RNA-Seq de novo assembly and differential transcriptome analysis of the nematode Ascaridia galli in relation to in vivo exposure to flubendazole

Abstract

The nematode Ascaridia galli (order Ascaridida) is an economically important intestinal parasite responsible for increased food consumption, reduced performance and elevated mortality in commercial poultry production. This roundworm is an emerging problem in several European countries on farms with laying hens, as a consequence of the recent European Union (EU) ban on conventional battery cages. As infection is associated with slow development of low levels of acquired protective immunity, parasite control relies on repeated use of dewormers (anthelmintics). Benzimidazoles (BZ) are currently the only anthelmintic registered in the EU for use in controlling A. galli and there is an obvious risk of overuse of one drug class, selecting for resistance. Thus we developed a reference transcriptome of A. galli to investigate the response in gene expression before and after exposure to the BZ drug flubendazole (FLBZ). Transcriptional variations between treated and untreated A. galli showed that transcripts annotated as mitochondrial glutamate dehydrogenase and cytochrome P450 were significantly down-regulated in treated worms, whereas transcripts homologous to heat shock proteins (HSP), catalase, phosphofructokinase, and a multidrug resistance P-glycoprotein (PGP1) were significantly up-regulated in treated worms. Investigation of candidate transcripts responsible for anthelmintic resistance in livestock nematodes led to identification of several tubulins, including six new isoforms of beta-tubulin, and several ligand-gated ionotropic receptors and ABC-transporters. We discovered several transcripts associated with drug binding and processing genes, but further characterisation using a larger set of worms exposed to BZs in functional assays is required to determine how these are involved in drug binding and metabolism.

Transcriptional responses in Parascaris univalens after in vitro exposure to ivermectin, pyrantel citrate and thiabendazole

“…So far only two β-tubulin isotypes (isotype 1 and 2) have been described in Parascaris spp. [48], though more might be present in the genome since six putative β-tubulin genes were found in the closely related parasite Ascaridia galli [49]. It has also been mentioned that the Ascaris suum genome contains at least nine β-tubulin genes [50].…”

“…It should be noted that the isoforms of ascarid β-tubulins should not be directly compared to the strongyle nematodes as phylogenetic studies have shown that they are evolutionary separated [48,49]. In a study by Martis et al [49], where the transcriptomes of A. galli were compared before and after in vivo exposure to flubendazole, Fig. 4 Venn diagrams showing the number of differentially expressed genes unique to and shared among Parascaris univalens worms exposed to three different concentrations of the anthelmintic drugs ivermectin (a), pyrantel citrate (b) and thiabendazole (c).…”

“…It has also been mentioned that the Ascaris suum genome contains at least nine β-tubulin genes [ 50 ]. It should be noted that the isoforms of ascarid β-tubulins should not be directly compared to the strongyle nematodes as phylogenetic studies have shown that they are evolutionary separated [ 48 , 49 ]. In a study by Martis et al [ 49 ], where the transcriptomes of A. galli were compared before and after in vivo exposure to flubendazole, no differential expression of β-tubulins was found, whereas a study by Tyden et al [ 51 ] showed an upregulation of β-tubulin isotype 1 in Parascaris spp.…”

Transcriptome Profile Changes Associated With Heat Shock Reaction in the Entomopathogenic Nematode, Steinernema carpocapsae

“…In nematodes, toxic chemicals activate the immune response and can change their fecundity and life span. RNA-Seq analyses demonstrated that some differentially expressed genes (DEGs) were related to the immune response ( Martis et al, 2017 ; Di et al, 2018 ). Changes of gene expression, in different life cycles, were detected by transcriptome sequencing ( Seiml-Buchinger and Matveeva, 2019 ).…”

Transcriptional responses in Parascaris univalens after in vitro exposure to ivermectin, pyrantel citrate and thiabendazole

“…It has also been mentioned that the Ascaris suum genome contains at least nine β-tubulin genes (50). It should be noted that the isoforms of ascarid β-tubulins should not be directly compared to the strongyle nematodes as phylogenetic studies have shown that they are evolutionary separated (48,49). In a study by Martis et al (49) where the transcriptomes of A. galli were compared before and after in vivo exposure to flubendazole, no differential expression of β-tubulins was found, whereas a study by Tyden et al (51) showed an upregulation of β-tubulin isotype 1 in Parascaris spp.…”