Investigating the interplay of indoor microbial diversity with pollutant variables and human health profile in Indian slums: a metagenomic approach

Microbial diversity within the airway microbiome in chronic pediatric lung diseases

Every year at the onset of winter season (October-November), crop residue/parali/stubble burning starts in Punjab and Haryana, leading to heavy air pollution in Delhi, and adversely affecting human and environmental health. During this time, the combination of unfavourable meteorological conditions, additional emissions from stubble burning, and firework activities in this area causes the air quality to furtherdeteriorate. In this study, we have attempted to understand the influence of parali and firecracker incidents on air pollutants' variability over Delhi during the last three years (2020 to 2022). For this purpose, daily average particulate matter and gaseous pollutants data were fetched from the Central Pollution Control Board (CPCB), and daily total fire counts and fire radiative power (FRP) data were retrieved from NASA's Fire Information for Resource Management System (FIRMS). A bigger area of severe burning is suggested by higher FRP values and higher fire counts in the middle of November in all the years considered. Three years satellite-based FIRMS data over Punjab and Haryana show the highest number of active fire counts in 2021 (n = 80,505) followed by 2020 (n = 75,428), and 2022 (n = 49,194). More than 90% parali burning incidents were observed in Punjab state only despite the considerable variability in numbers among the years. The significant effect of parali burning was seen on pollutant concentration variability. As the number of fire count increases or decreases in Punjab and Haryana, there is a corresponding increase or decrease in the particulate matter concentration with a time lag of few days (1 to 2days). The trend in backward air mass trajectories suggests that the variable response time of pollutants' concentration is due to local and distant sources with different air mass speeds. Our estimates suggest that stubble burning contributes 50-75% increment in PM2.5 and 40 to 45% increase in PM10 concentration between October and November. A good positive correlation between PM2.5, PM10, NOX, and CO and fire counts (up to 0.8) suggests a strong influence of stubble burning on air quality over Delhi. Furthermore, the firecracker activities significantly increase the concentration of particulate matter with ~100% increment in PM2.5 and ~55% increment in PM10 mass concentrations for a relatively shorter period (1 to 2days).

Chapter 6.7 - Microbial Communities Driving Pollution Degradation in Contaminated Environments: Metagenomic Insights

Metagenomic diagnostics for the simultaneous detection of multiple pathogens in human stool specimens from Côte d'Ivoire: a proof-of-concept study

Soil pollution occurring at mining sites has adverse impacts on soil microbial diversity. New approaches, such as metagenomics approach, have become a powerful tool to investigate biodiversity of soil microbial communities. In the current study, metagenomics approach was used to investigate the microbial diversity of soils contaminated with different concentrations of lead (Pb) and zinc (Zn). The contaminated soils were collected from a Pb and Zn mine. The soil total DNA was extracted and 16S rDNA genes were amplified using universal primers. The PCR amplicons were sequenced and bioinformatic analysis of metagenomes was conducted to identify prokaryotic diversity in the Pb- and Zn-contaminated soils. The results indicated that the ten most abundant bacteria in all samples were Solirubrobacter (Actinobacteria), Geobacter (Proteobacteria), Edaphobacter (Acidobacteria), Pseudomonas (Proteobacteria), Gemmatiomonas (Gemmatimonadetes), Nitrosomonas, Xanthobacter, and Sphingomonas (Proteobacteria), Pedobacter (Bacterioidetes), and Ktedonobacter (Chloroflexi), descendingly. Archaea were also numerous, and Nitrososphaerales which are important in the nitrogen cycle had the highest abundance in the samples. Although, alpha and beta diversity showed negative effects of Pb and Zn contamination on soil microbial communities, microbial diversity of the contaminated soils was not subjected to a significant change. This study provided valuable insights into microbial composition in heavy metals-contaminated soils.

