62 days. The experiment was designed as a factorial design with two factors, totaling four experimental groups: (1) No addition of microalgae and no addition of fish feed; (2) no addition of microalgae and addition of fish feed; (3) addition of microalgae and no addition of fish feed; and (4) addition of microalgae and addition of fish feed. In all groups shrimp were fed according to a feeding table. Shrimp (2.16 ± 0.01 g) and fish (1.53 ± 0.12 g) were cultured under densities of 400 shrimp m−3 and 522 fish m−3 in 800 L and 90 L (useful volumes) tanks maintained under water recirculation. Microalgae was added two times a week for a final concentration of 5 mg L−1 in the units, while a commercial fish feed was provided at the rate of 1% of the fish biomass daily. At the end of the trial, the fish and shrimp were sampled for the hemato-immunological analyses. As regards the shrimp, there was a significant effect of the interaction on total hemocyte count (p < 0.05), in which the highest value occurred in the group with microalgae/no fish feed addition. There was also a significant effect of microalgae addition on serum protein (p < 0.05), with the highest values occurring in the treatments that employed microalgae. In the case of the fish, the monocyte count was significantly affected by the fish feed addition (p < 0.05), the highest values being observed when the feed was provided. There were also significant effects of the interaction and of microalgae addition on mean corpuscular volume and mean corpuscular hemoglobin concentration (p < 0.05), with the highest values occurring in the group that received microalgae but no fish feed and in the groups that did not receive microalgae, respectively. Overall, these results suggest an enhanced immunological status for the shrimp due to increases in total hemocyte count and high serum protein concentration when microalgae was added and an improved nutritional status for the fish that received feed due to increased circulating monocytes, along with possible greater erythrocyte stability when microalgae was added.

Bioinsecticide spinosad poses multiple harmful effects on foragers of Apis mellifera.

There are multifactorial causes for the recent decline in bee populations, which has resulted in compromised pollination and reduced biodiversity. Bees are considered one of the most important non-target insects affected by insecticides used in crop production. In the present study, we investigated the effects of acute oral exposure to spinosad on the survival, food consumption, flight behavior, respiration rate, activity of detoxification enzymes, total antioxidant capacity (TAC), brain morphology, and hemocyte count of Apis mellifera foragers. We tested six different concentrations of spinosad for the first two analyses, followed by LC50 (7.7mg L-1) for other assays. Spinosad ingestion decreased survival and food consumption. Exposure to spinosad LC50 reduced flight capacity, respiration rate, and superoxide dismutase activity. Furthermore, this concentration increased glutathione S-transferase activity and the TAC of the brain. Notably, exposure to LC50 damaged mushroom bodies, reduced the total hemocyte count and granulocyte number, and increased the number of prohemocytes. These findings imply that the neurotoxin spinosad affects various crucial functions and tissues important for bee performance and that the toxic effects are complex and detrimental to individual homeostasis.