| 1. |
- Ekblom, Charlotta, et al.
(författare)
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Early Changes in Crayfish Hemocyte Proteins after Injection with a beta-1,3-glucan, Compared to Saline Injected and Naive Animals
- 2021
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Ingår i: International Journal of Molecular Sciences. - : MDPI. - 1661-6596 .- 1422-0067. ; 22:12
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Tidskriftsartikel (refereegranskat)abstract
- Early changes in hemocyte proteins in freshwater crayfish Pacifastacus leniusculus, in response to an injection with the fungal pattern recognition protein beta-1,3-glucan (laminarin) were investigated, as well as changes after saline (vehicle) injection and in naive animals. Injection of saline resulted in rapid recruitment of granular hemocytes from surrounding tissues, whereas laminarin injection on the other hand induced an initial dramatic drop of hemocytes. At six hours after injection, the hemocyte populations therefore were of different composition. The results show that mature granular hemocytes increase in number after saline injection as indicated by the high abundance of proteins present in granular cell vesicles, such as a vitelline membrane outer layer protein 1 homolog, mannose-binding lectin, masquerade, crustin 1 and serine protease homolog 1. After injection with the beta-1,3-glucan, only three proteins were enhanced in expression, in comparison with saline-injected animals and uninjected controls. All of them may be associated with immune responses, such as a new and previously undescribed Kazal proteinase inhibitor. One interesting observation was that the clotting protein was increased dramatically in most of the animals injected with laminarin. The number of significantly affected proteins was very few after a laminarin injection when compared to uninjected and saline-injected crayfish. This finding may demonstrate some problematic issues with gene and protein expression studies from other crustaceans receiving injections with pathogens or pattern recognition proteins. If no uninjected controls are included and no information about hemocyte count (total or differential) is given, expressions data for proteins or mRNAs are very difficult to properly interpret.
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| 2. |
- Hernandez-Perez, Ariadne, et al.
(författare)
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Environmental concentrations of sulfamethoxazole increase crayfish Pacifastacus leniusculus susceptibility to White Spot Syndrome Virus
- 2020
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Ingår i: Fish and Shellfish Immunology. - : ACADEMIC PRESS LTD- ELSEVIER SCIENCE LTD. - 1050-4648 .- 1095-9947. ; 102, s. 177-184
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Tidskriftsartikel (refereegranskat)abstract
- Antibiotics used for humans and livestock are emerging as pollutants in aquatic environments. However, little is known about their effect on aquatic organisms, especially in crustaceans. In the present study, the freshwater crayfish Pacifastacus leniusculus was exposed during 21 days to environmental concentrations of sulfamethoxazole (SMX) (100 ng/L and 1 mu g/L). Subsequently, the crayfish susceptibility to infection was evaluated by using White Spot Syndrome Virus (WSSV) challenge, a well-known crustacean pathogen. The median survival time of the infected crayfish exposed to 100 ng/L SMX was one day, whereas the control and the group exposed to 1 mu g/L SMX survived for two and three days, respectively. In order to elucidate the effect of SMX upon the crayfish immune response, new sets of crayfish were exposed to the same SMX treatments to evaluate mRNA levels of immune-related genes which are expressed and present in hemocytes and intestine, and to perform total and differential hemocyte counts. These results show a significant down-regulation of the antimicrobial peptide (AMP) Crustin 3 in hemocytes from the 100 ng/L SMX group, as well as a significant up-regulation of the AMP Crustin 1 in intestines from the 1 mu g/L SMX group. Semigranular and total hemocyte cell number were observed to be significantly lower after exposure to 100 ng/L SMX in comparison with the control group. The present study demonstrates that environmentally relevant SMX concentrations in the water at 100 ng/L led to an increased WSSV susceptibility, that may have been caused by a reduction of circulating hemocytes. Nevertheless, SMX concentrations of 1 mu g/L could marginally and for a few days have an immunostimulatory effect.
