Acta Scientific Medical Sciences (ASMS)(ISSN: 2582-0931)

Research Article Volume 7 Issue 10

Characterization of Chromosomal VmeAB, MacAB and EmrD Multidrug-Efflux Genes of Vibrio parahaemolyticus to Design Diagnostic PCR Primers to Improve Associated Problems in Shrimp Aquaculture

Asit Kumar Chakraborty*, Uttam Maity and Chandan Halder

Department of Biotechnology and Biochemistry, Oriental Institute of Science and Technology-West Bengal, Vidyasagar University, India

*Corresponding Author: Asit Kumar Chakraborty, Department of Biotechnology and Biochemistry, Oriental Institute of Science and Technology-West Bengal, Vidyasagar University, India.

Received: March 14, 2023; Published: September 29, 2023


Vibrio species contamination in fish is a serious threat to human population worldwide. Aquaculture of shrimp has increased in India due to high demand in the Europe and America. Previously, we reported the presence of unique chromosomal blaCARB, pbp1B and CatC1 mdr genes involved in multi-resistance of Vibrio parahaemolyticus (Vp) and useful for diagnostic primers design. Escherichia coli AcrA-AcrB-TolC tripartite multidrug efflux genes were related to MexA-MexB-OprM genes of Pseudomonas aeruginosa and OqxAB-TolC genes of Klebsiella pneumoniae. Here, we showed that VmeAB and VmeYZ related to acrAB or mexAB as well as MacAB drug-efflux genes might be also responsible for multidrug resistance in Vp. Many heterogeneous Vme-isomers like vmeAB, vmeCD, vmeEF, vmeHI, vmeJK located in Ch-1 whereas vmeHI and vmeCD located in the complement strand (accession no. CP034294). The Ch-2 of Vp (accession no. CP020428) contained vmeYZ, vmePQ, vmeUV (RND-type) as well as macAB (ABC-type), EmrD (MFS-type) and mdtL (MATE-type) drug efflux genes. BLAST-2 analysis suggested only vmeB and vmeZ had similarities over 40% to acrB, mexB or oqxB types popular bacterial RND permease subunits. The macB protein of Vp has 56% similarity to E. coli macB protein and within the related Vibrio species (V. harveyi and V. alginolyticus) the similarity found to be round 98%. The Vp EmrD protein has 41.49% homology to E. coli while V. cholerae protein has only 29% homology. The MdtL protein of Vp has 61% similarity to E. coli and not suitable for primer design. We designed the primers using NCBI Primer Design Software and oligonucleotides were validated by Oligo Analyzer Software 3.2. Multi-alignment of vme-related sequences done by CLUSTAL-Omega software to confirm heterogeneities and BLASTN search of primers confirmed specificities to Vp Ch-1 or Ch-2. The P2F-vmeB/P2.1R-vmeB primers for VmeB gene and P2.1F-macB/P2R-macB primers for MacB gene should be useful with good species specificity. Such primers will be useful to detect V. parahaemolyticus contamination in fish aquaculture.

Keywords: Multidrug Efflux Genes; Diagnostic PCR Primers; Vibrio parahaemolyticus; Shrimp Aquaculture; Multidrug Resistance; acrAB and mexAB, macAB and emrCD


