Article Main

Rayan Mazin Faisal Rafal Mhaide Younis

Abstract

Pyocyanin is a blue-green phenazine pigment and one of the most virulent factors produced by the opportunistic pathogen Pseudomonas aeruginosa. It has a redox activity and a toxic impact on living cells, as it interacts with oxygen to produce reactive oxygen species (ROS). Using antibiotics at a sub-lethal dose has an unexpected influence on the expression of pyocyanin-producing genes. In this study, qPCR technique was performed to identify the effect of eight antibiotics (cefotaxime, ampicillin, amoxiclav, ceftazidime, ceftriaxone, chloramphenicol, kanamycin and tetracycline) on the gene expression level of pyocyanin synthetic genes in P. aeruginosa isolated from different clinical sources of a few hospitals in Mosul, Iraq using qPCR technique. It was found that when P. aeruginosa was grown in media containing cefotaxime (CTX 30 µg/mL), ampicillin (AM 25 µg/mL) or amoxiclav (AMC 30 µg/mL), up-regulated the expression of pyocyanin producing genes belonging to different operons thereby increased pyocyanin production. Overexpression occurred in (CTX) treatment in PhzA1 operon with 235.56 fold change and phzM and phzS genes with 340.14, 280.13 fold change, respectively. Lower expression levels showed in tetracycline (TE 30 µg/mL) treatment, which was a (1.44) fold change for phzA1 and a (1.64, 1.08) fold change for phzM and phzS genes. More caution should be considered when delivering antibiotics to treat P. aeruginosa infections, as using drugs that the bacteria resists or at sub-lethal concentrations may trigger up-regulation of virulence factors, aiding in the spread of the disease.


 