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| 3. |
- Hernandez-Perez, Ariadne, et al.
(författare)
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Gut microbiome alterations in the crustacean Pacifastacus leniusculus exposed to environmental concentrations of antibiotics and effects on susceptibility to bacteria challenges*
- 2022
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Ingår i: Developmental and Comparative Immunology. - : Elsevier. - 0145-305X .- 1879-0089. ; 126
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Tidskriftsartikel (refereegranskat)abstract
- Gut-associated microbiota in crustaceans are recognized as a key element for maintaining homeostasis and health in the animal. Since the richness of these microbial communities is strongly influenced by the local environment, especially in aquatic organisms, it is important to address to what extent environmental variations can affect these communities. In the present study, we used high-throughput 16S rRNA sequencing technology to study the composition of gut-associated microbiota of the crayfish Pacifastacus leniusculus after exposure to environmentally-relevant concentrations of an antibiotic, namely sulfamethoxazole. Also, we examined if alterations of microbiota caused by environmentally-relevant concentrations of this antibiotic affected the host susceptibility to bacterial diseases, including Vibrio species. As a result, we found high individual variability of bacterial abundance and composition in the intestinal microbiome of crayfish, in both antibiotic-exposed and antibiotic-free crayfish. However, an increase of chitinolytic bacteria including Vibrio spp. was detected in some animals exposed to the antibiotic. Moreover, when crayfish susceptibility to bacterial infections was tested, the antibiotic-exposed crayfish survived longer than the control crayfish group. This study represents the first approach for investigating the interplay between crayfish and intestinal bacteria during antibiotic-pollution scenarios. Results herein should be considered by scientists before planning experiments under laboratory conditions, especially to study environmental effects on aquatic animals' intestinal health and immune status.
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| 4. |
- Hernández-Pérez, Ariadne, et al.
(författare)
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Vibrio areninigrae as a pathogenic bacterium in a crustacean
- 2021
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Ingår i: Journal of Invertebrate Pathology. - : Elsevier. - 0022-2011 .- 1096-0805. ; 178
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Tidskriftsartikel (refereegranskat)abstract
- The occurrence of infectious diseases poses a significant threat to the aquaculture industry worldwide. Therefore, characterization of potentially harmful pathogens is one of the most important strategies to control disease outbreaks. In the present study, we investigated for the first time the pathogenicity of two Vibrio species, Vibrio metschnikovii, a foodborne pathogen that causes fatalities in humans, and Vibrio areninigrae, a bacteria isolated from black sand in Korea, using a crustacean model, the signal crayfish Pacifastacus leniusculus. Mortality challenges indicated that injection of V. metschnikovii (108 CFU/crayfish) has a mortality percentage of 22% in crayfish. In contrast, injection of P. leniusculus with 108 or 107 CFU of V. areninigrae resulted in 100% mortality within one and two days post-injection, respectively. V. areninigrae was successfully re-isolated from hepatopancreas of infected crayfish and caused 100% mortality when reinjected into new healthy crayfish. As a consequence of this infection, histopathological analysis revealed nodule formation in crayfish hepatopancreas, heart, and gills, as well as sloughed cells inside hepatopancreatic tubules and atrophy. Moreover, extracellular crude products (ECP’s) were obtained from V. areninigrae in order to investigate putative virulence factors. In vivo challenges with ECP’s caused >90% mortalities within the first 24 h. In vitro challenges with ECP’s of hemocytes induced cytotoxicity of hemocytes within the first hour of exposure. These findings represent the first report that V. areninigrae is a highly pathogenic bacterium that can cause disease in crustaceans. On the contrary, V. metschnikovii could not represent a threat for freshwater crayfish.
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| 5. |
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| 6. |
- Junkunlo, Kingkamon, et al.
