Yams are important tuber crops in the tropics. Nitrogen is one of the limiting factors in yam production. Yam accessions that can rely on biological nitrogen fixation must be identified if we are to achieve efficient and sustainable yam production. The aim of this study was to isolate and identify the endophytic diazotrophic bacteria of 19 accessions of two yam species. Forty-one endophytic bacteria were isolated from surface-sterilized roots, stems and leaves of yam accessions grown in nutrient-poor subsoil for 160 days. The 16S rRNA gene sequencing showed that the 41 isolates were grouped into 18 genera including Bacillus, Bradyrhizobium, Devosia, Ensifer, Mycobacterium, Neorhizobium, Paenibacillus, Pseudoxanthomonas, Rhizobium and Xanthomonas. Most endophytes showed a positive response to nitrogenase activity and were found in the stems (21) and the roots (14), while only six were found in the leaves. In Dioscorea esculenta, the acetylene reduction assay (ARA) values ranged from 4.1 to 57.4 nmol tube/24 h and from 4.1 to 164 nmol tube/24 h in Dioscorea alata. In addition, the endophyte species were different among yam species and among yam accessions and were affected by fertilization. These results indicate that the endophytic diazotrophic bacteria of yams are diverse.

Two strains, NIES-2215 and NIES-2243, have been labeled “Chlamydomonas (Cd.) gerloffii” and “Cd. subangulosa,” respectively. Although these two strains were previously suggested to belong to the Reticulata group of the genus Chloromonas (Cr.), detailed taxonomic examinations are lacking for both strains. Here we taxonomically re-examined the two strains based on light and transmission electron microscopy and molecular phylogenetic analysis using a combined dataset from 18S and 28S ribosomal RNA gene sequences and genes encoding ATP synthase β-subunit, P700 chlorophyll a-apoprotein A1 and P700 chlorophyll a-apoprotein A2. The “Cd. gerloffii” strain NIES-2215 differed from all other species of the Reticulata group in terms of phylogeny and ultrastructure of the pyrenoid and eyespot; it was morphologically identified as Cd. gerloffii. Thus, we propose Cr. difformis Tomo. Makino, Matsuzaki & Nozaki nom. nov., to encompass Cd. gerloffii, because of presence of the previous valid description of Cr. gerloffii for a different species. The “Cd. subangulosa” strain NIES-2243 was significantly different in morphology from the original description of Cd. subangulosa. It was identified as Cr. typhlos because NIES-2243 and Cr. typhlos strain SAG 26.86 formed a small robust clade with essentially the same morphological characteristics except for the presence or absence of an eyespot in the chloroplast.