Schistosomin: a pronase-sensitive agent in the hemolymph of Trichobilharzia ocellata-infected Lymnaea stagnalis inhibits the activity of albumen glands in vitro

SUMMARYMacrophage-like defence cells (haemocytes) of the pond snailLymnaea stagnalismediate cytotoxicity through reactive oxygen intermediates (ROIs). This activity is NADPH-oxidase dependent, as in mammalian phagocytes during the respiratory burst. In this study, mother sporocysts of schistosomes, the compatibleTrichobilharzia ocellataand the incompatibleSchistosoma mansonievokein vitroROI activities (detected by luminol dependent chemiluminescence, LDCL) fromL. stagnalishaemocytes.S. mansoniis encapsulated by haemocytes and eliminated, whereasT. ocellataescapes encapsulation and survives. Both schistosomes were equally susceptible toin vitrooxidative damage from exposure to hydrogen peroxide and to ROIs generated by a xanthine/xanthine oxidase system. Protocatechuic acid, a specific antagonist of NADPH-oxidase, delayed the killing ofT. ocellataandS. mansonisporocysts by haemocytes of resistant snails (Biomphalaria glabrata and L. stagnalis, respectively). We conclude that ROIs take part in haemocyte-mediated cytotoxicity. However, neither a snail's capability to generate ROIs, nor a schistosome's susceptibility to ROIs, determine snail/schistosome incompatibility. Snail/schistosome compatibility is rather determined by the parasite's ability modulate haemocyte behaviour such that effective encapsulation and the generation of lethal concentrations of ROIs are prevented.

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Excretory–secretory products of Trichobilharzia ocellata and their modulating effects on the internal defence system of Lymnaea stagnalis

Parasitology ◽

10.1017/s0031182096008360 ◽

1997 ◽

Vol 114 (2) ◽

pp. 135-144 ◽

Cited By ~ 4

Author(s):

P. E. NÚÑEZ ◽

M. J. MOLENAAR ◽

W. LAGEWEG ◽

K. W. LI ◽

M. DE JONG-BRINK

Keyword(s):

Lymnaea Stagnalis ◽

Freshwater Snail ◽

Bacterial Clearance ◽

Culture Time ◽

Defence System ◽

Molecular Fraction ◽

The Difference ◽

Secretory Products ◽

Trichobilharzia Ocellata

Excretory–secretory (E–S) products, released by Trichobilharzia ocellata cultured in vitro, were assessed for their modulatory effects on the internal defence system of the freshwater snail, Lymnaea stagnalis. E–S products were concentrated and fractionated using high-pressure liquid chromatography (HPLC) and bacterial clearance activity of haemocytes was taken as a parameter for the internal defence system. Two main fractions of E–S products were produced by T. ocellata, a low molecular fraction (low MW) and a high molecular fraction (high MW), weighing approximately 2 kDa and 40 kDa, respectively. Stage-dependent release of E–S products by transforming miracidia and mother sporocysts was indicated by the difference in proportions of these two fractions, which varied over time. After a culture time of 0–33 h, the smaller fraction was prominent whilst E–S products released between 33 and 72 h consisted of a higher proportion of the high MW fraction. After a culture of 72–96 h, lower and comparable quantities of both fractions were produced by the same batch of cultured parasites. Assaying the 2 fractions released during 0–33 h revealed that the high MW fraction had a suppressing effect on the bacterial clearance activity of haemocytes from juvenile snails and the low MW fraction activated haemocyte activity. Combined fractions obtained during 0–33 h had an activating effect on haemocyte activity but fractions separated from media collected at 33–72 h culture had a suppressing effect on the bacterial clearance activity of the haemocytes. The combined E–S fractions from the 72–96 h incubation had no effect on haemocyte activity. The present study shows that T. ocellata modulates the internal defence system of L. stagnalis by actively interfering with haemocyte function. MALDI-MS analysis of the low MW fraction revealed several components ranging from 450 to 1600 Da. Although it is as yet not clear which factor is responsible for the activating effect it is supposedly a peptide as it is pronase-sensitive and resistant to TFA and acetonitrile.

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Modulation of the activity of the internal defence system of the pond snail Lymnaea stagnalis by the avian schistosome Trichobilharzia ocellata

Parasitology ◽

10.1017/s0031182000060777 ◽

1992 ◽

Vol 104 (1) ◽

pp. 33-40 ◽

Cited By ~ 24

Author(s):

R. I. Amen ◽

J. M. C. Baggen ◽

P. D. Bezemer ◽

M. De Jong-Brink

Keyword(s):

Lymnaea Stagnalis ◽

Snail Host ◽

Parasite Development ◽

Pond Snail ◽

Phagocytosis Assay ◽

Time Points ◽

Defence System ◽

A Cell ◽

Trichobilharzia Ocellata

SUMMARYEffects of infection with the avian schistosome Trichobilharzia ocellata on the activity of the internal defence system of the intermediate snail host Lymnaea stagnalis were studied, utilizing an in vitro phagocytosis assay for determining haemocyte activity. A distinction was made between plasma- and cell-associated effects. The period immediately after penetration of the parasite into the snail host (1.5–72 h post-exposure (p.e.)) was extensively studied. In addition, several time-points coinciding with the later–successive–stages of parasite development (2, 4, 6, 8 and 10 weeks p.e.) were investigated. Plasma-associated enhancement of defence activity was found between 1.5 and 6 h p.e., followed by plasma-associated suppression between 12 and 72 h p.e. A cell-associated activation was found between 1.5 and 6 h p.e. and also at 8 and 10 weeks p.e. How these effects on the defence system may be related to phenomena observed in infected snails at these time-points is discussed.

