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1.
A modulation of the phagocytic activity of hemocytes from the common periwinkle Littorina littorea by secretory-excretory products (SEP) released by trematode rediae during axenic in vitro cultivation was studied. The SEP released by the parasites Himasthla elongata (Echinostomatidae) and Cryptocotyle lingua (Heterophyidae) were found to inhibit the phagocytosis of zymozan particles by periwinkle hemocytes. The specificity of SEP effects was assessed: SEP of Himasthla militaris and Cryptocotyle concavum, two trematodes belonging to the same genera but infecting another closely related prosobranch snail Hydrobia ulvae, were also shown to be able to suppress L. littorea hemocytes phagocytic activity. However, no decrease in phagocytosis rate was observed when SEP of H. elongata and C. lingua were applied to monolayers of hemocytes from the bivalve mollusc Mytilus edulis. SEP from H. elongata was fractionated; only those fractions containing proteins of molecular weight more than 50 kDa were shown to possess inhibitory activity. Different H. elongata SEP concentrations were tested in for their ability to suppress phagocytosis by L. littorea hemocytes. Even very low SEP concentrations were shown to retain their ability to decrease phagocytosis rate, the inhibitory effect being dose-dependent. Hemocytes derived from snails naturally infected with H. elongata were also found to have lower phagocytic ability as compared to healthy individuals. 相似文献
2.
For a better understanding of virus x host interactions, transmission electron microscopy was used to characterize the intrahaemocoelic infection of Anticarsia gemmatalis larval haemocytes by A. gemmatalis M nucleopolyhedrovirus (AgMNPV). At 12 h post-infection (h p.i.), we observed nuclear hypertrophy, budded virus assembling, and protrusion towards the cytoplasm, virion envelopment, and accumulation of fibrillar aggregates in the cytoplasm. Around 24 h p.i., fibrillar aggregates also appeared inside nuclei of infected cells. By 48 h p.i., virogenic stroma and polyhedra were visualised in nuclei and at 72 h p.i., widespread infection in haemocytes was observed. Cell remnants and free polyhedra were phagocytosed by granular haemocyte 1 and plasmatocytes. Entire cells were phagocytosed only by plasmatocytes. Necrosis of infected cells was quite common, suggesting a putative cytotoxic response. Granular haemocyte 1 presented a more exuberant protrusion of budded viruses in comparison to other haemocytes. All types of haemocytes were shown to be infected, and the intense virus replication in some of these cells reveals the importance of haemolymph for AgMNPV spread in its natural host, a critical factor for permissiveness. 相似文献
3.
Cricket haemocytes were derived from either haemolymph or haemopoietic organs (lymph glands) of insects and introduced to a primary culture. Varied isolation protocols, tissue culture vessels, media compositions and cell densities were tested to determine the optimal conditions for in vitro maintenance of haemocytes, and for subsequent light and electron microscopic analysis of monolayers. Freshly prepared Mitsuhashi and Maramorosh (MM;Sigma, Steinheim, Germany) insect medium (420 mOsm), buffered with sodium bicarbonate (pH 7.2) and supplemented with 10 % FCS, was found to be most appropriate for haemocyte maintenance. All tested tissue culture vessels (FLEXiperm units, multiwell plates and Thermanox slides, with the exception of Melineux agar plates), were suitable for cell attachment and haemocyte monolayers formation. Viability of cultured cells was confirmed by LIVE/DEAD Viability/Cytotoxity Kit for Eukaryotic Cells. Free circulating haemocytes were cultivated up to 27 days and then degraded. Infection with the microsporidian Paranosema grylli or the coccidian Adelina grylli caused noticeable swelling of host lymph glands (haemopoietic tissue) and increase in the number of cells comprising the glands. The cells derived from haemopoietic tissue were maintained for maximum 5 days; thereafter multiplication of bacteria normally inhabiting cricket lymph glands destroyed monolayers and killed the cells. Microsporidian and coccidian invasive stages (spores and sporozoites, respectively) were isolated from infected tissues, resuspended in MM medium and added to haemocyte monolayers in ratios 1 zoite per haemocytes or 10 spores per 1 haemocyte. Actively moving zoites contacted and penetrated the cultured cells. Unlike coccidian zoites, microsporidian spores were phagocytized by haemocytes. Application of fluorescent LIVE/DEAD kit allowed to visualize internalized parasites inside host cells as clearly shaped dark areas. The present study has demonstrated that 1) cricket haemocytes from both circulating haemolymph and lymph glands can be short-term cultivated on tissue culture vessel surfaces which made possible their further light and electron microscopic analysis; 2) short-term haemocyte cultures may be employed to study host-parasite interactions, in particular, to follow the initial steps of parasite internalization inside host cell; 3) Fluorescent assay with Viability/Cytotoxity Kit for Eukaryotic Cells (Molecular Probes, Oregon) allows to observe penetration of these parasites into cultured cells. 相似文献
4.
