Investigation of the Potential Association between Clustered Regularly Interspersed Short Palindromic Repeats (CRISPR) and Antibiotic Resistance Pattern of Bacterial Strains Isolated from Medical Waste and Environmental Water

Enterobacteriaceae are important human pathogens that cause many food and waterborne illness. Rapid emergence of multi-drug resistant bacteria in Bangladesh has become a serious problem. Phage-host interaction is now considered as the major driving force for the conversion of non-pathogenic bacteria to pathogenic ones. Evolution of highly pathogenic and antibiotic resistant bacteria largely depends upon the horizontal gene transfer by means of plasmid, megaplasmid and bacteriophages. Conversely, bacteria may acquire a novel defence mechanism called CRISPR (Clustered regularly interspersed short palindromic repeats) that can restrict horizontal transfer of plasmids and bacteriophages to limit the spread of antibiotic resistance genes among bacterial species. In this study, twenty bacterial strains were isolated from water of different medical waste and Buriganga river. Therefore, CRISPR locus was investigated following various biochemical and molecular analysis of those bacterial isolates. Identification of the bacterial isolates was conducted by Polymerase Chain Reaction (PCR) based assay of 16S rDNA extracted from those isolated strains. Results indicated that most strains were identified as Proteus mirabilis and Citrobacter freundii which mainly cause septicemia, and pneumonia in human. Thereafter, antibiogram of these strains was performed by using 11 different antibiotic discs where bacterial isolates from medical drainage system showed more resistant to antibiotics than the river water. In this study, CRISPR locus was also investigated within the genome of the isolated bacterial stains but unexpectedly, we did not find any CRISPR locus in their genome. In conclusion, we confirm that multi-drug resistant bacterial strains are devoid of CRISPR locus suggesting a possible negative association between CRISPR locus and antibiotic resistance. Further studies to pinpoint are required to elucidate the underlying mechanism of the association between CRISPR and antibiotic resistance in these isolated strains.

BioCluster:Tool for Identification and Clustering of Enterobacteriaceae Based on Biochemical Data

Presumptive identifcation of different Enterobaeteriaeeae species is routinely achieved based on biochemical properties. Traditional practice includes manual comparison of each biochem- ical property of the unknown sample with known reference samples and inference of its identity based on the maximum similarity pattern with the known samples. This process is labor- intensive, time-consuming, error-prone, and subjective. Therefore, automation of sorting and sim- ilarity in calculation would be advantageous. Here we present a MATLAB-based graphical user interface （GUI） tool named BioCluster. This tool was designed for automated clustering and iden- tification of Enterobacteriaceae based on biochemical test results. In this tool, we used two types of algorithms, i.e., traditional hierarchical clustering （HC） and the Improved Hierarchical Clustering （IHC）, a modified algorithm that was developed specifically for the clustering and identification of within this species. IHC takes into account the variability in result of 1-47 biochemical tests family. This tool also provides different options to optimize the clus- tering in a user-friendly way. Using computer-generated synthetic data and some real data, we have demonstrated that BioCluster has high accuracy in clustering and identifying enterobacterial species based on biochemical test data. This tool can be freely downloaded at http：//microbialgen.du.ac.bd/ biocluster/.