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Trichobilharzia ocellata infections in its snail host Lymnaea stagnalis: an in vitro study showing direct and indirect effects on the snail internal defence system, via the host central nervous system

Parasitology ◽

10.1017/s0031182000074588 ◽

1992 ◽

Vol 105 (3) ◽

pp. 409-416 ◽

Cited By ~ 6

Author(s):

R. I. Amen ◽

M. De Jong-Brink

Keyword(s):

Central Nervous System ◽

Nervous System ◽

Indirect Effects ◽

Phagocytic Activity ◽

Lymnaea Stagnalis ◽

Snail Host ◽

Defence System ◽

Trichobilharzia Ocellata

SUMMARYIn this in vitro study we investigated whether previously described in vivo plasma-associated effects, that occurred in the period shortly after penetration of Trichobilharzia ocellata into the snail host Lymnaea stagnalis (1.5–72 h post-exposure; p.e.) were direct and/or indirect effects of parasite-derived factor(s). It was investigated whether the effect is mediated by the central nervous system (CNS) of the host. Phagocytic activity of the haemocytes was taken as a parameter for the activity of internal defence of the host. A number of preliminary experiments were performed. When the supernatant of in vitro cultured parasites (33 h; corresponding with their developmental stage in vivo when plasma-associated activation was found) was applied directly to monolayers of haemocytes, it appeared to enhance their phagocytic activity. No direct effect, however, was found with a supernatant of parasites cultured for a longer period of time (72 h; when, in vivo, a plasma-associated suppression was found). In this case, indirect suppression was detected: the parasites appeared to have released a factor that induced the CNS of the host to release material suppressing the activity of the internal defence system of the host. To date the nature of this factor is unknown.

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Modulation of the bacterial clearance activity of haemocytes from the freshwater mollusc, Lymnaea stagnalis, by the avian schistosome, Trichobilharzia ocellata

Parasitology ◽

10.1017/s0031182000078331 ◽

1994 ◽

Vol 109 (3) ◽

pp. 299-310 ◽

Cited By ~ 20

Author(s):

P. E. Núñez ◽

C. M. Ademal ◽

M. De Jong-Brink

Keyword(s):

Lymnaea Stagnalis ◽

Colour Change ◽

Snail Host ◽

Aeromonas Salmonicida ◽

Bacterial Clearance ◽

Gastropod Mollusc ◽

Freshwater Mollusc ◽

Viable Bacteria ◽

Trichobilharzia Ocellata

SummaryThe ability of haemocytes, from the haemolymph of the gastropod mollusc Lymnaea stagnalis, to recognize and eliminate the bacterium Aeromonas salmonicida was shown using an in vitro bacterial clearance assay. The assay employs a dye which is reduced by A. salmonicida in direct proportion to the number of viable bacteria resulting in a colour change which can be determined spectrophotometrically. Addition of cytochalasin B resulted in a marked decrease in bacterial clearance, implicating both intracellular and extracellular cytotoxicity of haemocytes. A comparison of haemocytes from uninfected snails and snails infected with the avian schistosome parasite Trichobilharzia ocellata showed that both juveniles and adults of L. stagnalis were susceptible to infection with T. ocellata. After exposure to the trematode for 1·5 h the haemocytes from these infected snails had an enhanced clearance capacity, whilst cells obtained from snails with 24–96 h infections showed decreased clearance of the bacteria, indicating suppression by the parasite. Haemocytes, as well as plasma, which was tested on haemocytes from uninfected snails, were used and hence a distinction was made between cell and humoralassociated effects. The results show that both cellular and humoral components of immunity were activated, then suppressed, following exposure to the parasite. Infection with T. ocellata seems to have a modulating effect on the bactericidal activity of the internal defence system of the snail host, L. stagnalis.

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In vitro release of the anti-gonadotropic hormone, schistosomin, from the central nervous system of Lymnaea stagnalis is induced with a methanolic extract of cercariae of Trichobilharzia ocellata

Parasitology ◽

10.1017/s0031182000061758 ◽

1992 ◽

Vol 104 (2) ◽

pp. 309-314 ◽

Cited By ~ 8

Author(s):

H. D. F. H. Schallig ◽

M. J. M. Sassen ◽

M. De Jong-Brink

Keyword(s):

Central Nervous System ◽

Nervous System ◽

Lymnaea Stagnalis ◽

Methanolic Extract ◽

Parasitic Infection ◽

In Vitro Release ◽

Intermediate Hosts ◽

The Central Nervous System ◽

Trichobilharzia Ocellata

SUMMARYInfection with digenetic trematodes causes an inhibition or complete cessation of fecundity in their intermediate hosts, freshwater snails. It has been demonstrated in the host–parasite combination Lymnaea stagnalis–Trichobilharzia ocellata that the action of the female gonadotropic hormones upon their target organs is inhibited by the peptide schistosomin. Schistosomin is produced in the central nervous system of the snail and released upon parasitic infection. In order to study the in vitro release of schistosomin, a bioassay was developed. Central nervous systems were incubated with either an acetic acid or a methanolic extract of larval stages of Trichobilharzia ocellata (miracidia, mother sporocysts, cercariae). The incubation media were chromatographed using HPLC and released schistosomin (-like material) was tested for bioactivity in the calfluxin bioassay. The in vitro release of schistosomin was only induced with a methanolic extract of cercariae. The nature of the cercarial factor is discussed.

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