Investigation of Water Quality and Molecular Analysis (16s rRNA) of Bacterial Community Structure in a Rural Geothermal Spring: Ucler Hill (Azat Village), Kars, Türkiye

Abstract

This study was carried out to evaluate the chemical and microbiological properties of a natural water source located in Azat Village of Kars province and used as drinking water by the surrounding villagers. Water samples taken from three different sampling points were analyzed for pH, electrical conductivity, total hardness, alkalinity, nitrogenous compounds (NO₂-, NO₃-, NH₄⁺), metal contents (Fe, Cu, Al, Pb), sulfate and chloride levels. The data obtained were compared with World Health Organization (WHO) drinking water standards and one-sample t-test was applied for each parameter. Statistically significant differences were found in total hardness, NO₃-, Fe, Cu, SO₄²- and Cl- parameters (p < 0.05), while other parameters were found to be compatible with WHO reference values. In microbiological analyses, a total of eight different bacterial species were isolated by culture-dependent methods. These species were identified by 16S rRNA gene sequencing analysis and all of them were determined to be non-pathogenic environmental microorganisms. The identified species included Microbacterium spp., Bacillus spp., Corynebacterium spp., Lactobacillus spp., Rhodococcus spp., Pseudomonas spp., Leuconostoc spp. and Aeromonas spp. Although some species have the potential to be opportunistic pathogens, the overall microbiological profile was considered to be within acceptable limits for natural sources.

Keywords

• : Natural drinking water source
• Microbial diversity
• Rural water security
• Chemical water quality
• 16S rRNA sequence analysis

Kırsal Bir Jeotermal Kaynakta Su Kalitesi ve Moleküler Analiz (16s rRNA) ile Bakteriyel Topluluk Yapısının İncelenmesi: Üçler Tepesi (Azat Köyü), Kars, Türkiye

Abstract

Bu çalışma, Kars ilinin Azat Köyü'nde bulunan ve çevre köylüler tarafından içme suyu olarak kullanılan doğal bir su kaynağının kimyasal ve mikrobiyolojik özelliklerini değerlendirmek amacıyla gerçekleştirilmiştir. Üç farklı örnekleme noktasından alınan su örnekleri pH, elektriksel iletkenlik, toplam sertlik, alkalinite, azotlu bileşikler (NO₂-, NO₃-, NH₄⁺), metal içerikleri (Fe, Cu, Al, Pb), sülfat ve klorür seviyeleri açısından analiz edilmiştir. Elde edilen veriler Dünya Sağlık Örgütü (WHO) içme suyu standartları ile karşılaştırılmış ve her parametre için tek örneklem t-testi uygulanmıştır. Toplam sertlik, NO₃-, Fe, Cu, SO₄²- ve Cl- parametrelerinde istatistiksel olarak anlamlı farklılıklar bulunurken (p<0.05), diğer parametreler WHO referans değerleriyle uyumlu bulunmuştur. Mikrobiyolojik analizlerde, kültüre bağlı yöntemlerle toplam sekiz farklı bakteri türü izole edilmiştir. Bu türler 16S rRNA gen dizileme analizi ile tanımlanmış ve hepsinin patojen olmayan çevresel mikroorganizmalar olduğu belirlenmiştir. Tanımlanan türler arasında Microbacterium spp., Bacillus spp., Corynebacterium spp., Lactobacillus spp., Rhodococcus spp., Pseudomonas spp., Leuconostoc spp. ve Aeromonas spp. yer almaktadır. Bazı türler fırsatçı patojen olma potansiyeline sahip olsa da, genel mikrobiyolojik profilin doğal kaynaklar için kabul edilebilir sınırlar içinde olduğu düşünülmüştür.

Keywords

• Doğal içme suyu kaynağı
• Mikrobiyal çeşitlilik
• Kırsal su güvenliği
• Kimyasal su kalitesi
• 16S rRNA dizi analizi

