Unveiling the phylogeny of Contracaecum jorgei (Nematoda, Anisakidae) parasitizing Ardea cocoi (Aves, Ardeidae) in Argentina based on an integrative analysis

Abstract

Anisakidae nematodes of the Contracaecum genus are known for their wide host and geographic distribution. These parasites commonly infest aquatic organisms worldwide. The life cycles of Contracaecum species typically involve aquatic invertebrates and fish as intermediate and/or paratenic hosts, while piscivorous birds and mammals serve as definitive hosts. The aim of the present work is to identify the Contracaecumspecimens parasitizing Ardea cocoifrom the Magdalena city coast, Argentina, and to determine their phylogenetic relationships with its congeners based on both morphometric and phylogenetic analyses. One Cocoi Heron specimen was found dead at the Magdalena coast, Buenos Aires Province, and 53 nematodes were recovered from its esophagus and stomach. Some nematode specimens were cleared with lactophenol and studied using an optical microscope. Three males and females were examined in a scanning electron microscope. DNA extraction, PCR and sequencing of mtDNA cox2, ITS1, ITS2, and SSrRNA genes were performed from three Contracaecum specimens. Both morphometric and phylogenetic analyses of the present studied specimens allowed us to recognize the species Contracaecum jorgei. In the constructed tree using SSrRNA sequences, a node aligns with C. microcephalum sequences. Conversely, ITS1 and ITS2 sequences respectively, establish a robust node positioning our sequence as the sister species to a node consisting of Contracaecum pyripapillatum, C. multipapillatum, C. gibsoni, and C. quadripapillatum. The mtDNA cox2gene shows our sequences exhibiting concordance with C. jorgei. The p-distances calculated for the SSrRNA gene matrix displayed a distance of 14% from C. microcephalum and the present study specimens (PES). For ITS1, the calculated distance is 16% from both C. pyripapillatum and C. multipapillatum. In the case of ITS2, the distances are 35% from C. multipapillatum, and 38% from C. pyripapillatum. Finally, the mtDNA cox2gene displayed a distance of 1% observed for among our sequences and C. jorgei. According to the molecular analysis, PES clustered with the sister species C. microcephalum on the SSrRNA BI tree. Considering the BI analysis of ITS1-ITS2 genes, PES appear as a sister species to the node consisting of C. multipapillatum, C. pyripapillatum, C. quadripapillatum, and C. gibsoni. Finally, the topology of the mtDNA cox2 BI tree and the genetic distances between Contracaecumtaxa supports the identification of the PES as C. jorgei. Contracaeum jorgei is closely related to C. multipapillatum. The record of C. jorgei parasitizing A. coccoi is the second report of an Anisakidae for this host species in Argentina, and also for any Ardeidae. Integrative molecular studies including morphological and molecular tools are important to know the real host and geographical distribution of parasite diversity and establish specific correspondences to determine phylogenetic relationships on the Contracaecum species. This work represents the second report of C. jorgei from Argentina based on morphological analysis conducted using optical and scanning electron microscopy. This study shed light on the limited information available regarding this conspicuous nematode and sets the stage for further investigations into its life cycles.