Glycogen catabolism enzymes and protein fractions in the third and fourth larval stages ofAnisakis simplex

Abstract

Extracts of Anisakis simplex third (L3) and fourth (L4) larval stages were assayed for protein content and activity and properties of alpha-amylase, glucoamylase and glycogen phosphorylase. Protein content in L4 was twice that in L3. SDS-PAGE applied to both larval stages revealed 22 protein fractions in each, including five stage-specific fractions in each larval stage. The L3 extracts contained three amylase isoenzymes: alpha 1, alpha 2 and alpha 3; their molecular weights were 64, 29 and 21 kDa, respectively. Only one amylase isoenzyme (64 kDa) was found in the L4 extracts. Glycogen in L3 was found to be broken down mostly by hydrolysis because of low glycogen phosphorylase activity. The alpha-amylase activity in L4 was higher than that in L3 by half and the glycogen phosphorylase activity was ten times higher. In addition, the same enzymes isolated from L3 and L4 were found to differ in their properties. These differences could be manifestations of metabolic adaptations of A. simplex larvae to host switch from fish (L3) to mammals (L4), i.e. adaptations to a new habitat.

Expression of Genes Encoding the Enzymes for Glycogen and Trehalose Metabolism in L3 and L4 Larvae ofAnisakis simplex

“…An inverse relationship was noted between the predominance of GS transcripts over GP transcripts in A. simplex and the predominance of GP over GS in dauer larvae of C. elegans [ 42 ]. Similar results were reported in a previous study investigating the activity of glycogen enzymes in L4 larval stage [ 25 ], which revealed higher activity levels of GP than GS in older larvae of A. simplex ( Figure 3 ).…”

“…The aim of this study was to investigate and compare the expression of genes encoding the enzymes involved in trehalose synthesis (TPS and TPP), glycogen synthesis (GS), and breakdown (GP) in L3 and L4 larvae of A. simplex . This study also tests the hypothesis formulated in our previous work [ 25 ], namely, that changes in host environments accompanied by metabolic adaptations associated with changes in enzyme activities probably also induce changes at the genetic level.…”

“…Different letters (a, b, and c) above the bars represent significant differences between L3 (control) and L4 of A. simplex for gene expression and enzyme activity. ¥ Data from Łopieńska-Biernat et al [ 25 ], r = 0.95 for enzymes and mRNA of trehalose metabolism, r = −0.8 for glycogen metabolism in L4 of A. simplex ( P < 0.005) (as control 100% activity of enzymes was 0.43 ± 0.02 U/mg for TPS and 0.34 ± 0.03 for TPP U/mg in L3).…”

“…The properties of TPS and TPP have been described in stage L3 larvae of A. simplex [ 24 ]. The activity of GPa and b has been studied in A. simplex [ 25 ], but mRNA levels have not been investigated to date. The synthesis of trehalose and glycogen protects developing parasites against external stressors and enables them to survive in harsh conditions [ 19 , 26 ].…”

Histopathological changes in the stomach wall of the European grayling Thymallus thymallus associated with severe infection by the nematode Cystidicoloides ephemeridarum

“…2). Łopieńska-Biernat et al (11) suggest the metabolic differences were confirmed by the data of protein analysis in extract of L3 and L4 Anisakis simplex. The total protein pool showed both quantitative and qualitative differences reflected in the presence of five stage-specific fractions in extracts from larval stages.…”