Genotyping of clinical Serratia marcescens isolates using molecular techniques

Document Type : Research Paper

Authors

1 Department of Biology, College of Education, University of Al-Qadisiyah, Iraq

2 Department of Biology, College of Education, University of Al-Qadisiyah, Iraq.

10.22103/jab.2025.24830.1665

Abstract

Objective
Serratia marcescens is a Gram-negative, facultatively anaerobic bacillus belonging to the Enterobacteriaceae family. It is an opportunistic pathogen that has gained increasing attention due to its involvement in a wide range of nosocomial and community-acquired infections, including urinary tract infections, respiratory tract infections, bloodstream infections, and wound infections. The bacterium is also known for its ability to survive in diverse environmental conditions and its intrinsic resistance to several antibiotics, making its infections particularly challenging to treat in clinical settings. The 16S ribosomal RNA (16S rRNA) gene, which is highly conserved among bacteria but also contains hypervariable regions, is widely used for phylogenetic studies and bacterial identification. Genotyping using PCR-restriction fragment length polymorphism (PCR-RFLP) analysis of the 16S rRNA gene allows differentiation of bacterial strains based on the presence of specific restriction sites. This technique provides insights into genetic diversity and evolutionary relationships among isolates. The aim of this study was to genotype clinical isolates of S. marcescens obtained from various healthcare sources in Al-Diwaniyah Governorate, Iraq using techniques such as PCR, sequencing, and PCR-RFLP with the restriction enzymes AluI and MspI, and explore the genetic diversity of local isolates.
Materials and methods
A total of 200 clinical samples were collected from patients attending Diwaniyah Teaching Hospital as well as from several private medical clinics across Al-Diwaniyah Governorate, Iraq. Samples were inoculated onto standard selective and differential media. Genomic DNA was extracted from purified S. marcescens isolates using a commercial bacterial DNA extraction kit. Molecular identification was performed by amplifying the 16S rRNA gene using universal bacterial primers. PCR products of confirmed S. marcescens isolates were purified and sent to a commercial sequencing facility in South Korea. Phylogenetic trees were constructed using the neighbor-joining method. To assess genetic variability among S. marcescens isolates, PCR-restriction fragment length polymorphism (PCR-RFLP) analysis of the 16S rRNA gene was performed. Distinct RFLP patterns were analyzed visually, and the number of different genotypes was determined based on banding profiles for each enzyme.
Results
A total of 20 isolates were obtained from different sources as follows: 15 (75%) isolates from urinary tract infections, 3 (15%) isolates from burn and wound injuries, and 2 (10%) isolates from eye injuries.
Conclusions
The study showed that molecular techniques provided accurate data on the genetic makeup of Serratia marcescens, improving diagnostic accuracy, paving the way for the development of future diagnostic tools that are more sensitive and reliable.