  1. Putman M., et al. “Molecular properties of bacterial multidrug transporters”. Microbiology and Molecular Biology Reviews 64 (2000): 672-693. 
  2. Padilla E., et al. “Klebsiella pneumoniae AcrAB efflux pump contributes to antimicrobial resistance and virulence”. Antimicrobial Agents and Chemotherapy 54 (2010): 177-183.
  3. Pos KM. “Drug transport mechanism of the AcrB efflux pump”. Biochimica et Biophysica Acta 1794 (2009): 782-793.
  4. Yamaguchi A., et al. “Structural basis of RND-type multidrug exporters”. Frontiers in Microbiology 6 (2015):
  5. Nishino K., et al. “Virulence and drug resistance roles of multidrug efflux systems of Salmonella enterica serovar Typhimurium”. Molecular Microbiology 59 (2006): 126-141.
  6. Nehme D and Poole K. “Assembly of the MexAB-OprM multidrug pump of Pseudomonas aeruginosa: component interactions defined by the study of pump mutant suppressorsJournal of Bacteriology 189 (2007): 6118-6127.
  7. Nikaido H. “Multidrug efflux pumps of gram-negative bacteriaJournal of Bacteriology 178 (1996): 5853-5859. 
  8. Mima T., et al. “Gene cloning and properties of the RND-type multidrug efflux pumps MexPQ-OpmE and MexMN-OprM from Pseudomonas aeruginosa”Microbiology and Immunology 49 (2005): 999-1002. 
  9. Li Y., et al. “A new member of the tripartite multidrug efflux pumps, MexVW-OprM, in Pseudomonas aeruginosa”Journal of Antimicrobial Chemotherapy 52 (2003): 572-575. 
  10. Maseda H., et al. “Assignment of the substrate-selective subunits of the MexEF-OprN multidrug efflux pump of Pseudomonas aeruginosa”Antimicrobial Agents and Chemotherapy 44 (2000): 658-664. 
  11. Nishino K and Yamaguchi A. “Analysis of a complete library of putative drug transporter genes in Escherichia coli”Journal of Bacteriology 183 (2001): 5803-5812. 
  12. Nishino K., et al. “Roles of TolC-dependent multidrug transporters of Escherichia coliin resistance to beta-lactams”. Antimicrobial Agents and Chemotherapy 47 (2003): 3030-3033. 
  13. Poole K., et al. “Overexpression of the mexC-mexD-oprJefflux operon in nfxB-type multidrug-resistant strains of Pseudomonas aeruginosa”Molecular Microbiology 21 (1996): 713-724.
  14. Nesme J., et al. “Large-scale metagenomic-based study of antibiotic resistance in the environment”. Current Biology 24 (2014): 1096-1100.
  15. Mine T., et al. “Expression in Escherichia coliof a new multidrug efflux pump, MexXY, from Pseudomonas aeruginosa”. Antimicrobial Agents and Chemotherapy 43 (1999): 415-417. 
  16. Du D., et al. “Structure of the AcrAB-TolC multidrug efflux pump”. Nature 509 (2014): 512-515.
  17. Ma D., et al. “Molecular cloning and characterization of acrA and acrE genes of Escherichia coli”Journal of Bacteriology 19 (1993): 6299-6313.
  18. Chuanchuen R., et al. “Substrate-dependent utilization of OprM or OpmH by the Pseudomonas aeruginosaMexJK efflux pump”. Antimicrobial Agents and Chemotherapy 49 (2005): 2133-2136. 
  19. Aires JR and Nikaido H. “Aminoglycosides are captured from both periplasm and cytoplasm by the AcrD multidrug efflux transporter of Escherichia coli”Journal of Bacteriology 187 (2005): 1923-1929. 
  20. Koronakis V., et al. “Crystal structure of the bacterial membrane protein TolC central to multidrug efflux and protein export”. Nature6789 (2000): 914-919.
  21. Lin YT., et al. “MacABCsm, an ABC-type tripartite efflux pump of Stenotrophomonas maltophilia involved in drug resistance, oxidative and envelope stress tolerances and biofilm formation”. Journal of Antimicrobial Chemotherapy69 (2014): 3221-3226.
  22. Rouquette-Loughlin CE., et al. “Characterization of the MacA-MacB efflux system in Neisseria gonorrhoeae”. Journal of Antimicrobial Chemotherapy 56 (2005): 856-860.
  23. Shirshikova TV., et al. “The ABC-Type Efflux Pump MacAB Is Involved in Protection of Serratia marcescens against Aminoglycoside Antibiotics, Polymyxins, and Oxidative Stress”. mSphere 2 (2021): e00033-21.
  24. Chatterjee M., et al. “Antibiotic resistance in Pseudomonas aeruginosa and alternative therapeutic options”. International Journal of Medical Microbiology 306 (2016): 48-58.
  25. Makino K., et al. “Genome sequence of Vibrio parahaemolyticus: a pathogenic mechanism distinct from that of V cholerae”. Lancet 9359 (2003): 743-749.
  26. Haendiges J., et al. “Draft Genome Sequences of Clinical Vibrio parahaemolyticus Strains Isolated in Maryland (2010 to 2013)”. Genome Announcement 4 (2014): e00776-714.
  27. Lau DYL., et al. “What Whole Genome Sequencing has told us about Pathogenic Vibrios”. Advances in Experimental Medicine and Biology 1404 (2023): 337-352.
  28. Gonzalez-Escalona N., et al. “Genome sequence of the clinical O4:K12 serotype Vibrio parahaemolyticus strain 10329Journal of Bacteriology 193 (2011): 3405-3406.
  29. Matsuo T., et al. “Characterization of all RND-type multidrug efflux transporters in Vibrio parahaemolyticus”. Microbiology Open5 (2013): 725-742.
  30. Matsuo T., et al. “VmeAB, an RND-type multidrug efflux transporter in Vibrio parahaemolyticus”. Microbiology (Reading)153 (2007): 4129-4137.
  31. Liu M and Chen S. “Draft Genome Sequence of Vibrio parahaemolyticus V110, Isolated from Shrimp in Hong Kong”. Genome Announcement3 (2013): e00300-13.