A positive correlation exists between the pathogenicity of bacteria and fungi when evaluated in the insect Galleria mellonella and mice. This work sought to determine whether fluctuations in the number of haemocytes and the proliferation of yeast cells in infected larvae could be used to determine the relative pathogenicity of a range of yeast isolates. Larvae were inoculated with 1 x 10(6) stationary-phase yeast cells and incubated in the dark at 30 degrees C for 48 h. The results indicated that larvae inoculated with the most pathogenic isolates (i.e. those capable of killing >80% of infected larvae) showed a significant reduction in haemocyte density. Larvae inoculated with isolates of low pathogenicity (i.e. capable of killing <20% of infected larvae) demonstrated only a small fluctuation in haemocyte numbers. The most pathogenic yeast isolates proliferated in the larvae, whereas the isolates of low pathogenicity did not. These results demonstrate a relationship between the ability of yeast isolates to kill larvae and changes in haemocyte density and yeast cell density in infected larvae. These end points may extend the applicability of the G. mellonella system for use with a wider range of microbial isolates. 相似文献
5.
The first mollusc mRNA coding G-protein-coupled transmembrane receptor (GPСR), homologous to human receptors LUSTR 1 (GPR107) and LUSTR 2 (GPR108), was isolated from haemocytes of common periwinkle Littorina littorea. The analyses showed that the full-length cDNA is 1935 bp long and is predicted to encode a 614 amino acid protein (named Lit-LUSTR) with a calculated molecular mass of 69.6 kDa and theoretical isoelectric point 7.59. Pair-wise comparisons between Lit-LUSTR and LUSTR proteins from human or mouse have approximately 38% identity and 56% similarity. Lit-LUSTR clusters with LUSTR-A sub-family proteins and is a first characterization of proteins containing Lung7TM-R domain in Mollusca. Significant differences were found between the Lit-LUSTR mRNA levels in haemocytes of healthy periwinkles and those naturally infected with the echinostome trematode Himasthla elongata. Down regulated expression of the LUSTR-like receptor caused by infection illustrates modification of the haemocyte receptor system and may be attributed to the previously demonstrated greater numbers of “immature” haemocytes in the circulation of infected snails. 相似文献
6.
Live adult and juvenile entomopathogenic Steinernema carpocapsae DD136 (P. Nematoda) were not subjected to adhesion by haemocytes of lepidopteran insect larvae of Galleria mellonella or Malacosoma disstriain vitro or in vivo. In vitro freeze-killed nematodes exhibited haemocyte attachment, the intensity increasing with time. Accumulation of haemocytes on the dead nematodes was associated with host phenoloxidase activity; live nematodes and their exudates did not activate the enzyme whereas dead nematodes but not their exudate did activate phenoloxidase. Live-nematode exudate inhibited granular cell and some plasmatocyte adhesion to slides, increased granular cell but not plasmatocyte dissociation from preformed haemocyte monolayers and in vivo elevated total haemocyte counts and changed the floating haemocyte types while impairing bacterial removal from the haemolymph. Dead-nematode exudate did not affect these parameters thus immunosuppressant activity by live nematodes may represent the release of inhibitors not associated with their cuticle. The third stage juveniles released the inhibitors. 相似文献
7.
The effects of a xenobiotic on the circulating haemocytes of Lymnaea stagnalis were investigated after short-term (24 h, 96 h) and long-term (504 h) exposure of snails to environmental concentrations.