Article Details

Article Details

Keywords

Antibiotic, Cefotaxime, Phenazine, Psuedomonas aeruginosa, Pyocyanin, qPCR

References
Abdelaziz, A. A., Abo Kamer, A. M., Al-Monofy, K. B. & Al-Madboly, L.A. (2023). Pseudomonas aeruginosa’s greenish-blue pigment pyocyanin: its production and biological activities. Microbial Cell Factories, 22, 110. doi: 10.1186/s12934-023-02122-1
Abdulrazzaq, R. & Faisal, R. (2022). Efficiency of hichrome Enterococcus faecium agar in the isolation of Enterococcus spp. and other associated bacterial genera from water. Journal of Life and Bio Sciences Research, 3, 1, 01-06. https://doi.org/10.38094/jlbsr30151
Alatraktchi, F., Dimaki, M., Støvring, N., Johansen, H. K., Molin, S. & Svendsen, W. E. (2020). Nanograss sensor for selective detection of Pseudomonas aeruginosa by pyocyanin identification in airway samples. Anal Biochem., 593: 113586. doi. Org/10.1016/j.ab.2020.113586
Aleanizy, F. S., Alqahtani, F. Y., Eltayb, E. K., Alrumikan, N., Almebki, R., Alhossan, A., Almangour, T.A. & AlQahtani, H. (2021). Evaluating the effect of antibiotics sub-inhibitory dose on Pseudomonas aeruginosa quorum sensing dependent virulence and its phenotypes. Saudi Journal of Biological Sciences, 28: 550–559. doi. Org/10.1016/j.sjbs.202010.040
Al-Shamary, M. M. K. (2018). The relationship between biofilm formation and pyocyanin producing genes in Pseudomonas aeruginosa isolated from clinical sources. Mustansiriyah University, Iraq, Thesis.
Azam, M. W. & Khan, A. U. (2019). Updates on the pathogenicity status of Pseudomonas aeruginosa. Drug discovery today, 24(1): 350-359. doi: 10.1016/j.drudis.2018.07.003
Babić, F.,Venturi, V. & Maravić-Vlahoviček, G. (2010).Tobramycin at subinhibitory concentration inhibits the RhlI/R quorum sensing system in a Pseudomonas aeruginosa environmental isolate. BMC Infectious Diseases, 10: 148. doi: 10.1186/1471-2334-10-148
Batrich, M., Maskeri, L., Schubert, R., Ho, B., Kohout, M., Abdeljaber, M., Abuhasna, A., Kholoki, M., Psihogios, P., Razzaq, T., Sawhney, S., Siddiqui, S., Xoubi, E., Cooper,A., Hatzopoulos,T. & Putonti, C. (2019). Pseudomonas diversity within urban freshwaters. Frontiers in microbiology, 10: 195. doi: 10.3389/fmicb.2019.00195
Chen, L., Xu, X., Fan, C., Zhang, R., Ji, Y., Yu, Z., Qu, H., Feng, Z., Chi, X., Shiwei Cheng, S. & Ge, Y. (2020). Pip serves as an intermediate in RpoS-modulated phz2 expression and pyocyanin production in Pseudomonas aeruginosa. Microbial Pathogenesis, 147, 104409. doi: 10.1016/j.micpath.2020.104409
Couce, A. & Blazquez, J. (2009). Side effects of antibiotics on genetic variability, article review. FEMS Microbiol Rev., 531–538. doi:10.1111/j.1574-6976.2009.00165.x
Cui, Q., Lv, H., Qi, Z., Jiang, B., Xiao, B. & Liu, L. (2016). Cross‐Regulation between the phz1 and phz2 operons maintain a balanced level of phenazine biosynthesis in Pseudomonas aeruginosa PAO1. PLoS One, 11: e0144447. doi.org/10.1371/journal.pone.0144447
Davies, J., Spiegelman, G. B. & Yim, G. (2006). The world of subinhibitory antibiotic concentrations. Current Opinion in Microbiology, 9: 445–453. doi: 10.1016/j.mib.2006.08.006
Dong, L., Pang, J., Wanga, X., Zhang, Y., Lia, G., Hua, X., Yang, X., Lub, C., Li, C. & You, X. (2020). Mechanism of pyocyanin abolishment caused by mvaT mvaU double knockout in Pseudomonas aeruginosa PAO1. Virulence, 11 (1): 57-67. doi: 10.1080/21505594.2019.1708052
Essar, D. W., Eberly, L., Hadero, A. & Crawford, I. P. (1990). Identification and characterization of genes for a second anthranilate synthase in Pseudomonas aeruginosa: interchangeability of the two anthranilate synthase and evolutionary implications. Journal of Bacteriology, 172: 884–900. doi: 10.1128/jb.172.2.884-900.1990
