The molecular profile of Paratrajectura longcementglandatus Amin, Heckmann et Ali, 2018 (Acanthocephala: Transvenidae) from percid fishes in the marine waters of Iran and Iraq

Pichelin & Cribb (2001) described the family Transvenidae with two genera: monotypic Transvena with T. annulospinosaPichelin et Cribb, 2001, and Trajectura with two species, T. ikedai (Machida, 1992) and T. perinsolensPichelin et Cribb, 2001. Specimens of the two genera were recovered from wrasses (Labridae, Perciformes) in the Pacific off southern Australia and southern Japan. Lisitsyna et al. (2019) described two other species of the family Transvenidae, namely Transvena pichelinaeLisitsyna, Kudlai, Cribb et Smit, 2019, and Pararhadinorhynchus sodwanensisLisitsyna, Kudlai, Cribb et Smit, 2019 from the marine fishes from the Sodwana Bay, South Africa. The other genus of this family, Paratrajectura, was established by Amin et al. (2018). It comprises one species Paratrajectura longcementglandatus Amin, Heckmann et Ali, 2018, which was described on the basis of worms from the Japanese threadfin bream Nemipterus japonicus Bloch (Nemipteridae) and the tigertooth croacker, Otolithes ruber Bloch et Schneider (Sciaenidae, Perciformes) caught in the marine territorial waters of Iraq and Iran, the Persian Gulf (Amin et al., 2018). The genus Paratrajectura is characterised by having apical proboscis cone, long, tubular cement glands, short lemnisci, prominent roots on all proboscis hooks, subterminal female gonopore, and males with long pre-equatorial testes.

While, several studies have been published about sequence data for acanthocephalans including two Transvena spp. (Westram et al., 2011; Garcia-Varela, et al., 2013; Pinacho-Pinacho et al., 2014; Lisitsyna et al., 2019), no sequence data has been published for P. longcementglandatus whose phylogenetic relationship with other acanthocephalans and related families was unknown. In this paper, we report the molecular profile of P. longcementglandatus, validate its generic affiliations, and explore its evolutionary relationships with related and other species and taxa based on partial 18S rDNA and cox1 genes.

The authors declare compliance with all relevant ethical standards.

The specimens of P. longcementglandatus successfully presented amplifications of about 1234 bp for the 18S rDNA gene and 664 bp for the cox1 gene. Comparisons of the18S rDNA and cox1 sequences from this parasite with other available acanthocephalan sequences in GenBank, using multiple sequence alignment, showed that it had the highest similarity with T. annulospinosa based on18S rDNA (98 %) and cox1 (77 %) genes. The 18S rDNA dataset (1129 nt) included 26 sequences for species of seven families within the Echinorhynchida and the novel sequence of P. longcementglandatus. The cox1 dataset (538 nt) included 39 sequences for species of nine families of Echinorhynchida and the sequence of P. longcementglandatus.

The phylogenetic reconstruction based on the partial sequence spanning the 18S rDNA showed that our sequence of P. longcementglandatus is clustered with Transvena annulospinosa (AY830153), T. pichelinae (MN105736 and MN105737), P. sodwanensis (MN105738) and an unidentified species of Pararhadinorhynchus (HM545903) with strong support forming a clade of the family Transvenidae. Also, the species of Gymnorhadinorhynchus sp. (MK014866) (Gymnorhadinorhynchidae) and Rhadinorhynchus laterospinosus (MK457183) (Rhadinorhynchidae) are very closely related with the family Transvenidae in the tree with 100 % of bootstrap support. The sequence of Gymnorhadinorhynchus decapteri (KJ590123) (Gymnorhadinorhynchidae) is located at the basal position to the members of the clade. Other families of the order Echinorhynchida including Rhadinorhynchidae, Pomphorhynchidae, Cavisomidae, Arhythmacanthidae and Echinorhynchidae located in a major sister clade (Fig. 1). Inter-generic differences are noted between P. longcementglandatus and T. annulospinosa, T. pichelinae, P. sodwanensis and Pararhadinorhynchus sp. from Transvenidae based on partial 18S rDNA sequence were 2.4 % (18 nt), 2.8 % (21 nt), 2.9 % (22 nt) and 2.9 % (22 nt), respectively. According to phylogenetic analyses based on the cox1 gene, our sequence of P. longcementglandatus (MK770615) is grouped with T. annulospinosa (DQ089711) and T. pichelinae (MN104895 and MN104896) with strong support in a clade of the family Transvenidae. The species of G. decapteri (KJ590125) and Gymnorhadinorhynchus sp. (MK012665) (Gymnorhadinorhynchidae), Neorhadinorhynchus nudus (MG757444) (Cavisomidae), R. laterospinosus (MK572744) and Rhadinorhynchus sp. (DQ089712) (Rhadinorhynchidae), appear as a sister group of the family Transvenidae (Fig. 2). The interspecific divergence between P. longcementglandatus and T. annulospinosa, T. pichelinae based on partial cox1 gene was 23.4 % (141 nt), 27.3 % (144 nt), respectively.