Fomesafen, a pro-oxidant generator led to the activation of the haemocyte apoptotic program by promoting reactive oxygen species
(ROS). Cells entering apoptosis underwent a series of events, both on the plasma membrane and in the mitochondria; these events
were quantified by flow cytofluorometry. The data showed a loss of mitochondrial transmembrane potential (Δψm), which was
dose-dependent and time-dependent and related to an increased release of superoxide anions. The phosphatidylserine that was
exposed at the outer plasma membrane was not related to the disruption of either ROS or Δψm but was strongly correlated with
the haemocyte concentration (total haemocyte count). This cascade of apoptotic processes occurred in a dose-independent manner
and was not strengthened over time. The increase of circulating haemocytes depended upon the life span of the cells and might
have reflected either facilitated cell turn-over or the accompanying presence of haemocytes phagocytosing apoptotic cells. 相似文献
8.
Preliminary study on haemocyte response to white spot syndrome virus infection in black tiger shrimp Penaeus monodon 总被引:4,自引:0,他引:4
van de Braak CB Botterblom MH Huisman EA Rombout JH van der Knaap WP 《Diseases of aquatic organisms》2002,51(2):149-155
White spot syndrome virus (WSSV) has been a major cause of shrimp mortality in aquaculture in the past decade. In contrast to extensive studies on the morphology and genome structure of the virus, little work has been done on the defence reaction of the host after WSSV infection. Therefore, we examined the haemocyte response to experimental WSSV infection in the black tiger shrimp Penaeus monodon. Haemolymph sampling and histology showed a significant decline in free, circulating haemocytes after WSSV infection. A combination of in situ hybridisation with a specific DNA probe for WSSV and immuno-histochemistry with a specific antibody against haemocyte granules in tissue sections indicated that haemocytes left the circulation and migrated to tissues where many virus-infected cells were present. However, no subsequent haemocyte response to the virus-infected cells was detected. The number of granular cells decreased in the haematopoietic tissue of infected shrimp. In addition, a fibrous-like immuno-reactive layer appears in the outer stromal matrix of tubule walls in the lymphoid organ of infected shrimp. The role of haemocytes in shrimp defence after viral infection is discussed. 相似文献
9.
The number of circulating haemocytes, the size of the haematopoietic organ, and the size of haemocyte capsules around the parasite were studied in M-line Biomphalaria glabrata snails exposed to 100 or 400 first-stage larvae of Angiostrongylus cantonensis. The number of haemocytes in exposed snails increased significantly at 1 day post-exposure, decreased to control value, and then increased again. The decrease in number of circulating haemocytes is probably due to the removal of cells from the circulation to participate in encapsulation of larvae. The majority of circulating haemocytes in M-line B. glabrata are fully-spread granulocytes, which increase significantly in number in snails following exposure to A. cantonensis larvae. However, populations of partially-spread granulocytes, round cells, hyalinocytes and miscellaneous haemocytes were relatively constant. The size of capsules around the parasite increased during the 42-day interval of the experiment. The haematopoietic organ increased in size in response to infection. 相似文献
10.
Vladimír Skála Alena ?erníková Zuzana Jindrová Martin Ka?ny Martin Vostry Anthony J. Walker Petr Horák 《PloS one》2014,9(11)
Radix lagotis is an intermediate snail host of the nasal bird schistosome Trichobilharzia regenti. Changes in defence responses in infected snails that might be related to host-parasite compatibility are not known. This study therefore aimed to characterize R. lagotis haemocyte defence mechanisms and determine the extent to which they are modulated by T. regenti. Histological observations of R. lagotis infected with T. regenti revealed that early phases of infection were accompanied by haemocyte accumulation around the developing larvae 2–36 h post exposure (p.e.) to the parasite. At later time points, 44–92 h p.e., no haemocytes were observed around T. regenti. Additionally, microtubular aggregates likely corresponding to phagocytosed ciliary plates of T. regenti miracidia were observed within haemocytes by use of transmission electron microscopy. When the infection was in the patent phase, haemocyte phagocytic activity and hydrogen peroxide production were significantly reduced in infected R. lagotis when compared to uninfected counterparts, whereas haemocyte abundance increased in infected snails. At a molecular level, protein kinase C (PKC) and extracellular-signal regulated kinase (ERK) were found to play an important role in regulating these defence reactions in R. lagotis. Moreover, haemocytes from snails with patent infection displayed lower PKC and ERK activity in cell adhesion assays when compared to those from uninfected snails, which may therefore be related to the reduced defence activities of these cells. These data provide the first integrated insight into the immunobiology of R. lagotis and demonstrate modulation of haemocyte-mediated responses in patent T. regenti infected snails. Given that immunomodulation occurs during patency, interference of snail-host defence by T. regenti might be important for the sustained production and/or release of infective cercariae. 相似文献
11.
