A sustainable environment relies on the hydrosphere, which covers approximately 75% of the Earth’s crust, yet only a limited amount of it is available as freshwater [1] . While a significant portion of the Earth’s surface is covered by water, merely 3% of that is freshwater, and the majority of terrestrial organisms cannot directly use the salty water that constitutes the rest [2] . The availability of freshwater is crucial for the survival of terrestrial forms as well as other forms of life. Lakes, rivers, streams, and wetlands are examples of freshwater ecosystems that provide humans with valuable ecosystem services, including drinking water, fishing, and enjoyment. Water bodies serve as natural barriers between people and different kinds of organisms, but they can be contaminated by a variety of sources, which can change the water’s quality and condition [3] . Therefore, protecting aquatic ecosystems and priceless water resources is crucial [1] . In freshwater habitats, the activities of autotrophic and heterotrophic microbes regulate the natural cycles of oxygen, carbon, and other nutrients [4] .

Freshwater ecosystems harbor vast microbial communities, playing critical roles in nutrient cycling, ecosystem stability, and overall ecological balance [5] . According to Fierer et al. [6] , these communities comprise bacteria, archaea, fungi, viruses, and other micro-eukaryotes, collectively forming a complex microbial consortium that drives essential biogeochemical processes. Studying the composition, structure, and functional potential of these communities is crucial for understanding the dynamics of freshwater ecosystems and their responses to environmental perturbations [7] [8] . The state of freshwater resources has deteriorated considerably in recent times, and the impact of water pollution events on the safety of water quality and the well-being of local populations is now being recognized on a global scale [1] . Presently, no studies investigating the microbial community structure of the freshwater bodies in Enugu State, Nigeria are available. Even though data regarding the composition of microbial communities plays a crucial role in freshwater ecosystems, such studies subjected to other water bodies of different regions of the world are available in the literature. Traditionally, the study of microbial communities relied on culture-based approaches, which were limited in capturing the true extent of diversity, mostly due to the nonculturable nature of most microorganisms [9] . Thus, the advent of high-throughput sequencing technologies, particularly metagenomics, has revolutionized the ability to explore the genetic and functional diversity of microorganisms, including the VBNC (viable but nonculturable), within their natural habitats [10] [11] .

Notably, the cumulative effects of anthropogenic stress on freshwater ecosystems are becoming more noticeable and concerning. In addition to natural stressors, anthropogenic stressors like increased water use, overuse of hydrobiological resources, nutrient inputs, hydrological stress, and the disposal of industrial, urban, and agricultural waste are damaging freshwater ecosystems [12] [13] . The accelerated changes in our ecosystem due to climate change and anthropogenic activities have created an urgent need for monitoring, empirical studies, and conservation of our biodiversity [14] . Thus, one of the key steps towards ensuring healthy conditions of a freshwater ecosystem is to have a good understanding of its microbial community structure [15] . According to Brar et al. [1] , there is a growing demand for routine aquatic ecosystem monitoring and assessment in order to take further actions to maintain the sustainability of these resources.

In recent years, metagenomic studies have shed light on the vast diversity of freshwater microbial communities, revealing a huge reservoir of untapped genetic resources [16] . Furthermore, this technique has enabled the identification of key functional attributes, such as nutrient cycling, secondary metabolite production, and stress response mechanisms, which are crucial for understanding the ecological roles of these microorganisms [17] - [19] . Quince et al. [20] emphasized that by directly sequencing DNA extracted from environmental samples, metagenomics offers a culture-independent alternative that allows for the comprehensive characterization of microbial communities in their entirety. Therefore, this study seeks to address this gap by conducting a comparative analysis of freshwater bodies’ microbial communities from Enugu State, Nigeria using metagenomic approaches. The complete microbial profile of the three freshwater bodies studied is analyzed for the first time to the best of our knowledge. Thus, by characterizing their taxonomic composition and metabolic diversity, we aim to gain deeper insights into the factors shaping the structure and ecological functions of these microorganisms.

Our research offers the initial descriptive examination of the microbial community profiles in the freshwater bodies of Enugu East, which is an essential step in understanding the microbial ecology of reservoirs. Microbial structures or communities, primarily bacteria, of the water system contribute significantly to the aquatic ecosystem due to their role in biogeochemical processes [23] . In recent years, with the development of industrialization, the lake environment has deteriorated to varying degrees, and lake health has become a hot research topic [24] [25] . The result of the analysis of the taxonomic community showed that the freshwater bodies were dominated by prokaryotic organisms (92% bacteria and 8% archaea). This distribution highlights prokaryotes’ importance in maintaining ecological balance and their potential roles in biogeochemical processes within freshwater ecosystems. Prokaryotes are considered the main drivers of transformation and the cycling of most biologically active elements in aquatic ecosystems via the degradation and mineralization of organic compounds to their inorganic constituents. The abundance of bacteria in the present findings agrees with other reports. Bacteria are the most common organisms on Earth; their makeup and abundance significantly influence ecosystem functions and stability, whether they exist in host-associated communities, soil, grasslands, or oceans [1] .

The two most dominant phyla in the present study are Proteobacteria and Acidobacteria. The abundance of the two phyla may be attributed to human interference and the discharge of human wastes in freshwater bodies. The dominance of these groups is in agreement with other freshwater studies [26] - [28] . Therefore, utilizing the studied freshwater bodies (Iyiube lake, Abala lake, and Iyioku river) for drinking water presents significant health hazards for residents of those regions, as human activities contribute to environmental contamination. Proteobacteria, which primarily consist of gram-negative bacteria, have previously been identified as the dominant taxa in surface water bodies defined by anthropogenic pollution and waste disposal [29] . Additionally, it has been observed that the phylum Proteobacteria is predominantly found in environments with low salinity and minimal electrical conductivity [3] . Proteobacteria mainly consist of rapidly growing copiotrophs that flourish in nutrient-rich environments, and they are believed to play an essential role in nitrogen cycling, connecting iron-carbon biogeochemistry, carbon sequestration, nutrient flux, and various biochemical processes [1] [30] .