  32. Rahman MM., et al. “Molecular cloning and characterization of all RND-type efflux transporters in Vibrio choleraenon-O1”. Microbiology and Immunology 51 (2007): 1061-1070. 
  33. Bina JE., et al. “Characterization of the Vibrio cholerae vexABand vexCD efflux systems”. Archives of Microbiology 186 (2006): 171-181. 
  34. Yeung M and Thorsen T. “Development of a More Sensitive and Specific Chromogenic Agar Medium for the Detection of Vibrio parahaemolyticus and Other Vibrio Species”. Journal of Visualized Experiments 117 (2016):
  35. Maniatis T., et al. “Molecular Cloning-A laboratory manual”. (Cold Spring Harbor Laboratory Press, Cold spring harbour, NY, USA (1982).
  36. Tinwongger S., et al. “Development of PCR diagnosis for shrimp acute hepatopancreatic necrosis disease (AHPND) strain of Vibrio parahaemolyticus”. Fish Pathology 49 (2014): 159-164.
  37. Bej AK., et al. “Detection of total and hemolysin-producing Vibrio parahaemolyticus in shellfish using multiplex PCR amplification of tl, tdh and trh”. Journal of Microbiological Methods 3 (1999): 215-225.
  38. Seeger MA., et al. “Structural asymmetry of AcrB trimer suggests a peristaltic pump mechanism”.Science 313 (2006): 1295-1298.
  39. Janganan TK., et al. “Evidence for the assembly of a bacterial tripartite multidrug pump with a stoichiometry of 3:6:3”. Journal of Biological Chemistry 286 (2011): 26900-26912. 
  40. Hinchliffe P., et al. “Structure and operation of bacterial tripartite pumps”. Annual Review of Microbiology 67 (2013): 221-242.
  41. Chakraborty AK. “High mode contamination of multi-drug resistant bacteria in Kolkata: mechanism of gene activation and remedy by heterogeneous phyto-antibiotics”. IndianJournal of Biotechnology 14 (2015): 149-159.
  42. Gavilan RG., et al. “Vibrio parahaemolyticus Epidemiology and Pathogenesis: Novel Insights on an Emerging Foodborne Pathogen”. Advances in Experimental Medicine and Biology 1404 (2023): 233-251.
  43. Cabanillas-Beltran H., et al. “Outbreak of gastroenteritis caused by the pandemic Vibrio parahaemolyticusO3:K6 in Mexico”. FEMS Microbiology Letter 265 (2006): 76-80. 
  44. Chakraborty AK., et al. “A saponin-polybromophenol antibiotic (CU1) from Cassia fistula bark targeting RNA polymerase”. Current Research Pharmacology and Drug Discovery 3 (2022):
  45. Chakraborty AK., et al. “Multidrug- Resistant Bacteria with activated and diversified MDR Genes in Kolkata Water: Ganga Action Plan and Heterogeneous Phyto-Antibiotics tackling superbug spread in India”. American Journal of Drug Delivery and Therapeutics1 (2018): 2.
  46. Rajpara N., et al. “A Highly Promiscuous Integron, Plasmids, Extended Spectrum Beta Lactamases and Efflux Pumps as Factors Governing Multidrug Resistance in a Highly Drug Resistant Vibrio fluvialisIsolate BD146 from Kolkata, India”. Indian Journal of Microbiology1 (2018): 60-67.
  47. Elmahdi S., et al. “Antibiotic resistance of Vibrio parahaemolyticus and Vibrio vulnificus in various countries: A review”. Food Microbiology 57 (2016): 128-134.
  48. Stephen J., et al. “Membrane Efflux Pumps of Pathogenic VibrioSpecies: Role in Antimicrobial Resistance and Virulence”. Microorganisms2 (2022): 382.
  49. Furniss RCD., et al. “Breaking antimicrobial resistance by disrupting extracytoplasmic protein folding”. eLife 11 (2022): e57974.
  50. Li R., et al. “First detection of AmpC β-lactamase blaCMY-2 on a conjugative IncA/C plasmid in a Vibrio parahaemolyticus isolate of food origin”. Antimicrobial Agents and Chemotherapy7 (2015): 4106-4111.
  51. Wang T., et al. “Whole genome sequencing and antimicrobial resistance analysis of Vibrio parahaemolyticus Vp2015094 carrying an antimicrobial-resistant plasmid”. Journal of Global Antimicrobial Resistance 30 (2022): 47-49.


Citation: Asit Kumar Chakraborty., et al. “Characterization of Chromosomal VmeAB, MacAB and EmrD Multidrug-Efflux Genes of Vibrio parahaemolyticus to Design Diagnostic PCR Primers to Improve Associated Problems in Shrimp Aquaculture”.Acta Scientific Medical Sciences 7.10 (2023): 71-85.


Copyright: © 2023 Asit Kumar Chakraborty., et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.


Acceptance rate30%
Acceptance to publication20-30 days
Impact Factor1.403

Indexed In

News and Events

  • Certification for Review
    Acta Scientific certifies the Editors/reviewers for their review done towards the assigned articles of the respective journals.
  • Submission Timeline for Upcoming Issue
    The last date for submission of articles for regular Issues is June 25, 2024.
  • Publication Certificate
    Authors will be issued a "Publication Certificate" as a mark of appreciation for publishing their work.
  • Best Article of the Issue
    The Editors will elect one Best Article after each issue release. The authors of this article will be provided with a certificate of "Best Article of the Issue"
  • Welcoming Article Submission
    Acta Scientific delightfully welcomes active researchers for submission of articles towards the upcoming issue of respective journals.

Contact US