Identification and specificity validation of unique and antimicrobial resistance genes to trace suspected pathogenic AMR bacteria and to monitor the development of AMR in non-AMR strains in the environment and clinical settings

Bhagwan Narayan Rekadwad; Nanditha Pramod; Manik Prabhu Narsing Rao; Abeer Hashem; Graciela Dolores Avila-Quezada; Elsayed Fathi Abd Allah

doi:10.1016/j.sjbs.2023.103869

Identification and specificity validation of unique and antimicrobial resistance genes to trace suspected pathogenic AMR bacteria and to monitor the development of AMR in non-AMR strains in the environment and clinical settings

Saudi J Biol Sci. 2023 Dec;30(12):103869. doi: 10.1016/j.sjbs.2023.103869. Epub 2023 Nov 3.

Authors

Bhagwan Narayan Rekadwad¹, Nanditha Pramod², Manik Prabhu Narsing Rao³, Abeer Hashem⁴, Graciela Dolores Avila-Quezada⁵, Elsayed Fathi Abd Allah⁶

Affiliations

¹ Microbe AI Lab, Department of Microbiology and Biotechnology, Yenepoya Research Centre, Yenepoya (Deemed to be University), Mangalore 575018, Karnataka, India.
² Department of Biochemistry and Molecular Biology, Pondicherry University, Pondicherry 605014, India.
³ Instituto de Ciencias Químicas Aplicadas, Facultad de Ingeniería, Universidad Autónoma de Chile, Sede Talca, Talca 3460000, Chile.
⁴ Botany and Microbiology Department, College of Science, King Saud University, P.O. Box. 2460, Riyadh 11451, Saudi Arabia.
⁵ Facultad de Ciencias Agrotecnológicas, Universidad Autónoma de Chihuahua, 31350 Chihuahua, Chihuahua, México.
⁶ Plant Production Department, College of Food and Agricultural Sciences, King Saud University, P.O. Box. 2460, Riyadh 11451, Saudi Arabia.

Abstract

The detection of developing antimicrobial resistance (AMR) has become a global issue. The detection of developing antimicrobial resistance has become a global issue. The growing number of AMR bacteria poses a new threat to public health. Therefore, a less laborious and quick confirmatory test becomes important for further investigations into developing AMR in the environment and in clinical settings. This study aims to present a comprehensive analysis and validation of unique and antimicrobial-resistant strains from the WHO priority list of antimicrobial-resistant bacteria and previously reported AMR strains such as Acinetobacter baumannii, Aeromonas spp., Anaeromonas frigoriresistens, Anaeromonas gelatinfytica, Bacillus spp., Campylobacter jejuni subsp. jejuni, Enterococcus faecalis, Escherichia coli, Haemophilus influenzae, Helicobacter pylori, Klebsiella pneumonia subsp. pneumoniae, Pseudomonas aeruginosa, Salmonella enterica subsp. enterica serovar Typhimurium, Thermanaeromonas toyohensis, and Vibrio proteolyticus. Using in-house designed gene-specific primers, 18 different antibiotic resistance genes (algJ, alpB, AQU-1, CEPH-A3, ciaB, CMY-1-MOX-7, CMY-1-MOX-9, CMY-1/MOX, cphA2, cphA5, cphA7, ebpA, ECP_4655, fliC, OXA-51, RfbU, ThiU2, and tolB) from 46 strains were selected and validated. Hence, this study provides insight into the identification of strain-specific, unique antimicrobial resistance genes. Targeted amplification and verification using selected unique marker genes have been reported. Thus, the present detection and validation use a robust method for the entire experiment. Results also highlight the presence of another set of 18 antibiotic-resistant and unique genes (Aqu1, cphA2, cphA3, cphA5, cphA7, cmy1/mox7, cmy1/mox9, asaI, ascV, asoB, oxa-12, acr-2, pepA, uo65, pliI, dr0274, tapY2, and cpeT). Of these sets of genes, 15 were found to be suitable for the detection of pathogenic strains belonging to the genera Aeromonas, Pseudomonas, Helicobacter, Campylobacter, Enterococcus, Klebsiella, Acinetobacter, Salmonella, Haemophilus, and Bacillus. Thus, we have detected and verified sets of unique and antimicrobial resistance genes in bacteria on the WHO Priority List and from published reports on AMR bacteria. This study offers advantages for confirming antimicrobial resistance in all suspected AMR bacteria and monitoring the development of AMR in non-AMR bacteria, in the environment, and in clinical settings.

Keywords: Antimicrobial resistance; Genome-based detection method; Molecular detection of pathogenic bacteria; Multiplex PCR; Pathogenic bacterial strains; Strain-specific genes.