(författare)
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A transcription factor glial cell missing (Gcm) in the freshwater crayfish Pacifastacus leniusculus
- 2020
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Ingår i: Developmental and Comparative Immunology. - : Elsevier BV. - 0145-305X .- 1879-0089. ; 113
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Tidskriftsartikel (refereegranskat)abstract
- The transcription factor glial cell missing, Gcm, is known to be an important protein in the determination of glial cell fate as well as embryonic plasmatocyte differentiation in Drosophila melanogaster. So far, no function for Gcm in crustaceans has been reported. In this study, we show the cDNA sequence of a Gcm homologue in the freshwater crayfish Pacifastacus leniusculus. The P. leniusculus Gcm transcript is expressed exclusively in brain and nervous tissue, and by in situ hybridization we show that the expression is restricted to a small number of large cells with morphology similar to neurosecretory cells. Furthermore, we show that the expression of Gcm coincides with the expression of a Repo homologue, that is induced in expression by Gcm in Drosophila. Moreover, the Gcm transcript is increased shortly and transiently after injection of cystamine, a substance that inhibits transglutaminase and also strongly affects the movement behavior of crayfish. This finding of Gcm transcripts in a subpopulation of brain cells in very low numbers may enable more detailed studies about Gcm in adult crustaceans.
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| 7. |
- Junkunlo, Kingkamon, et al.
(författare)
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Transglutaminase 1 and 2 are localized in different blood cells in the freshwater crayfish Pacifastacus leniusculus
- 2020
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Ingår i: Fish and Shellfish Immunology. - : ACADEMIC PRESS LTD- ELSEVIER SCIENCE LTD. - 1050-4648 .- 1095-9947. ; 104, s. 83-91
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Tidskriftsartikel (refereegranskat)abstract
- In the present study we show that hemocytes in the freshwater crayfish Pacifastacus leniusculus express two different transglutaminases. We describe the sequence of a previously unknown TGase (Pl_TGase1) and named this as Pl_TGase2 and compared this sequence with similar sequences from other crustaceans. The catalytic core domain is similar to the previously described TGase in P. leniusculus, but Pl_TGase2 has significant differences in the N-terminal and C-terminal domains. Further, we show conclusive evidences that these different transglutaminases are specific for different hemocyte types so that Pl_TGase1 is expressed in the hematopoietic tissue and in the cytoplasm of semigranular hemocytes, while Pl_TGase2 is expressed in vesicles in the granular hemocytes. By in situ hybridization we show that both Pl_TGase1 and Pl_TGase2 mRNA are present only in a subset of the respective hemocyte population. This observation indicates that there may be different subtypes of semigranular as well as granular hemocytes which may have different specific functions.
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| 8. |
- Sirikharin, Ratchanok, et al.
(författare)
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The N-terminal peptide generated after activation of prophenoloxidase affects crayfish hematopoiesis
- 2020
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Ingår i: Developmental and Comparative Immunology. - : ELSEVIER SCI LTD. - 0145-305X .- 1879-0089. ; 108
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Tidskriftsartikel (refereegranskat)abstract
- The circulating hemocytes of invertebrates are important mediators of immunity, and hemocyte homeostasis is of high importance for survival and health of crustaceans. The prophenoloxidase (proPO)-activating system is one of the most essential immune reactions, which can be activated by pattern recognition proteins from microorganisms. Activation of proPO by the proPO activating enzyme generates an N-terminal peptide, with cleavage site after Arg176, as well as the active enzyme phenoloxidase, which is the key enzyme for melanization. In the present study we demonstrate a role for the N-terminal proPO-peptide in hematopoiesis. Injection of this proPO-peptide increased the number of circulating hemocytes and especially granular hemocytes. We also show that the reactive oxygen species (ROS) production in the anterior proliferative center was enhanced after proPO peptide injection, which is a prerequisite for rapid hemocyte release from the hematopoietic tissue. Moreover, this peptide had an effect on ROS production in in vitro cultured hematopoietic cells and induced spreading of these cells within 72 h. Taken together, our findings show a role of the N-terminal proPO peptide in stimulation of hematopoiesis in crayfish, Pacifastacus leniusculus.