Keywords

References

Anfal, M. K., Suhaila, S., & Mohammed, A. F. (2011). Isolation of Multi Antibiotic Resistance Serratia marcescens and the Detection of AmpC & GESL Genes by Polymerase Chain Reaction Technique. Al-Mustansiriyah Journal of Science 22(6), 329-346. https://doi.org/10.23851/mjs.v22i6
Barman, S., Bhattacharya, S. S., & Mandal, N. C. (2020). Serratia. In N. Amaresan, M. S. Kumar, K. Annapurna, K. Kumar, & A. Sankaranarayanan (Eds.), Beneficial microbes in agro-ecology (pp. 27–36). Academic Press. https://doi.org/10.1016/B978-0-12-823414-3.00003-4
Cristina, M. L., Sartini, M., & Spagnolo, A. M. (2019). Serratia marcescens infections in neonatal intensive care units (NICUs). International Journal of Environmental Research and Public Health, 16(4), 610. https://doi.org/10.3390/ijerph16040610
Daham, R. I. (2021). Daham, R. I. (2021). Immunological and biological effects of Serratia marcescens extracellular protease isolated from UTI infections (Master’s thesis, Mustansiriyah University).
Ghorbanalizadgan, M., Bakhshi, B., Shams, S., & Najar-Peerayeh, S. (2019). Ghorbanalizadgan, M., Bakhshi, B., Shams, S., & Najar-Peerayeh, S. (2019). Pulsed-field gel electrophoresis fingerprinting of Campylobacter jejuni and Campylobacter coli strains isolated from clinical specimens, Iran. International Microbiology, 22(3), 391–398. https://doi.org/10.1007/s10123-019-00073-2
Hamzah, A. S., & Awayid, H. S. (2023). Hamzah, A. S., & Awayid, H. S. (2023). Analysis of virulence genes sequencing of Serratia marcescens in Iraqi hospitals. Cellular and Molecular Biology, 69(11), 162–166. https://doi.org/10.14715/cmb/2023.69.11.27
Hanczvikkel, A., Tóth, Á., Németh, I. A. K., Bazsó, O., Závorszky, L., Buzgó, L., ... & Hajdu, Á. (2024). Hanczvikkel, A., Tóth, Á., Németh, I. A. K., Bazsó, O., Závorszky, L., Buzgó, L., ... & Hajdu, Á. (2024). Nosocomial outbreak caused by disinfectant-resistant Serratia marcescens in an adult intensive care unit, Hungary, February to March 2022. Eurosurveillance, 29(26), 2300492. https://doi.org/10.2807/1560-7917.ES.2024.29.26.2300492
Idris, A. B., Hassan, H. G., Ali, M. A. S., Eltaher, S. M., Idris, L. B., Altayb, H. N., Abass, A. M., Ibrahim, M. M. A., Ibrahim, E. M., & Hassan, M. A. (2020). Molecular phylogenetic analysis of 16S rRNA sequences identified two lineages of Helicobacter pylori strains detected from different regions in Sudan suggestive of differential evolution. International Journal of Microbiology, 2020, 8825718. https://doi.org/10.1155/2020/8825718
Javanmard, A., Mohammadabadi, M. R., Zarrigabayi, G. E., & Gharahedaghi, A. A. (2008). Polymorphism within the intron region of the bovine leptin gene in Iranian Sarabi cattle (Bos taurus). Russian Journal of Genetics, 44(4), 495-497. https://doi.org/10.1134/S1022795408040169
Kamali, A., Ferguson, D., Dowless, H., Ortiz, N., Mukhopadhyay, R., Schember, C., ... & Kimura, A. (2024). Kamali, A., Ferguson, D., Dowless, H., Ortiz, N., Mukhopadhyay, R., Schember, C., ... & Kimura, A. (2024). Outbreak of invasive Serratia marcescens among persons incarcerated in a state prison, California, USA, March 2020–December 2022. Emerging Infectious Diseases, 30(Suppl 1), S41. https://doi.org/10.3201/eid3001.231234
Kashash, R. R. (2021). Kashash, R. R. (2021). Isolation and molecular identification of Serratia marcescens from different clinical sources in humans and birds (Master’s thesis, University of Baghdad).
Kljakić, D., Milosavljević, M. Z., Jovanović, M., Popović, V. Č., & Raičević, S. (2020). Kljakić, D., Milosavljević, M. Z., Jovanović, M., Popović, V. Č., & Raičević, S. (2020). Serratia marcescens as a cause of unfavorable outcome in the twin pregnancy. Open Medicine, 16(1), 81–86. https://doi.org/10.1515/med-2021-0001
Matys, J., Kensy, J., Gedrange, T., Zawiślak, I., Grzech-Leśniak, K., & Dobrzyński, M. (2024). Matys, J., Kensy, J., Gedrange, T., Zawiślak, I., Grzech-Leśniak, K., & Dobrzyński, M. (2024). A molecular approach for detecting bacteria and fungi in healthcare environment aerosols: A systematic review. International Journal of Molecular Sciences, 25(8), 4154. https://doi.org/10.3390/ijms25084154