References

1. Mishra, R.K., Fresh water availability and its global challenge, British Journal of Multidisciplinary and Advanced Studies 4(3) 2023; 1-78.
2. Till, D., G. McBride, A. Ball, K. Taylor, E. Pyle, Large-scale freshwater microbiological study: rationale, results and risks, Journal of Water and Health 6(4) 2008; 443-460.
3. Verma, S., S. Verma, R. Ramakant, V. Pandey, A. Verma, Comprehensive Assessment of Physico-Chemical and Biological Parameters in Water Quality Monitoring: A Review of Contaminants, Indicators, and Health Impacts, International Journal of Horticulture, Agriculture and Food science 9(2) 2025; 611019.
4. Ulucay, O., Potential Health Risks of the Çakmak Village Thermal Spring (Turkey, Kars) for Grazing Animals: Characterization of Bacterial Communities and Water Quality, Journal of Environmental Chemical Engineering 2025; 118059.
5. Reysenbach, A.-L., E. St. John, J. Meneghin, G.E. Flores, M. Podar, N. Dombrowski, A. Spang, S. L’Haridon, S.E. Humphris, C.E.J. De Ronde, Complex subsurface hydrothermal fluid mixing at a submarine arc volcano supports distinct and highly diverse microbial communities, Proceedings of the National Academy of Sciences 117(51) 2020; 32627-32638.
6. Uluçay, O., A. Görmez, C. Öziç, Determination of Physical, Chemical Properties of the Hot Springs Waters and Comparison of Bacteria: Hot Springs in the East and Southeast Anatolia Region of Turkey, Journal of Agriculture 4(1) 2021; 24-29.
7. Hamilton, T.L., E. Koonce, A. Howells, J.R. Havig, T. Jewell, J.R. de la Torre, J.W. Peters, E.S. Boyd, Competition for ammonia influences the structure of chemotrophic communities in geothermal springs, Appl Environ Microb 80(2) 2014; 653-661.
8. Dippong, T., M.-A. Resz, Heavy metal contamination assessment and potential human health risk of water quality of lakes situated in the protected area of Tisa, Romania, Heliyon 10(7) 2024.