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| 9. |
- Söderhäll, Irene, et al.
(författare)
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Blood cell formation in crustaceans.
- 2022
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Ingår i: Fish and Shellfish Immunology. - : Elsevier. - 1050-4648 .- 1095-9947. ; 131, s. 1335-1342
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Tidskriftsartikel (refereegranskat)abstract
- In crustacean animals the hemocytes are key players in immunity and of crucial importance for the health of the animals. Hemocytes are mainly produced in the hematopoietic tissue and from there released into the circulation where they finally mature. In this review we summarize the latest findings about crustacean hemocyte formation. The role of the extracellular matrix and crosslinking enzyme transglutaminase is discussed. Moreover, important growth factors, transcriptional regulation and recent findings about inducers of hematopoiesis are covered. Finally, we discuss the use of different markers for classification of crustacean hemocytes.
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| 10. |
- Söderhäll, Irene, et al.
(författare)
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Characterization of hemocytes and hematopoietic cells of a freshwater crayfish based on single-cell transcriptome analysis
- 2022
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Ingår i: iScience. - : Cell Press. - 2589-0042. ; 25:8
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Tidskriftsartikel (refereegranskat)abstract
- Crustaceans constitute a species-rich and ecologically important animal group, and their circulating blood cells (hemocytes) are of critical importance in immunity as key players in pathogen recognition, phagocytosis, melanization, and antimicrobial defense. To gain a better understanding of the immune responses to different pathogens, it is crucial that we identify different hemocyte subpopulations with different functions and gain a better understanding of how these cells are formed. Here, we performed single-cell RNA sequencing of isolated hematopoietic tissue (HPT) cells and hemocytes from the crayfish Pacifastacus leniusculus to identify hitherto undescribed hemocyte types in the circulation and show that the circulating cells are more diversified than previously recognized. In addition, we discovered cell populations in the HPT with clear precursor characteristics as well as cells involved in iron homeostasis, representing a previously undiscovered cell type. These findings may improve our understanding of hematopoietic stem cell regulation in crustaceans and other animals.
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| 11. |
- Bagchi, Basabi, et al.
(författare)
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Sexual conflict drives micro- and macroevolution of sexual dimorphism in immunity
- 2021
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Ingår i: BMC Biology. - : BioMed Central (BMC). - 1741-7007. ; 19:1
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: Sexual dimorphism in immunity is believed to reflect sex differences in reproductive strategies and trade-offs between competing life history demands. Sexual selection can have major effects on mating rates and sex-specific costs of mating and may thereby influence sex differences in immunity as well as associated host-pathogen dynamics. Yet, experimental evidence linking the mating system to evolved sexual dimorphism in immunity are scarce and the direct effects of mating rate on immunity are not well established. Here, we use transcriptomic analyses, experimental evolution and phylogenetic comparative methods to study the association between the mating system and sexual dimorphism in immunity in seed beetles, where mating causes internal injuries in females.RESULTS: We demonstrate that female phenoloxidase (PO) activity, involved in wound healing and defence against parasitic infections, is elevated relative to males. This difference is accompanied by concomitant sex differences in the expression of genes in the prophenoloxidase activating cascade. We document substantial phenotypic plasticity in female PO activity in response to mating and show that experimental evolution under enforced monogamy (resulting in low remating rates and reduced sexual conflict relative to natural polygamy) rapidly decreases female (but not male) PO activity. Moreover, monogamous females had evolved increased tolerance to bacterial infection unrelated to mating, implying that female responses to costly mating may trade off with other aspects of immune defence, an hypothesis which broadly accords with the documented sex differences in gene expression. Finally, female (but not male) PO activity shows correlated evolution with the perceived harmfulness of male genitalia across 12 species of seed beetles, suggesting that sexual conflict has a significant influence on sexual dimorphisms in immunity in this group of insects.CONCLUSIONS: Our study provides insights into the links between sexual conflict and sexual dimorphism in immunity and suggests that selection pressures moulded by mating interactions can lead to a sex-specific mosaic of immune responses with important implications for host-pathogen dynamics in sexually reproducing organisms.