The discoveries made in the area of microbiology using metagenomic techniques during the last 20 years reveal that we have not been able to cultivate and characterize more than 1% of the microbes present in nature in the 100 years since the birth of microbiology. The “great plate count anomaly” is the discrepancy between populations estimated by dilution plating and by microscopy. If we compare direct microscopic cell count after 4-6-diamidino-2-phenylindole (DAPI) staining of bacteria with the number of the microbial colonies growing on nutrient agar, it appears that in natural samples less than one cell in thousand produces a colony. Aman et al. [1] have reported that only fraction of microorganisms are found to be cultivable in any environmental sample. Accessing this uncultivable microbial resource would not have been possible but for the novel idea proposed by Torsvik et al. [2] and Pace et al. [3] who proposed and demonstrated that DNA can directly be isolated, manipulated and characterized from any environmental sample, ranging from hot springs to Antarctic soil, deep sea vents to clouds and from gut of the insect to human gut. Metagenomics, the term coined by Jo Handlesman in 2004 is defined as the culture independent analysis of microbial genomes [4], 91 years after Bergey categorically suggested that no microbe can be characterized until it is cultivated [5]. The glimpse of microbial diversity of various chosen environmental samples unraveled (left out by routine cultivation) using metagenomics in the last 10 years has been summarized in Fig. 1. Fig. 1 Metagenomic analysis of chosen environmental niches Metagenomics has not only been used to catalogue microbial diversity but it has been a useful technique for isolation of genes encoding novel biomolecules i.e. metaexploration. It is beyond the scope of this article to catalogue the numerous novel genes that have been isolated using metagenomic techniques so far. To give the reader some idea of the usefulness of metagenomics as a potent tool for prospecting the microbial wealth in nature, here is a brief lay down of few products isolated in the last 10 years (Table 1). Table 1 Novel microbial genes isolated by metagenomic approach in the last 10 years The metagenomic analysis of microbiome associated with human body has revealed that only 10% of the cells are human cells and rest are microbial cells. These astonishing results have been published in “Nature” 2010 June issue [17] wherein Zhao had drawn his conclusions primarily on the basis of work done by Qin et al. [18] that has also been published in “Nature” 2010 March issue. Recently, in an interesting study, metagenomics has been used to compare the genes in the microbial communities in the gut of obese mice with their leaner relatives. Researchers transplanted gut microbial communities harvested from both obese and lean animals into germ free wild type mice with the result that the leaner mice gained more fat. So there is an interesting point here that even the calorific value of the food we eat may vary depending upon the composition of the person’s gut microbes [19]. Metagenomic approach was also used to analyze the symbiotic microbial community in the eukaryotic host Olavius algarvensis. Two nearly complete and two partial genomes of the oligochaetes’ predominant symbionts were assembled using shotgun sequencing of a bacteria-enriched sample combined with nucleotide-signature based binning. Metabolic pathway reconstruction from the sequenced genomes revealed all the four symbionts to be capable of autotrophic carbon fixation and provide multiple sources of organic carbon to their host as two of them are sulfur-oxidizing and two sulfate-reducing bacteria [20]. The discoveries made using metagenomic approach about novel microbial genes, genomes and their association with other genomes has changed the way genomes and their functions will be studied now onwards.

The interactive relationship between plant and microbe starts on the surface of the plant, such as on the roots, leaves, or other parts. Various different types of microbes thrive on different parts of the plant. Microbes may also enter plant cells and are then known as endophytes; this relationship consists of an intricate interaction between plant and microbe. This intricate interaction occurs also at a genomic level. The plant–microbiome interaction may be beneficial or harmful for both, and sometimes it may be neutral. The presence of another microbe in the vicinity also changes the relationship between a microbe and its host. Now we understand that this type of communication or interface is very complicated, and, to understand this scenario, we need the help of modern techniques such as a metagenomic approach or next-generation sequencing. The soil and plant microbiome community plays a significant role in providing essential nutrients to the host plant and also in recycling nutrients and carbon in the environment. On the other hand, we do not know much about novel or nonisolated soil microbes; therefore, their functions are unknown to us. Using a metagenomic approach we can reveal the identity of an unknown soil microbe along with its functional gene information. Most of these associate microbes that enter into plant tissues communicate with their host very closely. This interaction influences metabolic aspects, nutritional uptake potential and transport, signaling of hormones, and stress mitigation, ultimately resulting in plant growth and development. In this respect, a metagenomic approach can be proficiently linked to other omics techniques to offer a multifarious picture of in-progress occurrences that exploit the communication between the plant and its total microbiome. The applied and practical purposes of these metagenomic studies, apart from providing a clearer vision of ecological biological diversity and ecological aspects of microbes, is to provide detailed and valuable tactics for enhancing crop production and protection against host pathogens.

Viruses have the greatest abundance and highest genetic diversity in marine ecosystems. The interactions between viruses and their hosts is one of the hot spots of marine ecology. Besides their important role in various ecosystems, viruses, especially bacteriophages and their gene pool, are of enormous interest for the development of new gene products with high innovation value. Various studies have been conducted in diverse ecosystems to understand microbial diversity and phage–host interactions; however, the Black Sea, especially the Eastern coastal area, remains among the least studied ecosystems in this regard. This study was aimed at to fill this gap by analyzing microbial diversity and bacteriophage–host interactions in the waters of Eastern Black Sea using a metagenomic approach. To this end, prokaryotic and viral metagenomic DNA from two sampling sites, Poti and Gonio, were sequenced on the Illumina Miseq platform and taxonomic and functional profiles of the metagenomes were obtained using various bioinformatics tools. Our metagenomics analyses allowed us to identify the microbial communities, with Proteobacteria, Cyanobacteria, Actinibacteria, and Firmicutes found to be the most dominant bacterial phyla and Synechococcus and Candidatus Pelagibacter phages found to be the most dominant viral groups in the Black Sea. As minor groups, putative phages specific to human pathogens were identified in the metagenomes. We also characterized interactions between the phages and prokaryotic communities by determining clustered regularly interspaced short palindromic repeats (CRISPR), prophage-like sequences, and integrase/excisionase sequences in the metagenomes, along with identification of putative horizontally transferred genes in the viral contigs. In addition, in the viral contig sequences related to peptidoglycan lytic activity were identified as well. This is the first study on phage and prokaryote diversity and their interactions in the Eastern coastal area of the Black Sea using a metagenomic approach.