9. Lynch, R.C., A.J. King, M.E. Farías, P. Sowell, C. Vitry, S.K. Schmidt, The potential for microbial life in the highest‐elevation (> 6000 masl) mineral soils of the Atacama region, Journal of Geophysical Research: Biogeosciences 117(G2) 2012.
10. Smrhova, T., K. Jani, P. Pajer, G. Kapinusova, T. Vylita, J. Suman, M. Strejcek, O. Uhlik, Prokaryotes of renowned Karlovy Vary (Carlsbad) thermal springs: phylogenetic and cultivation analysis, Environmental Microbiome 17(1) 2022; 48.
11. Arvanitidou, M., A. Papa, T.C. Constantinidis, V. Danielides, V. Katsouyannopoulos, The occurrence of Listeria spp. and Salmonella spp. in surface waters, Microbiological Research 152(4) 1997; 395-397.
12. Ghilamicael, A.M., H.I. Boga, S.E. Anami, T. Mehari, N.L.M. Budambula, Potential human pathogenic bacteria in five hot springs in Eritrea revealed by next generation sequencing, PLoS One 13(3) 2018; e0194554.
13. Rahel, C., R. Adriyani, H.A. Nurfitria, Health risk in hot springs: A literature review, Journal of Health Science and Prevention 5(2) 2021; 88-99.
14. Deza, M.A., M. Araujo, M.J. Garrido, Inactivation of Escherichia coli, Listeria monocytogenes, Pseudomonas aeruginosa and Staphylococcus aureus on stainless steel and glass surfaces by neutral electrolysed water, Lett Appl Microbiol 40(5) 2005; 341-346.
15. Ulucay, O., A. Gormez, C. Ozic, Identification, characterization and hydrolase producing performance of thermophilic bacteria: geothermal hot springs in the Eastern and Southeastern Anatolia Regions of Turkey, Antonie van Leeuwenhoek 2022; 1-18.
16. Uluçay, O., & Öziç, C. (2025). Molecular identification and phylogenetic profiling of keratinolytic bacteria isolated from goose and chicken farms in a cold region of Türkiye. International Journal of Agriculture Environment and Food Sciences, 9(2), 436-445. https://doi.org/10.31015/2025.2.17
17. Kars Maps, 2025. https://www.harita.gov.tr/urun/kars-fiziki-il-haritasi/382. (Accessed 25.06.2025 2025).
18. Edition, F., Guidelines for drinking-water quality, WHO chronicle 38(4) 2011; 104-8.
19. Oztas Gulmus, E., A. Gormez, Identification and characterization of novel thermophilic bacteria from hot springs, Erzurum, Turkey, Curr Microbiol 77(6) 2020; 979-987.
20. Ahmadpour, A., Y. Ghosta, Z. Alavi, F. Alavi, L.M. Hamidi, P.H. Rampelotto, Morphological and Molecular Characterization of a New Section and Two New Species of Alternaria from Iran, Life 15(6) 2025; 870.
21. Javadi, A., M. Shamaei, L.M. Ziazi, M. Pourabdollah, A. Dorudinia, S.M. Seyedmehdi, S. Karimi, Qualification study of two genomic DNA extraction methods in different clinical samples, Tanaffos 13(4) 2014; 41.
22. Elkins, K.M., Determination of Quality and Quantity of DNA Using Agarose Gel Electrophoresis, Forensic DNA Biology. Oxford, USA: Elsevier Inc 2013; 53-57.
23. Gurcan, K., F. Demirel, M. Tekin, S. Demirel, T. Akar, Molecular and agro-morphological characterization of ancient wheat landraces of turkey, Bmc Plant Biol 17(Suppl 1) 2017; 171.
24. Chen, C., P.H. Qu, Q. Gu, B. Huang, W.Z. Zhang, X.P. Mu, L. Zhang, M.R. Chen, L.X. Wang, Y.J.Y. E Sm, Application of broad-spectrum PCR amplification and direct sequencing for identification of the infrequent bacterial cultures from clinical sources, targeting the bacterial 16S rRNA gene with universal primers, Chin J Lab Med 35 2012; 612-619.
25. Drancourt, M., C. Bollet, A. Carlioz, R. Martelin, J.-P. Gayral, D. Raoult, 16S ribosomal DNA sequence analysis of a large collection of environmental and clinical unidentifiable bacterial isolates, Journal of clinical microbiology 38(10) 2000; 3623-3630.
26. Meyers, L.S., G.C. Gamst, A.J. Guarino, Performing data analysis using IBM SPSS, John Wiley & Sons2013.
27. Sahoo, P.K., K. Kim, M.A. Powell, Managing groundwater nitrate contamination from livestock farms: implication for nitrate management guidelines, Current Pollution Reports 2(3) 2016; 178-187.
28. Lawniczak, A.E., J. Zbierska, B. Nowak, K. Achtenberg, A. Grześkowiak, K. Kanas, Impact of agriculture and land use on nitrate contamination in groundwater and running waters in central-west Poland, Environmental monitoring and assessment 188(3) 2016; 172.
29. Crerar, D., S. Wood, S. Brantley, A. Bocarsly, Chemical controls on solubility of ore-forming minerals in hydrothermal solutions, The Canadian Mineralogist 23(3) 1985; 333-352.
30. Zampella, M., P. Adamo, L. Caner, S. Petit, D. Righi, F. Terribile, Chromium and copper in micromorphological features and clay fractions of volcanic soils with andic properties, Geoderma 157(3-4) 2010; 185-195.
31. Farrar Jr, W.E., Serious infections due to “non-pathogenic” organisms of the genus bacillus: Review of their status as pathogens, The American journal of medicine 34(1) 1963; 134-141.
32. Hajmeer, M.N., D.Y.C. Fung, Infections with other bacteria, Foodborne Infections and Intoxications (3rd Ed.) Elsevier, UK 2006; 341-367.
33. Oliveira, A., L.C. Oliveira, F. Aburjaile, L. Benevides, S. Tiwari, S.B. Jamal, A. Silva, H.C.P. Figueiredo, P. Ghosh, R.W. Portela, Insight of genus Corynebacterium: ascertaining the role of pathogenic and non-pathogenic species, Frontiers in microbiology 8 2017; 1937.
34. Ivshina, I.B., M.S. Kuyukina, A.V. Krivoruchko, E.A. Tyumina, Responses to ecopollutants and pathogenization risks of saprotrophic Rhodococcus species, Pathogens 10(8) 2021; 974.
35. Janda, J.M., S.L. Abbott, The genus Aeromonas: taxonomy, pathogenicity, and infection, Clinical microbiology reviews 23(1) 2010; 35-73.
36. Fernández-Bravo, A., M.J. Figueras, An update on the genus Aeromonas: taxonomy, epidemiology, and pathogenicity, Microorganisms 8(1) 2020; 129.
37. De Bentzmann, S., P. Plésiat, The Pseudomonas aeruginosa opportunistic pathogen and human infections, Environmental microbiology 13(7) 2011; 1655-1665.