Fang, Y. L., Cui, Y., Zhou, L., Thawai, C., Naqvi, T.A., Zhang, H. Y. & He, Y. W. (2021). H‐NS family protein MvaU downregulates phenazine‐1‐carboxylic acid (PCA) biosynthesis via binding to an AT‐rich region within the promoter of the phz2 gene cluster in the rhizobacterium Pseudomonas strain PA1201. Synth Syst Biotechnol., 6: 262–71. doi: 10.1016/j.synbio.2021.09.006
Hata, E., Harada, T. & Itoh, M. (2019). Relationship between antimicrobial susceptibility and multilocus sequence type of Mycoplasma bovis isolates and development of a method for rapid detection of point mutations involved in decreased susceptibility to macrolides, lincosamides, tetracyclines, and spectinomycin. Appl Environ Microbiol., 85(13): e00575-19. doi: 10.1128/AEM.00575-19
He, Q., Feng, Z., Wang, Y., Wang, K., Zhang, K., Kai, L., Hao, X.,Yu, Z., Chen, L. & Ge, Y. (2019). LasR might act as an intermediate in overproduction of phenazines the absence of RpoS in Pseudomonas aeruginosa. J Microbiol Biotechnol., 29(8): 1299–309. doi: 10.4014/jmb.1904.04029
Higgins, S., Heeb, S., Rampioni, G., Fletcher, M. P., Williams, P. & Cámara, M. (2018). Differential regulation of the phenazine biosynthetic operons by quorum sensing in Pseudomonas aeruginosa PAO1‐N. Front Cell Infect Microbiol., 8: 252. doi 10.3389/fcimb.2018.00252
Hirakawa, H., Takita, A., Uchida, M., Kaneko, Y., Kakishima, Y., Tanimoto,. K, Kamitani, W. & Tomita, H. (2021). Adsorption of Phenazines produced by Pseudomonas aeruginosa using AST-120 decreases Pyocyanin-Associated cytotoxicity. Antibiotics, 10(4): 434. doi: 10.3390/antibiotics10040434
Irie, Y., L., Mensa, A., Murina, V., Hauryliuk, V., Tenson, T. & Shingler, V. (2020). Hfq assisted RsmA regulation is central to Pseudomonas aeruginosa biofilm polysaccharide PEL expression. Front Microbiol.,11: 482. doi: 10.3389/fmicb.2020.482585
Khaleel, A. M., Faisal, R. M. & Altaii, H. A. (2023). The efficiency of molecular methods compared to traditional methods in identifying bacteria from blood and cerebrospinal fluid samples. Malaysian Journal of Microbiology, 19(2). doi: 10.21161/mjm.220105
Kumar, L., Brenner, N., Brice, J., Klein-Seetharaman, J. & Sarkar, S.K. (2021). Cephalosporins Interfere with Quorum Sensing and Improve the Ability of Caenorhabditis elegans to Survive Pseudomonas aeruginosa Infection. Front. Microbiol., 12: 59849. doi.org/10.3389/fmicb.2021.598498
Laxmi, M. & Sarita, G. (2014). Reserch article diversity characterization of biofilm formin microorganismsn in food sampled local markets in Kochi, Kerala, India. Inter.J. of Rec.Sci. Res., 5(60): 1075-1070.
Meng, L., Cao, X., Li, C., Li, J., Xie, H., Shi, J., Han, M., Shen, H. & Liu, C. (2023). Housekeeping gene stability in Pesudomonas aeruginosa PAO1 under the pressure of commonly used antibiotics in molecular microbiology assays. Frontiers in Microbiology, 14: 1140515. doi: 10.3389/fmicb.2023.1140515
Mesquita, C., Soares-Castro, P. & Santos, P. (2013). Pseudomonas aeruginosa: Phenotypic flexibility and antimicrobial resistance. Formatex Research Center, 650-665. doi: 10.13140/2.1.3099.2167
Mojsoska, B., Ghoul, M., Perron, G. G., Håvard Jenssen, H. & Alatraktchi, F. (2021). Changes in toxin production of environmental Pseudomonas aeruginosa isolates exposed to sub-inhibitory concentrations of three common antibiotics. PLOS ONE, 16(3): e0248014. doi: 10.1371/journal.pone.0248014
Montelongo‐Martínez, L. F., Hernández‐Méndez, C., Muriel‐Millan, L. F., Hernández‐Estrada, R., Fabian‐Del Olmo, M. J., González-Valdez, A., Soberón‐Chávez, G. & Cocotl‐Yañez, M. (2022). Unraveling the regulation of pyocyanin synthesis by RsmA through MvaU and RpoS in Pseudomonas aeruginosa ID4365. J Basic Microbiol., 63: 51–63. doi.org/10.1002/jobm.202200432
Mukherjee, S., Moustafa, D., Smith, C. D, Goldberg, J. B. & Bassler, B. L. (2017). The RhlR quorum‐sensing receptor controls Pseudomonas aeruginosa pathogenesis and biofilm development independently of its canonical homoserine lactone autoinducer. PLoS Pathog.,13: e1006504. doi: 10.1371/journal.ppat.1006504