Mohammadabadi, M. (2016). Inter-simple sequence repeat loci associations with predicted breeding values of body weight in kermani sheep. Genetics in the Third Millennium, 14(4), 4386-4393. https://sciexplore.ir/Documents/Details/472-591-539-924
Mohammadabadi, M. R., Torabi, A., Tahmourespoor, M., & Baghizadeh, A. (2010). Analysis of bovine growth hormone gene polymorphism of local and Holstein cattle breeds in Kerman province of Iran using polymerase chain reaction restriction fragment length. African Journal of Biotechnology, 9(41), 6848-6852. https://www.ajol.info/index.php/ajb/article/view/130242
Mohammadabadi, M., Akhtarpoor, A., Khezri, A., Babenko, O., Stavetska, R. V., Tytarenko, I., Ievstafiieva, Y., Buchkovska, V., Slynko, V. and Afanasenko, V. (2024a). The role and diverse applications of machine learning in genetics, breeding, and biotechnology of livestock and poultry. Agricultural Biotechnology Journal, 16(4), 413-442. https://doi.org/10.22103/jab.2025.24662.1644 
Mohammadabadi, M., Babenko Ivanivna, O., Borshch, O., Kalashnyk, O., Ievstafiieva, Y. and Buchkovska, V. (2024c). Measuring the relative expression pattern of the UCP2 gene in different tissues of the Raini Cashmere goat. Agricultural Biotechnology Journal, 16(3), 317-332. https://doi.org/10.22103/jab.2024.24337.1627
Mohammadabadi, M., Kheyrodin, H., Afanasenko, V., Babenko Ivanivna, O., Klopenko, N., Kalashnyk, O., Ievstafiieva, Y., & Buchkovska, V. (2024b). The role of artificial intelligence in genomics. Agricultural Biotechnology Journal, 16(2), 195-279. https://doi.org/10.22103/jab.2024.23558.1575
Mohammadabadi, M., Meymandi, M.G., Montazeri, M., Afanasenko, V., & Kalashnyk, O. (2021a). Molecular characterization of Iranian dromedaries using microsatellite markers. Acta Agronomica, 69(4), 321-330. https://api.semanticscholar.org/CorpusID:249364533
Mohammadabadi, M., Oleshko, V., Oleshko, O., Heiko, L., Starostenko, I., Kunovskii, J., Bazaeva, A., & Roudbari, Z. (2021b). Using inter simple sequence repeat multi-loci markers for studying genetic diversity in guppy fish. Turkish Journal of Fisheries and Aquatic Sciences, 21, 603-613. http://doi.org/10.4194/1303-2712-v21_12_03
Mohammadifar, A., & Mohammadabadi, M. (2018). Melanocortin-3 receptor (MC3R) gene association with growth and egg production traits in fars indigenous chicken. Malaysian Applied Biology, 47(3), 85-90.
Mohammadifar, A., & Mohammadabadi, M. R. (2017). The effect of uncoupling protein polymorphisms on growth, breeding value of growth, and reproductive traits in the Fars indigenous chicken. Iranian Journal of Applied Animal Science, 7(4), 679-685. https://journals.iau.ir/article_535799.html
Mohammadinejad, F., Mohammadabadi, M., Roudbari, Z., & Sadkowski, T. (2022). Identification of Key Genes and Biological Pathways Associated with Skeletal Muscle Maturation and Hypertrophy in Bos taurus, Ovis aries, and Sus scrofa. Animals, 12(24), 3471. https://doi.org/10.3390/ani12243471
Noori, A. N., Behzadi, M. R. B., & Mohammadabadi, M. R. (2017). Expression pattern of Rheb gene in Jabal Barez Red goat. Indian Journal of Animal Science, 87(11), 1375-1378. https://doi.org/10.56093/ijans.v87i11.75890
Pazla R, Yanti G, Jamarun N, Zain M, Triani HD, Putri EM, Srifani A. Identification of phytase producing bacteria from acidifying Tithonia diversifolia: Potential for ruminant feed development. Saudi J Biol Sci. 2024 Jul;31(7):104006. https://doi.org/10.1016/j.sjbs.2024.104006.
Pazla R, Yanti G, Jamarun N, Zain M, Triani HD, Putri EM, Srifani A. Identification of phytase producing bacteria from acidifying Tithonia diversifolia: Potential for ruminant feed development. Saudi J Biol Sci. 2024 Jul;31(7):104006. https://doi.org/10.1016/j.sjbs.2024.104006.