Soil microorganisms play key roles in ecosystem functioning and are known to be influenced by biotic and abiotic factors, such as plant cover or edaphic parameters. New Caledonia, a biodiversity hotspot located in the southwest Pacific, is one-third covered by ultramafic substrates. These types of soils are notably characterised by low nutrient content and high heavy metal concentrations. Ultramafic outcrops harbour diverse vegetation types and remarkable plant diversity. In this study, we aimed to assess soil bacterial and fungal diversity in New Caledonian ultramafic substrates and to determine whether floristic composition, edaphic parameters and geographical factors affect this microbial diversity. Therefore, four plant formation types at two distinct sites were studied. These formations represent different stages in a potential chronosequence. Soil cores, according to a given sampling procedure, were collected to assess microbial diversity using a metagenomic approach, and to characterise the physico-chemical parameters. A botanical inventory was also performed. Our results indicated that microbial richness, composition and abundance were linked to the plant cover type and the dominant plant species. Furthermore, a large proportion of Ascomycota phylum (fungi), mostly in non-rainforest formations, and Planctomycetes phylum (bacteria) in all formations were observed. Interestingly, such patterns could be indicators of past disturbances that occurred on different time scales. Furthermore, the bacteria and fungi were influenced by diverse edaphic parameters as well as by the interplay between these two soil communities. Another striking finding was the existence of a site effect. Differences in microbial communities between geographical locations may be explained by dispersal limitation in the context of the biogeographical island theory. In conclusion, each plant formation at each site possesses is own microbial community resulting from multiple interactions between abiotic and biotic factors.

Plant microbial interactions play a crucial role in agriculture and ecological balancing. It is also being reported that soil health depends upon microbial biodiversity. Several reports have suggested that exploring microbial and bioengineering technologies can be employed to increase farm productivity and reduce environmental pollutants such as greenhouse gas (GHG) emissions. This chapter emphasizes the metagenomics, metatranscriptomics, and meta-metabolomics approaches to provide comprehensive knowledge of the plant microbial interactions necessary for newer interventions to increase the health of the soil. Further, metagenomics approaches provide a valuable tool to identify and select novel microbial or gene resources with beneficial traits, such as the discovery of novel genes from ecological niches, including meadows, crop fields, and others. With metagenomic approaches, a new dimension of characterization of the complex microbial community can be attained. Metagenomic approaches can be used to build a predictive understanding of how microbial diversity functions in nature. Hence, metagenomic approaches can be helpful to develop a designer plant technology to enhance resources with less environmental damage.

The rhizosphere, the soil region influenced by plant roots, represents a dynamic microenvironment where intricate interactions between plants and microorganisms shape soil health, nutrient cycling, and plant growth. Soil microorganisms are integral players in the transformation of materials, the dynamics of energy flows, and the intricate cycles of biogeochemistry. Considerable research has been dedicated to investigating the abundance, diversity, and intricacies of interactions among different microbes, as well as the relationships between plants and microbes present in the rhizosphere. Metagenomics, a powerful suite of techniques, has emerged as a transformative tool for dissecting the genetic repertoire of complex microbial communities inhabiting the rhizosphere. The review systematically navigates through various metagenomic approaches, ranging from shotgun metagenomics, enabling unbiased analysis of entire microbial genomes, to targeted sequencing of the 16S rRNA gene for taxonomic profiling. Each approach's strengths and limitations are critically evaluated, providing researchers with a nuanced understanding of their applicability in different research contexts. A central focus of the review lies in the practical applications of rhizosphere metagenomics in various fields including agriculture. By decoding the genomic content of rhizospheric microbes, researchers gain insights into their functional roles in nutrient acquisition, disease suppression, and overall plant health. The review also addresses the broader implications of metagenomic studies in advancing our understanding of microbial diversity and community dynamics in the rhizosphere. It serves as a comprehensive guide for researchers, agronomists, and policymakers, offering a roadmap for harnessing metagenomic approaches to unlock the full potential of the rhizosphere microbiome in promoting sustainable agriculture.

