Assessing the Genetic Diversity of Isolated Cd and Cu Resistant Non-fermenting Gram-negative Bacilli from Waste Water by Using Molecular Markers

Abstract

Introduction: One of the best methods to remove pollutions resulted from heavy metals in industrial wastewater is biological removing. The first step in this way is identifying and clustering microorganisms especially, resistant bacteria to these metals. Material and methods: In this research, 17 non-fermenting gram-negative bacilli were isolated and identified from industrial wastewater. Their MIC was determined by different concentrations of Cu and Cd. With CTAB and Dellaporta, methods were extracted bacterial DNA. Then evaluated genetic diversity by 7 RAPD, 3 ISSR markers. The similarity matrix was calculated by using NTSYS-pc software based on Jaccard’s coefficient for genotypes and dendrogram was drawn in UPGMA method. Results: 133 bands were identified by using 10 markers that they included 122 polymorphic bands and 11 monomorphic bands. This markers show high level of polymorphism among the bacteria studied. Discussion and Conclusion: In the study cluster analysis and dendrogram drawn based on RAPD and ISSR markers together, there was a significant match between genetic diversity of markers and genetic diversity based on the MIC values. The combined use of both markers for genetic clustering based on resistance to copper and cadmium was more accurate. Principal component analysis was performed to complete the results of cluster analysis and 2-D, 3-D plots were drawn. The results of comparing these two methods showed that these markers had been properly selected for studying genetic diversity, and it was recommended using cluster analysis.

Keywords

• Genetic Diversity
• ‎‏ ‏RAPD
• ISSR
• Non-fermenting gram-negative Bacilli
• heavy metals