Elsevier

Acta Tropica

Volume 208, August 2020, 105531
Acta Tropica

Morphological and molecular characterization of a new ciliate Nyctotheroides grimi n. sp. (Armophorea, Clevelandellida) from Chinese frogs

https://doi.org/10.1016/j.actatropica.2020.105531Get rights and content

Highlights

  • Nyctotheroides grimi n. sp. is described from the frog Fejervarya limnocharis

  • Genetic similarity between Nyctotheroides species suggest recent phylogenetic divergence

  • The morphological criteria differencing Nyctotherus and Nyctotheroides are revised

Abstract

A new species of clevelandellid ciliate, Nyctotheroides grimi n. sp., is described from the frog Fejervarya limnocharis. Light and scanning electron microscopy were used for the morphological studies, and the DNA encoding the SSU rRNA gene (SSU rDNA) and the ITS1-5.8S subunit rRNA-ITS2 region (ITS) were sequenced for genetic comparisons and phylogenetic analysis. The main distinctive morphological feature is a knob-like projection in the left-posterior end; other differential characters are the cell size, the length of the oral groove and the shape of the infundibulum. Nyctotheroides grimi possess an apical suture line in the left and right side of the anterior end and in the left side of the caudal end. In the phylogenetic analyses, the new species engroups with other Nyctotheroides species forming a monophyletic group. The high similarity in the SSU rDNA and ITS sequences between Nyctotheroides species suggests a relative recent divergence. The genetic data and the different host range support the separation of Nyctotheroides and Nyctotherus; however the morphological criterion based on the presence (in Nyctotheroides)/absence (in Nyctothterus) of an apical kinetal suture line should be modified to consider the presence of kinetal suture lines in the apical and/or the caudal left side in Nyctotheroides.

Introduction

The nyctotherids are a large group of clevelandellid ciliates (Armophorea: Clevelandellida), mainly inhabiting the digestive track of both invertebrates and vertebrates, including oligochaetes, insects (cockroach), myriapods (centipede, millipede), molluscs (shipworm), fishes, amphibians (frogs and toads) and reptiles (Albaret 1975; Jankowski 2007; Lynn 2008). They have been discovered thus far in marine, freshwater and terrestrial habitats as endocommensals of the fore-mentioned hosts.

The nyctotherids are classified into 15 genera within the family Nyctotheridae (Lynn 2008), being Nyctotheroides and Nyctotherus the most important ones. Nyctotheroides is restricted to amphibian hosts and Nyctotherus to invertebrates and non-amphibian vertebrates, such as reptiles (Albaret 1975; Lynn and Wright 2013). It is generally accepted that these genera can be distinguished by the apical kinetal suture systems (“système secant”): there are kinetal sutures on the right and left apical sides in Nyctotheroides, while Nyctotherus has a kinetal suture only on the right apical side (Albaret 1975; Affa'a et al. 2004). There are other suture systems in Nyctotheroides, but they are not present in all species and Affa'a (1980, 1988) proposed to subdivide the species of this genus into four lineages (cordiformis, judesi, anomalus and bisecatus).

There are more than 70 species described in Nyctotheroides (Albaret 1975; Edwards et al. 1982; Kalavati et al. 1991; Fokam et al. 2013) but to date, genetic data have been obtained from only five of them (van Hoek et al. 1998; Affa'a et al. 2004; Li et al. 2016, 2017). In the present study, we attempt to contribute to the knowledge of the genus Nyctotheroides by describing a new species discovered from the rectum of the frog Fejervarya limnocharis (=Rana limnocharis), based on both detailed morphological information and molecular data.

Section snippets

Host collection and ethical statement

The frogs (F. limnocharis) were captured from Diaocha Lake (30°24′-30°48′N; 113°42′-113°58′E), Hubei Province, China in May to August 2015. All the experimental procedures and animal care were performed according to the protocols approved by the Institutional Animal Care and Use Committee of the Institute of Hydrobiology, Chinese Academy of Sciences (PAOIHB12519, 5/2012). The frogs were transported alive to the laboratory for further examination. All frogs were euthanized with MS-222 and

Results

Based on our survey, 74 (22.8%) of 325 examined F. limnocharis were found to be infected with a new ciliate species described now. About 150 trophozoites/ml were found in average in the recta of examined frogs.

  • Species name: Nyctotheroides grimi n. sp.

  • Type Host: Fejervarya limnocharis Gravenhorst, 1829

  • Type Locality: Diaocha Lake (30°24′-30°48′N; 113°42′-113°58′E), Hubei Province, China.

  • Infection site: Rectum.

  • Type material: Protargol stained specimens have been deposited in the Key Laboratory of

Discussion

The occurrence of a new Nyctotheroides species inhabiting Chinese anuran amphibians is recorded herein, with its morphological and genetic data presented in detail. Nyctotheroides grimi is quite unique considering its noteworthy knob-like projection at left-posterior end, which can distinguish it from almost all other Nyctotheroides species. Nictotheroides pyriformis could also present a knob-like projection at the posterior end in some cells, but much less significant than that of N. grimi;

Conclusions

A new species of clevelandellid ciliate, Nyctotheroides grimi, is described from the rectum of the frog Fejervarya limnocharis. It has clear morphological differences with related species, but the genetic similarity in the SSU rDNA and the ITS region indicates a recent phylogenetic divergence. There are no differential genetic features (such as indels, sequence motifs, or extra helices in the secondary structure) in the SSU rDNA or in the ITS region that could be useful for genus barcoding in

CRediT authorship contribution statement

Ming Li: Conceptualization, Investigation, Methodology, Data curation, Formal analysis, Validation, Writing - original draft, Writing - review & editing. Guangran Hu: Investigation, Methodology. Can Li: Investigation, Methodology. Wei-shan Zhao: Investigation, Methodology. Hong Zou: Formal analysis, Validation. Wen-xiang Li: Formal analysis, Validation. Shan-gong Wu: Formal analysis, Validation. Gui-tang Wang: Funding acquisition, Resources, Supervision. Francisco Ponce-Gordo: Data curation,

Declaration of Competing Interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Acknowledgements

This work was supported by the National Natural Science Foundation of China (No. 31772429, 31471978), the Youth Innovation Promotion Association CAS (No. Y82Z01) and the earmarked fund for China Agriculture Research System (No. CARS-45-15).

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