Pérez-Viso, B., Aracil-Gisbert, S., Coque, T. M., Del Campo, R., Ruiz-Garbajosa, P., & Cantón, R. (2021). Evaluation of CHROMagar™-Serratia agar, a new chromogenic medium for the detection and isolation of Serratia marcescens. European Journal of Clinical Microbiology & Infectious Diseases, 40(12), 2593–2596. https://doi.org/10.1007/s10096-021-04319-9
Posluszny, J. A., Jr., Conrad, P., Halerz, M., Shankar, R., & Gamelli, R. L. (2011). Surgical burn wound infections and their clinical implications. Journal of Burn Care & Research, 32(3), 324–333. https://doi.org/10.1097/BCR.0b013e31820aaf4f
Prado, L. C. D. S., Giacchetto Felice, A., Rodrigues, T. C. V., Tiwari, S., Andrade, B. S., Kato, R. B., ... & Soares, S. D. C. (2022). New putative therapeutic targets against Serratia marcescens using reverse vaccinology and subtractive genomics. Journal of Biomolecular Structure and Dynamics, 40(20), 10106–10121. https://doi.org/10.1080/07391102.2021.1931022
Riegman, P. H. J., Becker, K. F., Zatloukal, K., Pazzagli, M., Schröder, U., & Oelmuller, U. (2019). How standardization of the pre-analytical phase of both research and diagnostic biomaterials can increase reproducibility of biomedical research and diagnostics. New Biotechnology, 53, 35–40. https://doi.org/10.1016/j.nbt.2018.03.002
Riley, L. W. (2018). Laboratory methods in molecular epidemiology: Bacterial infections. Microbiology Spectrum, 6(6), 10–1128. https://doi.org/10.1128/microbiolspec.ARBA-0022-2017
Rohit, A., Maiti, B., Shenoy, S., & Karunasagar, I. (2016). Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) for rapid diagnosis of neonatal sepsis. Indian Journal of Medical Research, 143(1), 72–78. https://doi.org/10.4103/0971-5916.178618
Saadatabadi, L. M., Mohammadabadi, M., Nanaei, H. A., Ghanatsaman, Z. A., Stavetska, R. V., Kalashnyk, O., Kochuk-Yashchenko, O. A., & Kucher, D. M. (2023). Unraveling candidate genes related to heat tolerance and immune response traits in some native sheep using whole genome sequencing data. Small Ruminant Research, 225, 107018. https://doi.org/10.1016/j.smallrumres.2023.107018
Sader, H. S., Farrell, D. J., Flamm, R. K., & Jones, R. N. (2014). Antimicrobial susceptibility of Gram-negative organisms isolated from patients hospitalized in intensive care units in United States and European hospitals (2009–2011). Diagnostic Microbiology and Infectious Disease, 78(4), 443–448. https://doi.org/10.1016/j.diagmicrobio.2013.11.021
Sciesielski, L. K., Osang, L. K., Dinse, N., Weber, A., Bührer, C., Kola, A., & Dame, C. (2023). Validation of a new PCR-based screening method for prevention of Serratia marcescens outbreaks in the neonatal intensive care unit. Neonatology, 120(2), 176–184. https://doi.org/10.1159/000529230
Shokri, S., Khezri, A., Mohammadabadi, M., & Kheyrodin, H. (2023). The expression of MYH7 gene in femur, humeral muscle, and back muscle tissues of fattening lambs of the Kermani breed. Agricultural Biotechnology Journal, 15(2), 217-236. https://doi.org/10.22103/jab.2023.21524.1486
Sulimova, G.E., Azari, M.A., Rostamzadeh, J., Mohammadabadi, M. R., & Lazebny, O. E. (2007). κ-casein gene (CSN3) allelic polymorphism in Russian cattle breeds and its information value as a genetic marker. Russian Journal of Genetics, 43, 73-79. https://doi.org/10.1134/S1022795407010115
Tarach, P. (2021). Application of polymerase chain reaction-restriction fragment length polymorphism (RFLP-PCR) in the analysis of single nucleotide polymorphisms (SNPs). Acta Universitatis Lodziensis. Folia Biologica et Oecologica, 17, 48–53. https://doi.org/10.18778/1730-536X.17.06
Vijayaraghavan, P., Primiya, R., & Vincent, S. (2013). Thermostable alkaline phytase from Alcaligenes sp. in improving bioavailability of phosphorus in animal feed: In vitro analysis. ISRN Biotechnology, 2013, Article 394305. https://doi.org/10.5402/2013/394305
Agricultural Biotechnology Journal
Volume 17, Issue 2
May 2025
Pages 263-284

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