Kristina Kokotović1*, Dragan Radnović1, Marijana Kragulj Isakovski2, Jelena Beljin2, Nina Đukanović2, Stanko Milić3, Snežana Maletić2 and Dragana Tamindžija1
1University of Novi Sad, Faculty of Sciences, Department of Biology and Ecology
2University of Novi Sad, Faculty of Sciences, Department of chemistry, biochemistry and environmental protection
3Institute of field and vegetable crops Novi Sad
kristina.kokotovic [at] dbe.uns.ac.rs
Abstract
This study investigated the dynamics of soil microbial communities under phytoremediation in heavy metal-contaminated soil. Soil used for the pot experiment was collected from Srpski Itebej, a site affected by canal dredging and characterized by elevated concentrations of heavy metals. A controlled pot experiment was established with two plant species, Sorghum bicolor (forage sorghum) and Cannabis sativa (hemp), combined with commercial plant growth promoting rhizobacteria and fungi (PGPR) treatments. Samples were collected at the beginning and after three months. Soil DNA was extracted, and 16S rRNA gene amplicon sequencing was performed. Sequence data were processed and analyzed in R using established bioinformatics workflows for ASV inference, taxonomic classification, and community analysis. Total counts of heterotrophic, ammonium-oxidizing, denitrifying, and nitrogen-fixing bacteria were higher in the final samples, in contrast to nitrifiers, which exhibited greater abundance at the start of the experiment. Catalase and acid phosphatase activities were more pronounced at the beginning of the study, whereas alkaline phosphatase and dehydrogenase activities were slightly more prominent at the conclusion of the experiment. Alpha diversity analyses revealed higher diversity at the beginning of the experiment, particularly in control samples and those treated with PGPR. Beta diversity analysis indicated that dominant taxa were broadly similar across samples and timepoints; however, Thermoproteota consistently dominated, while Cyanobacteriota were more abundant in samples collected at the end of the experiment. Principal coordinate analysis (PCoA) based on Bray-Curtis distances revealed clear separation of samples according to both treatment and experimental stage. PERMANOVA confirmed that both treatment and time significantly influenced microbial community structure. Principal component analysis (PCA) integrating chemical and microbial parameters showed that heavy metals were the dominant drivers at the initial stage of the experiment. In contrast, reduced metal concentrations at the final stage were associated with recovery and stabilization of functional microbial groups, including denitrifiers and ammonifiers. Overall, the results demonstrate that phytoremediation can promote the recovery and stabilization of soil microbial communities under heavy metal stress, highlighting the importance of integrative bioinformatics approaches for understanding microbe-environment interactions.
Keywords: diversity, taxonomic composition, 16S rRNA
Acknowledgement: This research was supported by the Science Fund of the Republic of Serbia, #GRANT No 6769, Natural based efficient solution for remediation and revitalization of contaminated locations using energy crops – ReNBES.