Pourciau, C., Lai, Y. J., Gorelik, M., Babitzke, P. & Romeo, T. (2020). Diverse mechanisms and circuitry for global regulation by the RNA- binding protein CsrA. Frontiers in Microbiology, 11: 601352. doi.org/10.3389/fmicb.2020.601352
Rampioni, G., Schuster, M., Greenberg, E. P., Zennaro, E. & Leoni, L. (2009). Contribution of the RsaL global regulator to Pseudomonas aeruginosa virulence and biofilm formation. FEMS Microbiol Lett., 301: 210–7. doi: 10.1111/j.1574-6968.2009.01817.x
Reiner, K. (2010). Catalase test protocol. American Society for Microbiology,1-6.
Saleem, H., Mazhar, S., Syed, Q., Javed, M.Q. & Adnan, A. (2021). Bio-characterization of food grade pyocyanin bio-pigment extracted from chromogenic Pseudomonas species found in pakistani native fora. Arab J Chem, 14(3): 103005. doi.org/10.1016/j.arabjc.2021.103005
Schulmeyer, K. H., Diaz, M. R., Bair, T. B., Sanders, W., Gode, C. J., Laederach, A., Wolfgang, M. C. & Yahr, T. L. (2016). Primary and secondary sequence structure requirements for recognition and discrimination of target RNAs by Pseudomonas aeruginosa RsmA and RsmF. J Bacteriol., 198(18): 2458–69. doi: 10.1128/JB.00343-16
Shaan, L. G. & Robert E. W. (2013). Pseudomonas aeruginosa: new insights into pathogenesis and host defenses. Pathogens and Disease, 27(2): 2049-2057. doi: 10.1111/2049-632X.12033
Skindersoe, M. E., Alhede, M., Phipps, R., Yang, L., Jensen, P. O., Rasmussen, T. B., Bjarnsholt, T., Tolker-Nielsen, T., Hoiby, N. & Givskov, M. (2008). Effects of Antibiotics on Quorum Sensing in Pseudomonas aeruginosa. Antimicrob Agents Chemother, 52(10): 3648-3663. doi: 10.1128/AAC.01230-07
Soto-Aceves, M. P., Cocotl-Yañez, M., Servín-González, L. & SoberónChávez, G. (2021). The Rhl quorum-sensing system is at the top of the regulatory hierarchy under phosphate-limiting conditions in Pseudomonas aeruginosa PAO1. J Bacteriol., 203(5): eoo475–20. doi: 10.1128/JB.00475-20
Su, H. C., Ramkissoon, K., Doolittle, J., Clark, M., Khatun, J., Secrest, A., Wolfgang, M. C. & Giddings, M. C. (2010). The Development of Ciprofloxacin Resistance in Pseudomonas aeruginosa Involves Multiple Response Stages and Multiple Proteins. Antimicrobial Agents and Chemotherapy, 54(11): 4626 LP– 4635. doi: 10.1128/AAC.00762-10
Wang, K., Kai, L., Zhang, K., Hao, M., Yu, Y., Xu, X., Yu, Z., Chen, L., Chi, X. & Ge, Y. (2020). Overexpression of phzM contributes to much more production of pyocyanin converted from phenazine‐1‐carboxylic acid in the absence of RpoS in Pseudomonas aeruginosa. Archives of Microbiology, 202 (6), 1507-1515. doi:10.1007/s00203-020-01837-8
Weiner, L. M., Webb, A. K., Limbago, B., Dudeck, M. A., Patel, J., Kallen, A. J., Edwards, J. R. & Sievert, D. M. (2016). Antimicrobial-resistant pathogens associated with healthcare-associated infections: Summary of data reported to the National Healthcare Safety Network at the Centers for Disease Control and Prevention, 2011-2014. Infect Control Hosp Epidemiol, 37(11), 1288-1301. doi: 10.1017/ice.2016.174
Winn, W., Allen, S. & Janda, W. (2006). Koneman's color atlas and textbook of diagnostic microbiology. 6th ed. Lippincott Williams and Wilkins. Philadelphia. Pp: 947-982.
Zhao, K., Li, J., Yang, X., Zeng, Q., Liu, W., Wu, Y., Zhou, H., Prithiviraj, B., Wang, X., Zhou, X. & Chu, Y.(2022). Subinhibitory Cefotaxime and Levofloxacin Concentrations Contribute to Selection of Pseudomonas aeruginosa in Coculture with Staphylococcus aureus. Applied and Enviromental Microbiology, 88(12): e0059222 doi: 10.1128/aem.00592-22
Section
Research Articles

How to Cite

Effect of antibiotics on the expression of pyocyanin synthetic genes in Pseudomonas aeruginosa isolated from different clinical sources of a few hospitals in Mosul, Iraq. (2024). Journal of Applied and Natural Science, 16(2), 812-819. https://doi.org/10.31018/jans.v16i2.5590