Species Diversity Of Interstitial Ciliates In Brackish Water Off The Coast Of Yantai

Interstitial ciliates are kinds of benthic protozoa living in the sand grains,and they are important functional groups in the benthic microbial food web.However,due to its high species diversity and difficulty in identification,there is still a lot of blank in the study of morphology and taxonomy of interstitial ciliates.From September 2016 to October 2018,the species diversity of interstitial ciliates in brackish water off the coast of Yantai,was studied in detail.A total of 24 species of interstitial ciliates were identified,belonging to 6 class,9 order,14 families and 18 genera(Phialina,Lacrymaria,Helicoprorodon,Trachelotractus,Paraspathidium,Tracheloraphis,Kovalevaia,Zosterodasys,Dysteria,Loxophyllum,Cardiostomatella,Pleuronema,Condylostoma,Peritromus,Uronychia,Diophrys,Aspidisca,Euplotes),respectively.The main results are as follows:1.Establishment of novel species and novel combination.A novel species and a new combination were established based on live observations,protargol staining method and SSU rRNA gene sequences.The morphological diagnoses are described below:(1)Phialina clampi Wang et al.,2019: Extended cells about 80~300 × 18~50 ?m in vivo;body cylindrical;head carrying oral bulge,distinctly separated from trunk by a constriction in fully extended cell;macronucleus usually ovoidal,situated in mid-body;contractile vacuole located caudally;two types of extrusomes: type I about 20~35 × 1~4 ?m in size,type II 1.5~3.0 × 0.7~1.0 ?m in size and attached to somatic cortex;27~31 somatic kineties,each composed of 2~4 dorsal brush anteriorly and 71~163 somatic monokinetids posteriorly.(2)Phialina caudata(Kahl,1933)Wang et al.,2019: Extended cells about 80~170 × 20~45 ?m in vivo;body cylindrical;outline radish-shaped with a tail-like posterior end;head carrying conical oral bulge,distinctly separated from trunk in fully extended cell;macronucleus usually ovoidal,situated in mid-body;micronucleus ovoidal or globular,near or attached to macronucleus;contractile vacuole located subterminally between mid-body and base of tail;20~24 somatic kineties,each composed of 2~4 dorsal brush anteriorly and 34~74 somatic monokinetids posteriorly.2.The SSU rRNA gene sequences of Phialina clampi Wang et al.,2019 and Phialina caudata(Kahl,1933)Wang et al.,2019 were sequenced for the first time.The phylogenetic trees were constructed by maximum likelihood(ML)analyses and Bayesian inference(BI),and then the systematic positions of two species was evaluated.The main conclusions are as follows:(1)The systematic positions of Phialina clampi and Phialina caudata was confirmed.In the phylogenetic trees,P.caudata clusters with Phialina sp.before meeting with P.clampi,and occupies the basal position within the family Lacrymariidae.(2)Phylogenetic analyses based on SSU rRNA gene sequences indicate that the genus Phialina is non-monophyletic.(3)Based on the morphological features and phylogeny of the genera Phialina and Lacrymaria,it is speculated that the non-contractile “neck” in Phialina should be an ancestral character state,whereas the contractile “neck” in Lacrymaria should be a derived character state.3.An optimized protocol of protargol stainingThe Wilbert protargol staining method,a key technology for ciliate morphology and taxonomy research,was optimized to improve the success rate of staining and optimize the staining effect.