Whole genome sequencing of the multidrug-resistant pathogen Citrobacter werkmanii recovered from a urinary tract infection patient (case in Mosul, Iraq)

S. M. Alomari; A. N. Al-Najim; S. G. Fadhil

doi:10.15421/0225038

S. M. Alomari Northern Technical University
A. N. Al-Najim Northern Technical University
S. G. Fadhil Northern Technical University

Keywords: antibiotic resistance, Citrobacter werkmanni, whole genome sequencing.

Abstract

Citrobacter werkmanii is an emerging opportunistic human pathogen increasingly widespread in poor nations, r e sponsible for wound, urinary tract, and bloodstream infections. The whole genome sequence of C. werkmanii SAS had a size of 5,072,546 bp and yield ed a GC content of 51.99% distributed within 61 contigs; the largest contig was 941,705 bp, and the smallest was 528 bp with an N 50 value of 351,029. Using the Rapid Annotation System Technology (RAST) server, 4793 coding sequences were detected in addition to 71 RNA genes from different categories. The phylogenetic taxonomy tree of C. werkmanii SAS, generated using the Type Strain Genome Server (TYGS), identified the closest type strains as C. cronae Tue2-1T (accession number: NZ_VOSQ01000380.1). A search for antibiotic resistance genes was conducted in the genome of C. werkmanni SAS utilizing the Comprehensive Antibiotic Resistance Database (CARD). The findings indicated that the genome harbored five key genes associated with resistance to various classes of antibiotics, such as fluoroquinolone, cephamycin, cephalosporin, glycopeptides, polypeptide antibiotics, and tetracyclines. A consi s tent GC content was noted in the SAS genome; however, regions with low GC content were identified at multiple loc a tions. Those regions are an indicative of a possible horizontal gene transfer or insertions in such genomic locations.

References

Abdulrazzaq, S. E., Faisal, R. M., & Hazem, E. G. (2024). Plasposon mutagenesis in Klebsiella pneumoniae isolates reveals the function of a hypothetical protein in biofilm formation. Malaysian Journal of Microbiology, 20(6), 175–184.

Aguirre-Sánchez, J. R., Quiñones, B., Ortiz-Muñoz, J. A., Prieto-Alvarado, R., Vega-López, I. F., Martínez-Urtaza, J., Lee, B. G., & Chaidez, C. (2023). Comparative genomic analyses of virulence and antimicrobial resistance in Citrobacter werkmanii, an emerging opportunistic pathogen. Microorganisms, 11(8), 2114.

Alcock, B. P., Raphenya, A. R., Lau, T. T. Y., Tsang, K. K., Bouchard, M., Edalatmand, A., Huynh, W., Nguyen, A. V., Cheng, A. A., Liu, S., Min, S. Y., Miroshnichenko, A., Tran, H. K., Werfalli, R. E., Nasir, J. A., Oloni, M., Speicher, D. J., Florescu, A., Singh, B., Faltyn, M., … McArthur, A. G. (2020). CARD 2020: Antibiotic resistome surveillance with the comprehensive antibiotic resistance database. Nucleic Acids Research, 48(D1), D517–D525.

An, J., Guo, L., Zhou, L., Ma, Y., Luo, Y., Tao, C., & Yang, J. (2016). NDM-producing Enterobacteriaceae in a Chinese hospital, 2014–2015: Identification of NDM-producing Citrobacter werkmanii and acquisition of blaNDM-1-carrying plasmid in vivo in a clinical Escherichia coli isolate. Journal of Medical Microbiology, 65(11), 1253–1259.

Aurilio, C., Sansone, P., Barbarisi, M., Pota, V., Giaccari, L. G., Coppolino, F., Barbarisi, A., Passavanti, M. B., & Pace, M. C. (2022). Mechanisms of action of carbapenem resistance. Antibiotics, 11(3), 421.

Aziz, R. K., Bartels, D., Best, A. A., DeJongh, M., Disz, T., Edwards, R. A., Formsma, K., Gerdes, S., Glass, E. M., Kubal, M., Meyer, F., Olsen, G. J., Olson, R., Osterman, A. L., Overbeek, R. A., McNeil, L. K., Paarmann, D., Paczian, T., Parrello, B., Pusch, G. D., … Zagnitko, O. (2008). The RAST server: Rapid annotations using subsystems technology. BMC Genomics, 9, 75.

Bankevich, A., Nurk, S., Antipov, D., Gurevich, A. A., Dvorkin, M., Kulikov, A. S., Lesin, V. M., Nikolenko, S. I., Pham, S., Prjibelski, A. D., Pyshkin, A. V., Sirotkin, A. V., Vyahhi, N., Tesler, G., Alekseyev, M. A., & Pevzner, P. A. (2012). SPAdes: A new genome assembly algorithm and its applications to single-cell sequencing. Journal of Computational Biology, 19(5), 455–477.

Campana, E. H., Kraychete, G. B., Montezzi, L. F., Xavier, D. E., & Picão, R. C. (2022). Description of a new non-Tn4401 element (NTEKPC-IIe) harboured on IncQ plasmid in Citrobacter werkmanii from recreational coastal water. Journal of Global Antimicrobial Resistance, 29, 207–211.

Fathi, N. A., & Faisal, R. M. (2024). Efficiency of HiCrome Acinetobacter Agar in identifying A. baumannii from different clinical specimens. Rafidain Journal of Science, 33(4), 34–42.

