Walaa F Alsanie1, Ebaa M Felemban2, Mona A. Farid3
Mohamed M Hassan4,5, Ayman Sabry4,6 and Ahmed Gaber4,7*

1Department of Clinical Laboratories, Faculty of Applied Medical Sciences,
Taif University, Saudi Arabia.
2Department of Nursing, Faculty of Applied Medical Sciences, Taif University, Saudi Arabia.
3Genetics Department, Faculty of Agriculture, Kafrelsheikh Univ., 33516, Kafr El-Sheikh, Egypt.
4Department of Biology, Faculty of Science, Taif University, Saudi Arabia.
5Department of Genetics, Faculty of Agriculture, Minufiya University, Egypt.
6Department of Cell Biology, National Research Center, Dokki, Giza, Egypt.
7Department of Genetics, Faculty of Agriculture, Cairo University, Egypt.

Abstract

In the present study, 30 multidrug-resistant bacterial samples were isolated from different hospitals in the Taif governorate in Saudi Arabia. Given its discriminating power as a universal phylogenetic marker, the 16S rDNA gene was sequenced in a comprehensive diversity study to determine the evolutionary and phylogenetic relationships among the bacterial isolates. The 16S rDNA genes of all isolates were successfully amplified using PCR, and comprehensive identification results were based on GenBank databases. Analysis revealed nucleotide identities ranging from 76% to 100% based on the consensus sequences of 21 species, namely, Bacillus cereus, Bacillus subtilis, Bacillus tequilensis, Caldimonas manganoxidans, Citrobacter freundii, Enterococcus faecium, Escherichia fergusonii, Klebsiella pneumoniae, Lactobacillus plantarum, Lactococcus garvieae, Leuconostoc mesenteroides, Myristica yunnanensis, Pantoea eucrina, Pseudomonas aeruginosa, Staphylococcus aureus, Staphylococcus capitis, Staphylococcus caprae, Staphylococcus epidermidis, Staphylococcus hominis, Staphylococcus petrasii, and Staphylococcus saccharolyticus. We observed high variability in terms of DNA length and GC content between and within species. Phylogenetic analysis clustered the isolates into three groups. The number of sites ranged from 827 (S. aureus) to 1,219 (L. mesenteroides). Estimation of nucleotide diversity (π) showed that all analyzed sequences were diverse site-wise and also exhibited high nucleotide diversity, with π values ranging from 0.17 to 0.94. All isolates showed significantly conserved regions (P>0.05). In conclusion, the observed variations in the sequences of the target bacterial strains can be attributed to resistance to antibiotics and gene transfer among bacterial strains in the hospital environment. Further sequence analyses of antibiotic resistance genes are warranted.

Keywords: Multi-drug resistant bacteria, 16s rDNA sequencing, PCR, nucleotide diversity.