A research project to compare productive traits (growth and mortality), disease susceptibility and immune capability between Ostrea edulis stocks was performed. This article reports the results on the immune capability and its relation with infection by the intrahaemocytic protozoan Bonamia ostreae. Four to five oyster spat families were produced from each of four European flat oyster populations (one from Ireland, one from Greece and two from Galicia, Spain) in a hatchery. The spat were transferred to a raft in the Ría de Arousa (Galicia) for on growing for 2 years. Total haemocyte count (THC) and differential haemocyte count (DHC) were estimated monthly through the second year of growing-out. Three types of haemocytes were distinguished: granulocytes (GH), large hyalinocytes (LHH) and small hyalinocytes (SHH). Significant correlations between the mean relative abundance of GH and SHH of the families and the mean prevalence of B. ostreae, the overall incidence of pathological conditions and the cumulative mortality of the families were found; these correlations supported the hypothesis that high %GH and low %SHH would enhance oyster immune ability and, consequently, would contribute to lower susceptibility to disease and longer lifespan. Infection by B. ostreae involved a significant increase of circulating haemocytes, which affected more markedly the LHH type. The higher the infection intensity the higher the %LHH. This illustrates the ability of B. ostreae to modulate the immune responses of the O. edulis to favour its own multiplication. A significant reduction of the phenoloxidase activity in the haemolymph of oysters O. edulis infected by B. ostreae was observed. Nineteen enzymatic activities in the haemolymph of O. edulis and Crassostrea gigas (used as a B. ostreae resistant reference) were measured using the kit api ZYM®, Biomerieux. Qualitative and quantitative differences in enzyme activities in both haemocyte and plasma fractions between B. ostreae noninfected O. edulis from different origins were recorded. However, no clear positive association between enzyme activity and susceptibility to bonamiosis was found. The only enzyme detected in the resistant species C. gigas that was not found in the susceptible one O. edulis was β-glucosidase (in plasma). B. ostreae infected O. edulis showed significant increase of some enzyme activities and the occurrence of enzymes that were not detected in noninfected oysters. These changes could be due to infection-induced enzyme synthesis by the host or to enzyme synthesis by the parasite. 相似文献
12.
Benjamin Morga Isabelle Arzul Bruno Chollet Tristan Renault 《Fish & shellfish immunology》2009,26(6):836-842
Bonamia ostreae is an intracellular protozoan parasite, infecting haemocytes of the European flat oyster Ostrea edulis. Oyster defence mechanisms mainly rely on haemocytes. In the present study in vitro interactions between parasites and flat oyster haemocytes were investigated using flow cytometry and light microscopy.Haemocyte parameters including: non specific esterase activity, reactive oxygen species (ROS) production and phagocytosis were monitored using flow cytometry after 2 h cell incubation with live and dead B. ostreae. Two ratios of parasites per haemocyte were tested (5:1 and 10:1), haemocytes alone were used as controls and the experiment was carried out three times. Flow cytometry revealed a decrease of non specific esterase activities and ROS production by haemocytes after incubation with live parasites, while there was little difference in phagocytosis activity when compared with controls. Similarly, dead parasites induced a decrease in haemocyte activities but to a lesser extent compared to live parasites. These results suggest that B. ostreae actively contributes to the modification of haemocyte activities in order to ensure its own intracellular survival. 相似文献
13.