Fig. 1

Fig. 2

Recently, molecular methods are applied for species identification, classification and phylogenetic analysis of acanthocephalan species (García-Varela et al., 2002). To date, molecular profile has been provided for few species of the family Transvenidae including T. annulospinosa, T. pichelinae, P. sodwanensis and Pararhadinorhynchus sp. (Pichelin & Cribb, 2001; Lisitsyna et al., 2019). In the current study, phylogenetic relationships of P. longcementglandatus as another genus of this family is described based on partial 18S rDNA and cox1 genes determining relationships with other acanthocephalan families.

This study showed that the interspecific variation between P. longcementglandatus and species of Transvena based on partial 18S rDNA was 2.4 % – 2.8 % (18 – 21 nt) and between it and species of Pararhadinorhynchus was 2.9 % (22 nt). Also based on cox1 gene, inter-generic variations between P. longcementglandatus and T. annulospinosa was 23.4 % (141 nt). These results illustrate that sequence differences between the genera of the family based on cox1 gene is higher than18S rDNA and it is appropriate to consider for taxonomic studies at the generic level.

The phylogenetic analysis of the 18S rDNA sequence (Fig. 1) showed that P. longcementglandatus is grouped in a highly supported clade with T. annulospinosa (AY830153), T. pichelinae (MN105736 and MN105737), P. sodwanensis (MN105738) and Pararhadinorhynchus sp. (HM545903) forming a clade of the family Transvenidae. In the clade, the family Transvenidae grouped close to R. laterospinosus (MK457183) and G. decapteri (KJ590123) (Gymnorhadinorhynchidae). Our phylogenetic tree for 18S rDNA is similar to those of García-Varela et al. (2002) and Lisitsyna et al. (2019) where the family Transvenidae grouped close to different species of Rhadinorhynchus (Rhadinorhynchidae) and Gymnorhadinorhynchus (Gymnorhadinorhynchidae).

Our phylogenetic analysis of the cox1 gene (Fig. 2) confirmed that P. longcementglandatus is grouped with T. annulospinosa (DQ089711) and T. pichelinae (MN104895 and MN104896) which made the clade of the family Transvenidae with good statistical support. Also, the families Rhadinorhynchidae, Gymnorhadinorhynchidae and Cavisomidae appear as a sister group with the clade of family Transvenidae. Other families of Echinorhynchida such as Pomphorhynchidae, Echinorhynchidae, Cavisomidae, Illiosentidae, Rhadinorhynchidae, Gymnorhadinorhynchidae, Diplosentidae and Arhythmacanthidae are well separated in the later clade. In the present study, the higher level of variation in cox1 gene compared to the 18S rDNA gene provides better resolution of the relationships within closely related taxa. While Amin et al. (2019a) presented relationships in their analysis of Rhadinorhynchus based on cox1 sequences, it was not clearer than 18S rDNA due to the lack of sufficient sequences of this gene in GenBank.

One of the most commonly used molecular markers for classification of acanthocephalans is the small subunit from RNA ribosomal gene or 18S rRNA. This gene displays a slow evolution rate and is highly conserved. It was used to infer phylogenetic relationships among the major classes of Acanthocephala (García-Varela & Pérez-Ponce de León, 2015). Most of phylogenetic studies of acanthocephalans similar to this research showed that 18S rDNA sequences appear to be suitable marker for phylogenies among acanthocephalans (García-Varela et al., 2000; Near, 2002; Herlyn et al., 2003; Verweyen et al., 2011; Amin et al., 2019b). Also, cox1 gene is commonly used for phylogenetic studies and to recognize and establish species limits in acanthocephalans (Guillen-Hernández et al., 2008; Alcántar-Escalera et al., 2013; García-Varela, et al., 2013). The present study confirmed that this gene has high genetic diversity among genera of the family and other families of Echinorhynchida which would be more particularly useful for phylogenetic analysis.

Finally, the genetic data collected in the current study provide a better understanding of the taxonomic status of P. longcementglandatus. Sequence variations within the family Transvenidae and among other families of Echinorhynchida based on cox1 gene is higher than18S rDNA that can be useful for achieving a proper assessment of biodiversity. More sequence data from other geographical isolates using more gene targets will be useful for exploring the phylogenetic relationships among species. On the other hand, using of molecular tools for identification of acanthocephalan species is still scarce due to the lack of sequences of different genera of acanthocephalans in GenBank (Amin et al., 2013; Salgado-Maldonado, 2013; Weaver & Smales, 2013; Amin et al., 2014; Smales, 2014; Gomes et al., 2015; Steinauer & Nickol, 2015). More molecular studies are recommended in order to elucidate acanthocephalans classification.