Phylogenetic MLSA and phenotypic analysis identification of three probable novel Pseudomonas species isolated on King George Island, South Shetland, Antarctica

Abstract

Antarctica harbors a great diversity of microorganisms, including bacteria, archaea, microalgae and yeasts. The Pseudomonas genus is one of the most diverse and successful bacterial groups described to date, but only eight species isolated from Antarctica have been characterized. Here, we present three potentially novel species isolated on King George Island. The most abundant isolates from four different environments, were genotypically and phenotypically characterized. Multilocus sequence analysis and 16S rRNA gene analysis of a sequence concatenate for six genes (16S, aroE, glnS, gyrB, ileS and rpoD), determined one of the isolates to be a new Pseudomonas mandelii strain, while the other three are good candidates for new Pseudomonas species. Additionally, genotype analyses showed the three candidates to be part of a new subgroup within the Pseudomonas fluorescens complex, together with the Antarctic species Pseudomonas antarctica and Pseudomonas extremaustralis. We propose terming this new subgroup P. antarctica. Likewise, phenotypic analyses using API 20 NE and BIOLOG® corroborated the genotyping results, confirming that all presented isolates form part of the P. fluorescens complex. Pseudomonas genus research on the Antarctic continent is in its infancy. To understand these microorganisms’ role in this extreme environment, the characterization and description of new species is vital.

Pseudomonas fildesensis sp. nov., a psychrotolerant bacterium isolated from Antarctic soil of King George Island, South Shetland Islands

“…S3). This data corroborates previously presented results with other isolates of members of the genus Pseudomonas from Antarctica [25] and it does not allow determination of closely related species and proposal of a clear classification of the strain within any of the taxonomic groups of the genus Pseudomonas established by Bennasar et al [22]. However, from the phylogenetic reconstruction based on the multilocus sequences, it was possible to clarify that KG01 T forms part of a monophyletic cluster, which is robust and restricted to P. veronii LMG 17761 T and P. extremaustralis DSM 17835 T .…”

“…Moreover, KG01 T is positive for catalase and oxidase activities and secretes fluorescent green pigments when it is grown for 36-48 h at 25 °C on King B agar. The ability of KG01 T to grow at different temperatures (4,15,20,25,30,35 and 37 °C) was tested in 250 ml Erlenmeyer flasks containing 50 ml LB. Tolerance to different NaCl concentrations (0-10 % w/v at increments of 1.0 % w/v) was evaluated in LA plates after incubation for 3 days at 25 °C.…”

Genetic and Phenotypic Characterization of Indole-Producing Isolates of Pseudomonas syringae pv. actinidiae Obtained From Chilean Kiwifruit Orchards

“…Escherichia coli DH5α, E. coli K12, Pseudomonas aeruginosa PAO1, Azospirillum brasilense SP7, Salmonella bongori X9617, and Cupriavidus metallidurans CH34 were obtained from the bacterial collection of the Laboratory of Microbiology of the Pontificia Universidad Católica de Valparaíso (PUCV). Pseudomonas antarctica S63 ( Vásquez-Ponce et al, 2018 ) was provided by Dr. Jorge Olivares from the PUCV. The bacteria were grown at 25°C in Luria-Bertani (LB) medium except when another medium is specified.…”

Comparative Genomic Analysis of Soil Dwelling Bacteria Utilizing a Combinational Codon Usage and Molecular Phylogenetic Approach Accentuating on Key Housekeeping Genes

“…To focus on bacterial phylogeny, ribosomal RNA is often envisaged as one of the best parameter due to its ubiquitous and informative nature (Morales et al, 2009) but it's not without its pitfalls. Phylogenetic analysis based on the 16S rRNA gene is generally used to articulate the evolutionary history (Rajendhran and Gunasekaran, 2011) but, several discrepancies have been observed in 16S rRNA based phylogeny (Vásquez-Ponce et al, 2018). Some of the major drawbacks include mosaicism due to horizontal gene transfer and recombination events (Eardly et al, 1996;Schouls et al, 2003), presence of polymorphic genes (Janda and Abbott, 2007;Lang et al, 2013), intra-genomic heterogeneity of the 16S rRNA genes, presence of multiple copies of the rRNA operon (Wang et al, 1997;Klappenbach et al, 2000), etc.…”

“…Therefore, it is a very resourceful proposition to compare the sequences of several housekeeping genes as well as the 16S rRNA genes portraying as a reference for evaluating a comprehensive bacterial phylogeny, and also for investigating the genetic as well as physiological relatedness within the species. A technique which combines analysis of several housekeeping genes called multilocus sequence analysis (MLSA) has emerged as a powerful and pragmatic molecular method to assess pertinent phylogenetic information in the field of polyphasic taxonomy (Das et al, 2014;Vásquez-Ponce et al, 2018). Several studies have been carried out using MLSA in order to get better resolution regarding the phylogenetic analysis and inter-species relatedness in a host of soil bacterial genera like Arthrobacter (Liu et al, 2018), Borrelia (Margos et al, 2009), Burkholderia (Estrada-De Los Santos et al, 2013), Ensifer (Martens et al, 2008), Micromonospora (Carro et al, 2012), Streptomyces (Rong and Huang, 2010), Nocardia (Barcellos et al, 2007;McTaggart et al, 2010;Serrano et al, 2010;Busse and Wieser, 2014), and some other rhizospheric bacteria like Mesorhizobium (Degefu et al, 2011) and Bradyrhizobium (Delamuta et al, 2012).…”