Following infection with Schistosoma mansoni larvae, haemocytes of resistant Biomphalaria glabrata snails execute a rapid defence during which they migrate towards and encapsulate the parasites. Such immediate and precise responses are thought to depend on signal transduction cascades though the signalling components involved remain largely unknown. It is proposed that mitogen-activated protein kinases may play a role in B. glabrata immune signalling, in particular p38 mitogen-activated protein kinases, which are known to be associated with stress and inflammatory signalling. Using degenerate PCR followed by Rapid Amplification of cDNA Ends a full-length p38 mitogen-activated protein kinase-like cDNA was cloned from both the B. glabrata embryonic (Bge) cell line (Bge-p38) and haemocytes (Bgh-p38). In addition, B. glabrata p38 mitogen-activated protein kinase activation was examined at the protein level in Western blot analyses using an antibody that specifically recognises activated/diphosphorylated p38 mitogen-activated protein kinase. Results showed that Bge cell p38 mitogen-activated protein kinase was activated/phosphorylated following 30 min incubation with anisomycin, an established p38 mitogen-activated protein kinase activator. Furthermore, p38 mitogen-activated protein kinase was also activated after only 5 min exposure to either the beta-glucan polymer laminarin or S. mansoni larval excretory-secretory products. In a comparative study, activated haemocyte p38 mitogen-activated protein kinase could also be detected using the anti-phosphorylated p38 antibody following cell treatment with anisomycin. However, in contrast with Bge cells, haemocyte p38 was not activated by either excretory-secretory products or laminarin treatments, suggesting fundamental differences in the role of p38 mitogen-activated protein kinase in signal transduction pathways between haemocytes and Bge cells. 相似文献
14.
Ballarin L Menin A Tallandini L Matozzo V Burighel P Basso G Fortunato E Cima F 《Cell and tissue research》2008,331(2):555-564
A recurrent blastogenetic cycle characterizes colonies of the ascidian Botryllus schlosseri. This cycle starts when a new zooid generation opens its siphons and ends with take-over, when adult zooids cease filtering
and are progressively resorbed and replaced by a new generation of buds, reaching functional maturity. During the generation
change, massive apoptosis occurs in the colony, mainly in the tissues of old zooids. In the present study, we have investigated
the behaviour of haemocytes during the colonial blastogenetic cycle, in terms of the occurrence of cell death and the expression
of molecules involved in the induction of apoptosis. Our results indicate that, during take-over, caspase-3 activity in haemocyte
lysates increases. In addition, about 20%–30% of haemocytes express phosphatidylserine on the outer leaflet of their plasma
membrane, show DNA fragmentation and are immunopositive for caspase-3. Senescent cells are quickly ingested by circulating
phagocytes that frequently, having once engulfed effete cells, in turn enter apoptosis. Dying cells and corpses are replaced
by a new generation of cells that appear in the circulation during the generation change.
This research was supported by the Italian M.I.U.R. (PRIN 2006) 相似文献
15.
When larvae of the ectoparasitic wasp Eulophus pennicornis were incubated for 4 h on balls of cotton wool soaked in tissue culture medium (TC-100), they released a variety of factors. Subsequent incubation of these larval wasp secretions with monolayers of haemocytes from their host, Lacanobia oleracea, demonstrated that they adversely affect haemocyte morphology, behaviour and viability. For instance, when monolayers of haemocytes were incubated for 18 h in TC-100, approximately 73% of the cells present, attached firmly to and spread over the tissue culture surface by extending pseudopods. By contrast, when incubated in TC-100 containing larval wasp secretions, only about 27% of the haemocytes present remained attached to the tissue culture surface after washing. The majority of these had a rounded configuration and neither spread nor extended pseudopods. Furthermore, viability assays indicated that approximately 36% of the attached haemocytes were dead, as opposed to 11-12% in the controls. The E. pennicornis secretions also significantly reduced the ability of L. oleracea haemocytes to move across the surface of the slide and form clumps (p≤0.0005) and to phagocytose FITC-labelled Escherichia coli in vitro (p≤0.0005). These results indicate that secretions from E. pennicornis larvae contain an anti-haemocyte factor(s) that can kill and/or alter the behaviour of host haemocytes. As a result, the ability of the haemocytes to execute important immune responses is compromised. Preliminary data suggest that the active molecules are proteins, and that their mechanism of action may involve inhibition of polymerization and/or disorganization of the haemocyte cytoskeleton. 相似文献
16.