Forsythe, S. J., Abbott, S. L., & Pitout, J. (2015). Klebsiella, Enterobacter, Citrobacter, Cronobacter, Serratia, Plesiomonas, and other Enterobacteriaceae. In: Jorgensen, J. H., Carroll, K. C., Funke, G., Pfaller, M. A., Landry, M. L., Richter, S. S., & Warnock, D. W. (Eds.). Manual of clinical microbiology. 11th Edition. Wiley. Pp. 714–737.‏

Gurevich, A., Saveliev, V., Vyahhi, N., & Tesler, G. (2013). QUAST: Quality assessment tool for genome assemblies. Bioinformatics, 29(8), 1072–1075.

Ibraheem, M. A., & Faisal, R. M. (2025). Whole genome sequencing of the multidrug-resistant Proteus mirabilis MORAY37 recovered from a urinary tract infection case in Mosul / Iraq. Rwanda Medical Journal, 82(1), 28–37.

Ibrahim, M. A., & Faisal, R. M. (2024). Molecular characterization of antibiotic resistance and virulence genes on plasmids of Proteus mirabilis isolated from urine samples of hospitals in Mosul City, Iraq. Journal of Applied and Natural Science, 16(2), 830–841.

Jabeen, I., Islam, S., Hassan, A. K. M. I., Tasnim, Z., & Shuvo, S. R. (2023). A brief insight into Citrobacter species – a growing threat to public health. Frontiers in Antibiotics, 2, 1276982.

Jean, S. S., Harnod, D., & Hsueh, P. R. (2022). Global threat of carbapenem-resistant gram-negative bacteria. Frontiers in Cellular and Infection Microbiology, 12, 823684.

Lefort, V., Desper, R., & Gascuel, O. (2015). FastME 2.0: A comprehensive, accurate, and fast distance-based phylogeny inference program. Molecular Biology and Evolution, 32(10), 2798–2800.

Meier-Kolthoff, J. P., & Göker, M. (2019). TYGS is an automated high-throughput platform for state-of-the-art genome-based taxonomy. Nature Communications, 10(1), 2182.

Meier-Kolthoff, J. P., Carbasse, J. S., Peinado-Olarte, R. L., & Göker, M. (2022). TYGS and LPSN: A database tandem for fast and reliable genome-based classification and nomenclature of prokaryotes. Nucleic Acids Research, 50(D1), D801–D807.

Mullineaux-Sanders, C., Sanchez-Garrido, J., Hopkins, E. G. D., Shenoy, A. R., Barry, R., & Frankel, G. (2019). Citrobacter rodentium-host-microbiota interactions: Immunity, bioenergetics and metabolism. Nature Reviews, Microbiology, 17(11), 701–715.

Overbeek, R., Olson, R., Pusch, G. D., Olsen, G. J., Davis, J. J., Disz, T., Edwards, R. A., Gerdes, S., Parrello, B., Shukla, M., Vonstein, V., Wattam, A. R., Xia, F., & Stevens, R. (2014). The SEED and the rapid annotation of microbial genomes using subsystems technology (RAST). Nucleic Acids Research, 42, D206–D214.

Papp, M., & Solymosi, N. (2022). Review and comparison of antimicrobial resistance gene databases. Antibiotics, 11(3), 339.

Peter, S., Bezdan, D., Oberhettinger, P., Vogel, W., Dörfel, D., Dick, J., Marschal, M., Liese, J., Weidenmaier, C., Autenrieth, I., Ossowski, S., & Willmann, M. (2018). Whole-genome sequencing enabling the detection of a colistin-resistant hypermutating Citrobacter werkmanii strain harbouring a novel metallo-β-lactamase VIM-48. International Journal of Antimicrobial Agents, 51(6), 867–874.

Prestinaci, F., Pezzotti, P., & Pantosti, A. (2015). Antimicrobial resistance: A global multifaceted phenomenon. Pathogens and Global Health, 109(7), 309–318.

Rafiqul, I. (2012). Isolation, characterization and genome sequencing of a soil-borne Citrobacter freundii strain capable of detoxifying trichothecene mycotoxins. University of Guelph, Guelph.

Saeed, H. K., Alsammak, E. G., & Haddad, M. F. (2024). Draft genome sequencing of Microcoleus sp. HI-ES isolated from freshwater in Iraq: Cyanobacterial strain. Biomedical and Biotechnology Research Journal, 8(1), 129–134.

Stothard, P., Grant, J. R., & Van Domselaar, G. (2019). Visualizing and comparing circular genomes using the CGView family of tools. Briefings in Bioinformatics, 20(4), 1576–1582.

Tamura, K., Stecher, G., & Kumar, S. (2021). MEGA11: Molecular evolutionary genetics analysis version 11. Molecular Biology and Evolution, 38(7), 3022–3027.

Thapa, B., Adhikari, P., Mahat, K., Chhetri, M. R., & Joshi, L. N. (2009). Multidrug-resistant nosocomial Citrobacter in a hospital in Kathmandu. Nepal Medical College Journal, 11(3), 195–199.

Younis, R. M., & Faisal, R. M. (2024). Plasposon mutagenesis in Pseudomonas aeruginosa isolates illustrates the role of ABC transporter in intrinsic resistance to antibiotics. Journal of Applied and Natural Science, 16(3), 1256–1264.

Zhou, G., Peng, H., Wang, Y. S., Huang, X. M., Xie, X. B., & Shi, Q. S. (2017). Complete genome sequence of Citrobacter werkmanii strain BF-6 isolated from industrial putrefaction. BMC Genomics, 18(1), 765.

Zhou, X., Lei, D., Tang, J., Wu, M., Ye, H., & Zhang, Q. (2022). Whole genome sequencing and analysis of fenvalerate degrading bacteria Citrobacter freundii CD-9. AMB Express, 12(1), 51.