Phylogenetic Affinities of Uvulifer Spp. (Digenea: Diplostomidae) in the Americas with Description of Two New Species from Peruvian Amazon

Abstract

Uvulifer Yamaguti, 1934, is a genus of diplostomoidean digeneans that parasitizes kingfishers worldwide. Species have a Neascus-type metacercaria that encysts in or on fish intermediate hosts, often causing black spot disease. Only 3 prior studies published DNA sequence data for Uvulifer species with only 1 including a single named species (Uvulifer spinatus López-Jiménez, Pérez-Ponce de León, & García-Varela, 2018). Herein we describe 2 new species of Uvulifer from the green-and-rufous kingfisher, Chloroceryle inda (Linnaeus), collected in Peru ( Uvulifer batesi n. sp. and Uvulifer pequenae n. sp.). Both new species are readily differentiated from their New World congeners by a combination of morphological characters including distribution of vitelline follicles and prosoma:opisthosoma length ratios. In addition, we used newly generated nuclear 28S rRNA and mitochondrial COI gene sequence data to differentiate among species and examine phylogenetic affinities of Uvulifer. This includes the 2 new species and Uvulifer ambloplitis (Hughes, 1927), as well as Uvulifer elongatus Dubois, 1988 , Uvulifer prosocotyle (Lutz, 1928), and Uvulifer weberi Dubois, 1985 , none of which have been part of prior molecular phylogenetic studies. Our data on Uvulifer revealed 0.1-2.2% interspecific divergence in 28S sequences and 9.3-15.3% in COI sequences. Our 28S phylogeny revealed at least 6 well-supported clades within the genus. In contrast, the branch topology in the COI phylogenetic tree was overall less supported, indicating that although COI sequences are a great tool for species differentiation, they should be used with caution for phylogenetic inference at higher taxonomic levels. Our 28S phylogeny did not reveal any clear patterns of host association between Uvulifer and particular species of kingfishers; however, it identified 2 well-supported clades uniting Uvulifer species from distant geographical locations and more than 1 biogeographic realm, indicating at least 2 independent dispersal events in the evolutionary history of the New World Uvulifer. Our results clearly demonstrate that the diversity of Uvulifer in the New World has been underestimated.

Keywords: Diplostomidae; Uvulifer batesi n. sp .; Uvulifer pequenae n. sp .; Amazon; Brazil; Chloroceryle inda; Kingfishers; Molecular Phylogeny; Peru; Uvulifer ambloplitis.

Figures 1, 2.

Uvulifer pequenae n. sp. (1) Ventral view of whole mount. Scale bar = 300 μm. (2) Ventral view of posterior body end. Scale bar = 100 μm. Abbreviations: c, ceca; cb, copulatory bursa; eg, egg; ep, ejaculatory pouch; ev, excretory vesicle; gc, genital cone; Mg, Mehlis' gland; o, ovary; pf, preputial fold; sv, seminal vesicle; t, testis; to, tribocytic organ; v, vitelline follicle; vr, vitelline reservoir.

Figures 3, 4.

Uvulifer batesi n. sp. (3) Ventral view of holotype. Scale bar 250 μm. (4) Ventral view of posterior body end of holotype with uterus omitted. Scale bar = 150 μm. Abbreviations: c, ceca; cb, copulatory bursa; eg, egg; ep, ejaculatory pouch; ev, excretory vesicle; gc, genital cone; Mg, Mehlis' gland; o, ovary; pf, preputial fold; sv, seminal vesicle; t, testis; to, tribocytic organ; v, vitelline follicle; vr, vitelline reservoir.

Figure 5.

Phylogenetic interrelationships among 14 Uvulifer taxa based on Bayesian Inference (BI) analysis of partial 28S rRNA gene sequences. Bayesian Inference posterior probability values lower than 70% (BI) are not shown. New sequences obtained in this study are in bold. Branch length scale bar indicates number of substitutions per site. GenBank accession numbers and the biogeographical realm, and geographic origin are provided after the names of species. Abbreviations for biogeographical realms: AF = Afrotropical realm, NA = Nearctic realm, NT = Neotropical realm. Abbreviations for geographic origin: BR La = Lábrea site in Brazil, BR Pa = Pantanal site in Brazil, MX = Mexico, PE = Peru, SA = South Africa, USA = United States of America.

Figure 6.

Phylogenetic interrelationships among 8 Uvulifer taxa based on Bayesian Inference (BI) analysis of partial COI mtDNA sequences. Bayesian Inference posterior probability values lower than 70% (BI) are not shown. New sequences obtained in this study are in bold. Branch length scale bar indicates number of substitutions per site. GenBank accession numbers, the biogeographical realm, and the geographic origin are provided after the names of species. Abbreviations for biogeographical realms: NA = Nearctic realm, NT = Neotropical realm. Abbreviations for geographic origin: BR Pa = Pantanal site in Brazil, CA = Canada, PE = Peru, USA = United States of America.