- Volume 66, Issue 12, 2016

Coccoid green algae are extremely diverse despite their simple coccoid phenotype, a phenotype that may be the result of convergent evolution. In this study, we used a polyphasic approach combining molecular phylogenetic analyses, morphology and ultrastructure to investigate isolated coccoid strains from China, and our results reveal three new lineages of Trebouxiophyceae: the novel genus and species Mysteriochloris nanningensis gen. et sp. nov., and the two novel species Phyllosiphon coccidium sp. nov. and Desertella yichangensis sp. nov. (Trebouxiophyceae, Chlorophyta). We provide a detailed characterization of the novel microalgae which they are autosporic coccoid unicells and have parietal chloroplasts. In phylogenies based on 18S rDNA sequences and the chloroplast ribulose-bisphosphate carboxylase gene (rbcL), these three algae are nested within the Watanabea clade and are different from any known algae. M. nanningensis FACHB-1787 is not really close to any known algae within the Watanabea clade. Phyllosiphoncoccidium FACHB-2212 is within the Phyllosiphon lineages. D. yichangensis FACHB-1793 is closely related to Desertella californica and described as a representative of a novel species of the genus Desertella.

A polyphasic taxonomic approach was applied to strains of the species Staphylococcus sciuri in order to clarify the taxonomic legitimacy of the delineation of S. sciuri into S. sciuri subsp. sciuri , S. sciuri subsp. carnaticus and S. sciuri subsp. rodentium . A group of 81 S. sciuri isolates obtained from human (n=62) and veterinary (n=17) clinical materials and foods (n=2) and ten reference and type strains obtained from the Czech Collection of Microorganisms were characterized by extensive biotyping using conventional tests and commercial identification kits (ID 32 Staph, STAPHYtest, Biolog Microbial ID System), matrix-assisted laser-desorption/ionization time-of-flight mass spectrometry, automated ribotyping with EcoRI restriction enzyme, 16S–23S rRNA gene intergenic transcribed spacer PCR fingerprinting and repetitive sequence-based PCR fingerprinting with the (GTG)5 primer. Selected strains representing different ribotypes were further characterized using sequencing of the β-subunit of RNA polymerase (rpoB) gene. Individual techniques revealed high heterogeneity within the analysed S. sciuri strains but differentiation of the investigated strains into groups corresponding to the aforementioned S. sciuri subspecies and supported by these techniques was not clearly revealed. Based on obtained results and data retrieved from literature we propose rejecting the separation of S. sciuri species into S. sciuri subsp. sciuri , S. sciuri subsp. carnaticus and S. sciuri subsp. rodentium and we suggest reclassification these subspecies as S. sciuri with the type strain W.E. Kloos SC 116T (=ATCC 29062T=BCRC 12927T=CCM 3473T=CCUG 15598T=CNCTC 5683T=DSM 20345T=JCM 2425T=NCTC 12103T).

Phytoplasmas are unculturable, phytopathogenic bacteria that cause economic losses worldwide. As unculturable micro-organisms, phytoplasma taxonomy has been based on the use of the 16S rRNA-encoding gene to establish 16Sr groups and subgroups based on the restriction fragment length polymorphism (RFLP) pattern resulting from the digestion of amplicon (in vitro) or sequence (in silico) with seventeen restriction enzymes. Problems such as heterogeneity of the ribosomal operon and the inability to differentiate closely related phytoplasma strains has motivated the search for additional markers capable of providing finer differentiation of phytoplasma strains. In this study we developed and validated a scheme to classify phytoplasmas based on the use of cpn60 universal target (cpn60 UT) sequences. Ninety-six cpn60 UT sequences from strains belonging to 19 16Sr subgroups were subjected to in silico RFLP using pDRAW32 software, resulting in 25 distinctive RFLP profiles. Based on these results we delineated cpn60 UT groups and subgroups, and established a threshold similarity coefficient for groups and subgroups classifying all the strains analysed in this study. The nucleotide identity among the reference strains, the correspondence between in vitro and in silico RFLP, and the phylogenetic relationships of phytoplasma strains based on cpn60 UT sequences are also discussed.

Two novel species names, Acinetobacter bohemicus and Acinetobacter pakistanensis , appeared on validation list no. 161 (January 2015) under priority numbers 26 and 28, respectively. As the published data suggested a high similarity of the organisms associated with these names, we aimed to define their taxonomic relationship. The study set included all strains used in the original nomenclatural proposals, i.e. 25 strains of A. bohemicus and one strain of A. pakistanensis . The average nucleotide identity values (95.9 and 96.1 % based on blast and MUMmer, respectively) between the whole-genome sequences of A. bohemicus ANC 3994T and A. pakistanensis KCTC 42081T supported the identity of these type strains at the species level. Based on the genus-wide comparative analyses of the rpoB sequences and whole-cell fingerprints generated by matrix-assisted laser desorption/ionization-time-of-flight MS, A. pakistanensis KCTC 42081T fell within the respective clusters formed by the 25 A . bohemicus strains. The same picture was obtained on the basis of comparative analysis of 16S rRNA gene sequences of KCTC 42081T and three A. bohemicus strains. Finally, the metabolic and physiological features of KCTC 42081T were found to be congruent with those of A. bohemicus . Based on these results, we conclude that Acinetobacter pakistanensis is a later heterotypic synonym of Acinetobacter bohemicus.