Lichens are self-supporting and ecologically obligate associations between symbiotic fungi, commonly an Ascomycete (mycobiont), and a photosynthesising organism (photobiont). Samples of Usnea longissima lichen were collected from Abies georgei in the Pudacuo National Park, Shangri-la County, Yunan Province, China, and their biological compositions were analysed by cultural and metagenomic approaches. We found that the Usnea longissima lichen symbiont contained a number of endolichenic fungi. The organisms identified included Usnea, Sydowia, Alectoria, Punctelia, Penicillium, Mucor, Hypocrea, Trichoderma, Elaphocordyceps, Arthrinium and Cladosporium. This is the first report of different genera of lichenized fungi, Usnea, Alectoria and Punctelia, co-existing in the same symbiont. This may be related to multiple origins for the fungi lichen symbiosis. Using algal-specific ITS rDNA primers, the photobiont of the Usnea longissima lichen was identified as Trebouxia. However, we also found that Poterioochromonas was present in the Usnea longissima lichen, through 18S rDNA clonal analysis of the metagenome. To date, there has been no report of two cross-phylum algae co-existing in a single lichen. This study provides new insight into the biological composition of the lichen Usnea longissima, highlighting its microbial diversity. Key words: Usnea longissima lichen, biological composition, cultural and metagenomic approach, Poterioochromonas.

Metagenomics is an approach for directly analyzing the genomes of microbial communities in the environment. The use of metagenomics to investigate novel enzymes is critical because it allows researchers to acquire data on microbial diversity, with a 99% success rate, and different kinds of genes encode an enzyme that has yet to be found. Basic metagenomic approaches have been created and are widely used in numerous studies. To promote the success of the advance research, researchers, particularly young researchers, must have a fundamental understanding of metagenomics. As a result, this review was conducted to provide a thorough insight grasp of metagenomics. It also covers the application and fundamental methods of metagenomics in the discovery of novel enzymes, focusing on recent studies. Moreover, the significance of novel biocatalysts anticipated from varied microbial metagenomes and their relevance to future research for novel industrial applications, the ramifications of Next-Generation Sequencing (NGS), sophisticated bio-informatic techniques, and the prospects of the metagenomic approaches are discussed. The current study additionally explores metagenomic research on enzyme exploration, specifically for key enzymes like lipase, protease, and cellulase of microbial origin.

Landfills are repository for complex microbial diversity responsible for bio-degradation of solid waste. To elucidate this complexity, samples from three different landfill sites of North India (sample V: Bhalswa near Karnal byepass road, New Delhi, India; sample T: Chandigarh, India and sample S3: Una, H.P., India) were analyzed using metagenomic approach. Selected landfill sites had different geographical location, varied in waste composition, size of landfill and climate zone. For comparison, one sample from high altitude (sample J) having less human interference was taken in this study. The aim of this study was to explore microbial diversity of communities responsible for degradation of landfill. Samples were characterized by 16S rRNA gene sequencing. Data from three landfill sites showed abundance of phylum Proteobacteria while less contaminated sample from high altitude showed abundance of phylum Cholroflexi followed by phylum Proteobacteria. The most abundant genus was unknown suggesting that these landfills could be repository for various novel bacterial communities. Sample T was relatively more active in terms of microbial activity. It was relatively abundant in enzymes responsible for dioxin degradation, styrene degradation, steroid degradation, streptomycin biosynthesis, carbapenem biosynthesis, monobactam biosynthesis, furfural degradation pathways while sample J was predicted to be enriched in plant cell wall degrading enzymes. Co-occurrence analysis revealed presence of complex interaction networks between microbial assemblages responsible for bio-degradation of hydrocarbons. The data provides insights about synergetic interactions and functional interplay between bacterial communities in different landfill sites which could be further exploited to develop an effective bioremediation process.

Metagenomic approaches have provided a better understanding of microbial diversity and function across the terrestrial biome. Initial studies on soil metagenomics involved construction of libraries and sequencing of cloned genes to know the product encoded, but now a days direct sequence-based information plays an important role in functional profiling of environmental DNA. The rich information obtained from soil metagenome provides new insight into the taxonomic and functional diversity of soil microorganism. Some of the techniques of molecular biology research such as clone-based gene sequence analysis, molecular fingerprinting, next-generation sequencing, and many others have proved very useful in analyzing unknown environmental DNA sample and opened a flux gate of exciting research finding. Additionally, development of new environmental DNA isolation method as well as improved cloning systems has accelerated the pace of research. More importantly, metagenomic tools have resulted in discovery of several novel genes coding for protease, lipase, amylase, alcohol oxidoreductase, antibiotic resistance, etc., from ecological niches including meadows, crop fields, and others. With metagenomic approaches, new dimension in the characterization of complex microbial community has been attained. Surely, metagenomic approaches can be used to build a predictive understanding of how microbial diversity and function vary across terrestrial biome.