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| 12. |
- Ekblom, Charlotta, 1991-
(författare)
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Immune defense reactions in freshwater crayfish : Cellular components
- 2024
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The freshwater signal crayfish, Pacifastacus leniusculus, is a well-established model for studying the immune system of invertebrates. Many important discoveries have been made in this species, among others related to the clotting reaction, hematopoiesis, the prophenoloxidase activating system, the functions of crustacean immune cells, and pathogen recognition. In this thesis, a small contribution is made to this body of work, with the focus on the crayfish cellular defence reactions to the fungal pattern recognition protein, beta-1,3,-glucans, and to an oomycete, the type of pathogen that causes crayfish plague. Through mapping the proteomic reaction in the hemocytes to beta-1,3,-glucan, and then studying some of the identified proteins in more detail, it brings us closer to understanding how these animals defend themselves against fungal infections without relying on adaptive immunity. A proteomic screening of hemocytes was conducted after an injection of laminarin, a beta-1,3,-glucan, and compared to the response to a saline injection and un-injected controls. When compared to both control groups, three proteins were specific to the laminarin group: a glycine-rich peptide, a Kazal-type proteinase inhibitor, and one putative chitin-binding protein; none of which had been previously described. Three other proteins were upregulated in both the saline and the laminarin group: one invertebrate-type (i-type) lysozyme, one crustin, and masquerade. The glycine-rich peptide and the i-type lysozyme were both investigated in detail for their potential functions in the immune- and wounding responses. The peptide was found to be expressed in several tissues, and have a specific activity against the crayfish plague pathogen Aphanomyces astaci, with no effect against any other tested species of oomycete, fungi or bacteria. The i-type lysozyme (Pl-ilys), which was muramidase deficient and therefore likely not involved in antibacterial defence, was found to be able to destabilise clots formed from the crayfish clotting protein and transglutaminase. This result indicated a possible new function for muramidase-deficient i-type lysozymes in crustaceans. A single-cell RNA-sequencing study was also conducted to investigate the types of blood cells and hematopoietic stem cells in P. leniusculus, the result of which shows that there are several potential subtypes among the of granular, semi-granular, hyaline and hematopoietic cells.
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| 13. |
- Hernandez-Perez, Ariadne, et al.
(författare)
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Intestinal microbiome in crayfish : Its role upon growth and disease presentation
- 2023
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Ingår i: Developmental and Comparative Immunology. - : ELSEVIER SCI LTD. - 0145-305X .- 1879-0089. ; 145
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Tidskriftsartikel (refereegranskat)abstract
- The intestine-associated microbiota in crustaceans are considered a key element for maintaining homeostasis and health within the organisms. Recently, efforts have been made to characterize bacterial communities of freshwater crustaceans, including crayfish, and their interplay with the host's physiology and the aquatic environments. As a result, it has become evident that crayfish intestinal microbial communities display high plasticity, which is strongly influenced by both the diet, especially in aquaculture, and the environment. Moreover, studies regarding the characterization and distribution of the microbiota along the gut portions led to the discovery of bacteria with probiotic potential. The addition of these microorganisms to their food has shown a limited positive correlation with the growth and development of crayfish freshwater species. Finally, there is evidence that infections, particularly those from viral etiology, lead to low diversity and abundance of the intestinal microbial communities. In the present article, we have reviewed data on the crayfish' intestinal microbiota, highlighting the most frequently observed taxa and emphasizing the dominance of phylum within this community. In addition, we have also searched for evidence of microbiome manipulation and its potential impact on productive parameters, and discussed the role of the microbiome in the regulation of diseases presentation, and environmental perturbations.
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