Identification of a phenoloxidase- and melanin-dependent defence mechanism in Achatina fulica infected with Angiostrongylus vasorum

Abstract

BackgroundAngiostrongylus vasorum has different freshwater aquatic and terrestrial gastropod molluscs as an intermediate host, e.g. Arion spp. The mollusc Achatina fulica is a danger to public health, given the large diversity of nematodes utilizing it as an intermediate host, such as the parasites of the genus Angiostrongylus, of importance in human and veterinary medicine. Achatina fulica has been shown to have an excellent capacity for maintaining outbreaks and natural infections with A. cantonensis in Asia. Within the mollusc, the nematode parasites activate haemocytes and/or haemolymph factors and in some invertebrates, phenoloxidase (PO), that induces the release of toxic elements and eliminates the parasites. Despite the importance of A. fulica in the life-cycle of nematodes, little is known regarding the defence mechanisms involving PO in molluscs infected with nematodes. Here, the presence of PO and nitric oxide (NO) in the haemolymph and haemocytes of A. fulica infected with first-stage (L1) larvae of Angiostrongylus vasorum was evaluated, together with the presence of melanin in the cephalopod mollusc tissue.ResultsAn increase in PO at one day post infection (dpi), in comparison with the control using the substrates L-tyrosine (F(4,90) = 6.73, P = 0.00006), L-DOPA (F(4,90) = 22.67, P = 0.02) and p-phenylenediamine (PPD) (F(4,90) = 27.58, P = 0.0019), was observed. PO increase coincided with the presence of melanin in the cephalopodal tissue. At 8 dpi, PO activity, compared to L-DOPA (F(4,90) = 22.67, P = 0.00002) and PPD (F(4,90) = 27.58, P = 0.079) decreased, while melanin increased. At 13 dpi, PO decreased with PPD (F(4,90) = 27.58, P = 0.000015) and also the amount of melanin observed in histology. At 30 dpi, PO increased along with the substrates L-DOPA and PPD, while melanin decreased. NO levels increased until 8 dpi, and decreased after 13 dpi.ConclusionsTo our knowledge, this is the first study that illustrates PO activity in a helminth-infected A. fulica and provides the first observation of an L-tyrosine dependent PO activity in molluscs infected with A. vasorum. This work suggests that PO pathway may help to control A. vasorum infection in A. fulica.

Morph-dependent nematode infection and its association with host movement in the land snail Cepaea nemoralis (Mollusca, Gastropoda)

“…Indeed, our initial hypothesis when designing this experiment (in 2019) was based on the oft-seen association between immunity and melanin (San-Jose & Roulin, 2018), and on the role of melanin in nematode defence in e.g. Lissachatina fulica (Coaglio et al ., 2018), although other pigments may also have links to immunity (Tan et al ., 2020). While it is still likely that melanin-associated pathways play a role in immune defence in Cepaea (Scheil et al ., 2014), the actual mechanism is probably not as simple as “banded snails constitutively produce more melanin, and thus are better protected”.…”

“…Moreover, as in insects (González-Santoyo & Córdoba-Aguilar, 2012; San-Jose & Roulin, 2018), melanin and the phenoloxidase pathway involved in melanisation may play a key role in snail defences against parasites. For instance, accumulation of melanin around nematodes encapsulated in the haemolymph is documented in the snail Lissachatina fulica (Coaglio et al ., 2018). Phenoloxidase activity has been shown to increase in L. fulica and Helix lucorum after infection by nematodes or trematodes, respectively (Coaglio et al ., 2018; Unlu & Ekici, 2021).…”

“…For instance, accumulation of melanin around nematodes encapsulated in the haemolymph is documented in the snail Lissachatina fulica (Coaglio et al ., 2018). Phenoloxidase activity has been shown to increase in L. fulica and Helix lucorum after infection by nematodes or trematodes, respectively (Coaglio et al ., 2018; Unlu & Ekici, 2021). While less studied, carotenoids may also contribute to immune responses in invertebrates (Tan et al ., 2020).…”

An Evolutionary Perspective of Dopachrome Tautomerase Enzymes in Metazoans

“…Melanin is widespread in the animal kingdom, where it is involved in several important biological functions including pigmentation and immune-defense [6,8,66]. Here, we now traced the distribution of the DCT and DCE/ yellow gene families through metazoan evolution.…”

“…Although melanotic encapsulation is mostly described in arthropods [7], the presence of dark, melanin-like granules surrounding pathogens has also been reported in several mollusk species. Examples for this come from the gastropod Achatina fulica infected by nematodes [8], Roseovarius crassostreae infecting Crassostrea virginica and Vibrio tapetis infecting Ruditapes phlilippinarum [9]. Here, all pallial epithelia of bivalves were capable of melanization [9], but most of the genes involved in the melanogenic pathway were highly expressed in the mantle edge [10,11], a tissue involved in both defense mechanisms and shell formation in bivalve and gastropod mollusks [12,13,14].…”