Five types of haemocytes have been identified in the haemolymph ofSpilostethus hospes. Their morphology and micrometric measurements have been provided. Changes in the total and differential haemocyte population
[total haemocyte count (THC) and differtial haemocyte count (DHC)] as well as in the absolute number of haemocytes in circulation
have been assessed in relation to eclosion, sex and mating. The haemogram profile was studied prior to and immediately after
eclosion and also prior to and after copulation. Though the THC was not significantly different immediately before and after
eclosion, there was a significant increase in total count prior to copulation. Mated females registered an increase in total
count but there was no appreciable change in the mated males. Granulocytes were the most abundant of the haemocyte types in
both the sexes and mating caused a significant increase in the plasmatocyte count in females. Changes in the blood volume
as well as the mitotic activity of the haemocytes is also discussed 相似文献
17.
Perkinsus marinus 《Fish & shellfish immunology》1996,6(8):581-597
The effects of extracellular products (ECP) and purified proteases from the protozoan parasitePerkinsus marinuson three host defence parameters (haemocyte motility, lysozyme and haemagglutinin) of the eastern oyster,Crassostrea virginica, were investigated. ECP with high proteolytic activities, as well as purified proteases, significantly decreased the random migration of haemocytes through micro-porous filters in Boyden chambers. Stimulation of haemocyte migration byP. marinuscells orP. marinuscell lysate was also dramatically reduced by ECP and purified proteases. Incubation of oyster plasma with ECP and purified proteases caused a significant decrease in lysozyme activity and also appeared to reduce haemagglutinin titres. These data suggest thatP. marinusECP, as well as the proteolytic fraction of the ECP, can modulate some defence parameters of oystersin vitro. 相似文献
18.
19.
Zahida Zahoor Angela J. Davies Ruth S. Kirk David Rollinson Anthony John Walker 《Cell stress & chaperones》2010,15(5):639-650
Synthesis of heat shock proteins (HSPs) following cellular stress is a response shared by many organisms. Amongst the HSP
family, the ∼70 kDa HSPs are the most evolutionarily conserved with intracellular chaperone and extracellular immunoregulatory
functions. This study focused on the effects of larval excretory-secretory products (ESPs) from the parasite Schistosoma mansoni on HSP70 protein expression levels in haemocytes (defence cells) from its snail intermediate host Biomphalaria glabrata. S. mansoni larval stage ESPs are known to interfere with haemocyte physiology and behaviour. Haemocytes from two different B. glabrata strains, one which is susceptible to S. mansoni infection and one which is resistant, both showed reduced HSP70 protein levels following 1 h challenge with S. mansoni ESPs when compared to unchallenged controls; however, the reduction observed in the resistant strain was less marked. The
decline in intracellular HSP70 protein persisted for at least 5 h in resistant snail haemocytes only. Furthermore, in schistosome-susceptible snails infected by S. mansoni for 35 days, haemocytes possessed approximately 70% less HSP70. The proteasome inhibitor, MG132, partially restored HSP70
protein levels in ESP-challenged haemocytes, demonstrating that the decrease in HSP70 was in part due to intracellular degradation.
The extracellular signal-regulated kinase (ERK) signalling pathway appears to regulate HSP70 protein expression in these cells,
as the mitogen-activated protein-ERK kinase 1/2 (MEK1/2) inhibitor, U0126, significantly reduced HSP70 protein levels. Disruption
of intracellular HSP70 protein expression in B. glabrata haemocytes by S. mansoni ESPs may be a strategy employed by the parasite to manipulate the immune response of the intermediate snail host. 相似文献
20.
Dr. P. J. S. Smith E. A. Howes C. A. Leech J. E. Treherne 《Cell and tissue research》1986,243(2):367-374
Summary Injection of physiologically inert particles (fluorescent microspheres) has a profound effect on neural repair of central nervous connectives of the cockroach Periplaneta americana following selective glial disruption. The injected particles, which do not gain direct access to the central nervous tissues, are taken up by a relatively small proportion (< 10%) of the haemocytes. This interference with haemocyte function virtually abolishes the appearance of the granule-containing cells (which are prominently involved in normal glial repair) and produces abnormal reorganization of the superficial glial elements. These results are interpreted as evidence that the granule-containing cells are derived from haemocytes which are critically involved in glial repair. 相似文献