Development of Genomic Tools for the Identification of Certain Pseudomonas up to Species Level

Abstract

Pseudomonas is a highly versatile bacterium at the species level with great ecological significance. These genetically and metabolically diverse species have undergone repeated taxonomic revisions. We propose a strategy to identify Pseudomonas up to species level, based on the unique features of their 16S rDNA (rrs) gene sequence, such as the frame work of sequences, sequence motifs and restriction endonuclease (RE) digestion patterns. A species specific phylogenetic framework composed of 31 different rrs sequences, allowed us to segregate 1,367 out of 2,985 rrs sequences of this genus, which have been classified at present only up to genus (Pseudomonas) level, as follows: P. aeruginosa (219 sequences), P. fluorescens (463 sequences), P. putida (347 sequences), P. stutzeri (197 sequences), and P. syringae (141 sequences). These segregations were validated by unique 30-50 nucleotide long motifs and RE digestion patterns in their rrs. A single gene thus provides multiple makers for identification and surveillance of Pseudomonas.

A Genome-Wide Profiling Strategy as an Aid for Searching Unique Identification Biomarkers for Streptococcus

“…Among the various genes based identification methods, the most widely employed has been the usage of rrs gene [12][13][14][15][16]. It has proved instrumental in bacterial identification; however, the major difficulty encountered is in the cases where the organism has multiple copies of rrs within the genome [13,14,[18][19][20].…”

“…: S. pneumoniae, S. pseudopneumoniae, S. mitis, and S. oralis show more than 99 % similarity leading to unsuccessful or misleading results [10]. Recent studies have elucidated certain latent but unique characteristics in rrs: (1) 30-50 nucleotides (nts) long signatures, and (2) restriction endonuclease (RE) digestion patterns [12][13][14][15][16]. These features enable easy identification of organisms up to the species level.…”

Phylogenetic Study of Methanol Oxidizers from Chilika-Lake Sediments Using Genomic and Metagenomic Approaches

“…For PCR amplification, final genomic DNA concentration was adjusted to 50 ng/ll. 16S rRNA gene was partially amplified using primers pA (5 0 -AGAGTTTGATCCTGGCTCAG3 0 ; E. coli position [8][9][10][11][12][13][14][15][16][17][18][19][20][21][22][23][24][25][26][27] and pH (5 0 -AAGGAGGTGATCCAGCCGCA3 0 ; E. coli position 1525-1544) [31]. Amplification was carried out on a thermal cycler (Biorad PTC0220) in 100 ll volume by mixing 50-90 ng template DNA with polymerase reaction buffer (109); 100 lM (each) dATP, dCTP, dTTP and dGTP; primers pA and pH (20 ng each) and 1.0 U Taq polymerase using following conditions: initial denaturation at 94°C for 1.5 min; 35 cycles at 95°C for 1.0 min, 55°C for 1.0 min, 72°C for 1.0 min; and final extension at 72°C for 5 min.…”

“…Prudent biological technologies can become lowcost ecofriendly alternative to physicochemical methods for GHGs abatement. Molecular phylogenetic studies of microbial diversity based on the conserved functional gene sequences have greatly expanded our knowledge [9,[20][21][22][23]. The mxaF gene is supposed to be a phylogenetic chronometer for methylotrophs.…”

Comparative Genomics Reveals Biomarkers to Identify Lactobacillus Species

“…Ribosomal database project (RDP) (https://rdp.cme.msu.edu/), is a depository of 3,224,600 rrs sequences. Using the unique signatures and restriction endonuclease (RE) digestion patterns of rrs, it has been possible to re-classify bacteria, which were identified so far only up to genus level to species level [1][2][3][4]. In spite of such a roaring success of rrs, there have been cases where the gene sequences are quite similar and don't prove effective in distinguishing closely related organisms.…”