One Gram-stain-positive, non-motile, non-spore-forming, catalase-negative, and coccobacilli-shaped strain, designated c10Ua161M^T, was isolated from a urine sample from a reproductive-age healthy woman. Comparative 16S rRNA gene sequence analysis indicated that strain c10Ua161M^T belonged to the genus Lactobacillus . Phylogenetic analysis based on pheS and rpoA gene sequences strongly supported a clade encompassing strains c10Ua161M^T and eight other strains from public databases, distinct from currently recognized species of the genus Lactobacillus. In silico Average Nucleotide Identity (ANI) and Genome-to-Genome Distance Calculator (GGDC), showed 87.9 and 34.3 % identity to the closest relative Lactobacillus jensenii , respectively. The major fatty acids of strain c10Ua161M^T were C_18 : 1ω9c (65.0%), C_16 : 0 (17.8%), and summed feature 8 (10.2 %; comprising C_18 : 1ω7c, and/or C_18 : 1ω6c). The DNA G+C content of the strains is 34.2 mol%. On the basis of data presented here, strain c10Ua161M^T represents a novel species of the genus Lactobacillus , for which the name Lactobacillus mulieris sp. nov. is proposed. The type strain is c10Ua161M^T (=CECT 9755^T=DSM 108704^T).

For several years, knowledge on the microbiome associated with marine invertebrates was impaired by the challenges associated with the characterization of bacterial communities. With the advent of culture independent molecular tools it is possible to gain new insights on the diversity and richness of microorganisms associated with marine invertebrates. In the present study, we evaluated if different preservation and processing methodologies (prior to DNA extraction) can affect the bacterial diversity retrieved from snakelocks anemone Anemonia viridis. Denaturing gradient gel electrophoresis (DGGE) community fingerprints were used as proxy to determine the bacterial diversity retrieved (H[prime]). Statistical analyses indicated that preservation significantly affects H[prime]. The best approach to preserve and process A. viridis biomass for bacterial community fingerprint analysis was flash freezing in liquid nitrogen (preservation) followed by the use of a mechanical homogenizer (process), as it consistently yielded higher H[prime]. Alternatively, biomass samples can be processed fresh followed by cell lyses using a mechanical homogenizer or mortar & pestle. The suitability of employing these two alternative procedures was further reinforced by the quantification of the 16S rRNA gene; no significant differences were recorded when comparing these two approaches and the use of liquid nitrogen followed by processing with a mechanical homogenizer.

Two Gram-stain-positive strains, c9Ua_26_M^Tand c11Ua_112_M^T, were isolated from voided urine samples from two healthy women. Comparative 16S rRNA gene sequences demonstrated that these novel strains were members of the genusLimosilactobacillus. Phylogenetic analysis based onpheSgene sequences and core genomes showed that each strain formed a separated branch and are closest toLimosilactobacillus vaginalisDSM 5837^T. The average nucleotide identity (ANI) and Genome-to-Genome Distance Calculator (GGDC) values between c9Ua_26_M^Tand the closest relative DSM 5837^Twere 90.7 and 42.9 %, respectively. The ANI and GGDC values between c11Ua_112_M^Tand the closest relative DSM 5837^Twere 91.2 and 45.0 %, and those among the strains were 92.9% and 51,0 %, respectively. The major fatty acids were C_12 : 0(40.2 %), C_16 : 0(26.7 %) and C_18 : 1ω9c(17.7 %) for strain c9Ua_26_M^T, and C_18 : 1ω9c(38.0 %), C_16 : 0(33.3 %) and C_12 : 0(17.6 %) for strain c11Ua_112_M^T. The genomic DNA G+C content of strains c9Ua_26_M^Tand c11Ua_112_M^Twas 39.9 and 39.7 mol%, respectively. On the basis of the data presented here, strains c9Ua_26_M^Tand c11Ua_112_M^Trepresent two novel species of the genusLimosilactobacillus, for which the namesLimosilactobacillus urinaemulierissp. nov. (c9Ua_26_M^T=CECT 30144^T=LMG 31899^T) andLimosilactobacillus portuensissp. nov. (c11Ua_112_M^T=CECT 30145^T=LMG 31898^T) are proposed.