Among the five classes of Proteobacteria (alpha, beta, gamma, delta, and Epsilon), the most abundant across all the freshwater bodies in this study is gamma-proteobacteria. The abundance of gamma-proteobacteria in freshwater bodies has been previously reported by Betiku et al. [31] . Gamma-proteobacteria comprise families of N-fixing bacteria [32] and favor high pH [33] . Gamma-proteobacteria are transient members of the bacterioplankton, brought in by surrounding environments, and the members of the group are naturally occurring in freshwater environments that are probably copiotrophs, which could be one of the main reasons for their disproportionate distribution among the studied sites [34] . Phylum Acidobacteriota has been reported in previous studies in freshwater lakes to thrive in settings with acidic systems [34] , limited nutritional availability, slower growth rates, high tolerance to toxic compounds [35] , and sulfidic conditions [36] . A significant prevalence of Acidobacteria has been documented in a water stream in Japan [37] .

Additionally, other prevalent phyla identified in all three freshwater ecosystems examined included Chloroflexi, Bacteroidota, Verrucomicrobiota, Myxococcota, and Nitrospirota, which align with observations from various other studies on freshwater bodies [3] [31] [38] . Similar trends in the prevalence of the primary bacterial groups have been observed globally across various freshwater ecosystems, even with differences in hydrology and water chemistry across the locations examined [39] [40] . Verrucomicrobia identified in examined freshwater ecosystems have been shown to possess a strong ability to degrade various forms of carbon, such as xylan, cellulose, pectin, sugars, and methane, and they play a beneficial role in nitrogen fixation in soils and polysaccharide breakdown in termites [31] [41] .

The identification of the phyla Fusobacteriota and Sumerlaeota for Abala Lake and DTB120, Deferrisomatota, and Hydrogenedentes for Iyioku River that are specific to a single location suggests that the freshwater bodies differ in terms of pollution, nutrient levels, pH, oxygen availability, and human influence, all of which affect the composition of the microbial communities. Fusobacteriota are rod-shaped, anaerobic, Gram-negative bacteria that are well-known for their capacity to flourish in low oxygen conditions [42] . While Sumerlaeota have been confirmed to exist in diverse natural and artificial environments, including freshwater and marine sediments, they are still not well understood because there are not enough cultured specimens [43] . The phylum Deferrisomatota is enriched in anoxic and highly salinized environments and is involved in anaerobic respiration and metal cycling, particularly the reduction of iron and sulfur. The discovery of the candidate phylum DTB120, which is mainly known from metagenomic data and lacks cultured representatives, emphasizes the hidden microbial diversity and the possibility of new metabolic capacities that have not yet been identified. McAllister et al. [44] noted DTB120 is still uncharacterized and was identified using metagenomics analysis.

Although there were some anticipated variations among the three freshwater sources from different locations, the findings revealed overlap in the composition of the microbial communities across the analyzed freshwater bodies. These findings align with other research that indicates freshwater bodies harbor similar microbial communities [29] [31] . Our study found that the alpha diversity at Abala Lake was lower than that of the other freshwater bodies examined and indicated a significantly lower level in comparison to the Iyioku River. The results highlight the abundance of microbial diversity in the Iyioku River. The Iyioku River represents a typical river-fed ecosystem, and the higher diversity here is probably due to the influx of foreign bacterial populations from other smaller freshwater sources that feed into the river.

Furthermore, a significant difference was also observed in the beta diversity between the Iyioku river and the other studied freshwater bodies. These data all demonstrated that the bacterial community structure of the Iyioku river was quite different from that of the Abala lake. Iyiube lakes show the highest abundance of Acidobacteria when compared to other sites, revealing it to be the most polluted site because of human interference and the discharge of human wastes near the lake bank. These observed human activities could have an impact on the river’s aquatic microbiome and fish health, based on a study that showed host-habitat as a primary determinant of gut microbiome in fish [45] . Adedire et al. [29] reported that the abundance of Acidobacteriota at the Idah river in Kogi State, Nigeria is as a result of industrial pollutants in the surrounding area. Future research on the studied freshwater bodies (Abala lake, Iyiube lake, and Iyioku river) should integrate metatranscriptomics to uncover microbial functions and pathways, while culture-based approaches remain essential for characterizing the ecological roles of uncultured or poorly defined taxa. Thus, the study was restricted to 16S rRNA-based taxonomic profiling of prokaryotic communities and did not capture other microbial groups such as fungi and protists, highlighting an important avenue for future investigation.

The initial metagenomic analysis of three freshwater ecosystems (Iyiube Lake, Abala Lake, and Iyioku River) in Enugu East, Nigeria, showed that Proteobacteria and Acidobacteria were the predominant phyla across all sampling sites, suggesting pollution from community waste, human sewage, and other anthropogenic sources. The identification of Acidobacteria in Iyiube Lake indicates that it is the most contaminated among the tested freshwater bodies, with a significant amount of sewage disposal. The three studied freshwater bodies (Iyiube Lake, Abala Lake, and Iyioku River) should not be used for drinking water due to environmental contamination, as consuming them poses substantial health risks for locals. Given that the water is used for agricultural purposes in growing vegetables, it is important that it be purified of pollutants through natural methods (slow sand and cloth filtration). The results of this study could serve as a reference or case study for future research on freshwater microbiomes and provide insights into the microbial communities of these three freshwater bodies.

The authors are thankful to the DNA Learning Center, New York, United States of America, for funding part of this project.