Table of Contents
Introduction
Fungi are appearing in almost all habitats including aquatic and soil, where they are known to maintain their symbiotic or parasitic correlation with different organisms (Cuadros-Orellana et al. 2013). They embody a huge fraction of the earth’s genetic diversity, and their activity is often vital as it influences the rate of a number of processes in the ecosystem; thus the structure of animals and plants. Fungi are primary players in the nutrient cycle process, including decomposition of organic matter; and are also critical sources of the active biological substances. Studies have shown that fungal biodiversity is fundamentally dissimilar in plants, animals, and even bacteria. The global number of fungal species has been approximated to be a few millions. Therefore, this research paper describes the fungal patterns of biodiversity by considering the drivers of this diversity in various environments. The research aims at proving the hypothesis that fungal diversity is greater in dry soil and plant habitats compared to aquatic or wet and animal habitats. The experiment entails the determination of fungal diversity in dry and moist habitats.
Background Information and Literature Review
Fungal diversity researches have conventionally relied on the phenotypic and morphologic characteristics, which were also the previous classification criteria for fungi. However, these techniques have changed over time due to their instability and overlapping nature; thus giving ways to other novel methods of fungi identification. Many terrestrial environments are characterized by fungal hyphae often twisting around animals, plants other microorganisms like bacteria and even soil. Fungi are not obvious to the nude eye, but they are immensely enmeshed in the evolutionary ecology of life just like any other organism. Fungi kingdom falls under the category of Eukaryotes, and consists of more than 1.5 million species (Peay, Kennedy and Talbot 2016). The category consists of diverse organisms that may be unicellular or multicellular. The fungi are often essential both ecologically and economically. For instance, the root mycorrhiza through mutualism has facilitated the land colonization by various plants, thus leading to fungal peroxidase evolution which is essential in lignin degradation. Furthermore, fungi can be observed to be influential in plant genomes human immune system functioning as well as soil chemistry. In particular, fungal pathogens have been significant in maintaining the great taxonomic diversity among the tropical forests. In spite of this immense influence, fungal diversity is not fully appreciated.
Fungi are of the monophyletic category which diverged from the animal kingdom several years ago. The organisms in the kingdom fungi, as well as their habitats, form the mycobiome. Also, fungal hallmark can be described by their unique hyphal growth, which enhances their ability to create a network of mycelium or filaments that interconnect to form somatic tissues. The single-celled fungi are mainly predominant in stressful habitats or liquid environments; for instance the deep marine sediments, anaerobic conditions like in gut rumen, or in the floral nectar. These habitats mainly prevent the growth of the fungal filaments. On the other hand, the hyphal fungi consist of the most diverse and abundant organisms in the natural environments. The hyphal element is a clear indication that the fungi have the ability to perceive as well as respond to the surrounding environment. Further, these fungi have the ability to share resources and also coordinate functions throughout the heterogeneous habitats that are reachable to the mycelia. Because of these mycelia, often the mycobiome may incorporate larges or older organisms in nature.
In addition, fungi are considered to be osmotrophs, meaning that they have the ability to decompose the organic matter efficiently. This is possible because they can produce diverse organic acids as well as enzymes that often dissolve and recycle the recalcitrant natural or synthetic substances. Therefore, the ecological and evolutionary success of fungi is often related to an uncommon combination of biochemical and morphological aspects that differentiate them from the other organisms.
The composition and diversity of mycobiome change significantly across different environments. Fungal diversity studies have been facilitated by various developments, thus enabling novel ways of identifying fungi from different habitats and samples. Most studies have provided immense insight into the population of fungi present in soils, plants, animals and even aquatic ecosystems. According to Peay, Kennedy, and Talbot, fungal diversity is immense in the soil as well as plants, but less in aquatic environments; thus fungi in soils and plants are associated with great functional contribution to the ecosystem. Generally, fungal diversity has been known to be high in the soils, and studies have shown that over 80,000 fungal species have been found in soil habitat ((Peay, Kennedy and Talbot 2016). This study simply involved a few grams of the soils, which detected thousands of the fungal species. High diversity of fungi in the soil can be attributed to the fungal taxa radiation into the open ecological niche especially during the development of the terrestrial environment. Almost all the vital branches of fungi can be isolated from soil, but the most abundant branch of fungi are the mycorrhizal fungi and the Basidiomycota.
The tissues of living plants are also rich in fungi, especially on the roots, leaves, and stems; which inhabit a number of the endophytic fungi, fungal pathogens, and mycorrhizal fungi. Peay, Kennedy, and Talbot (2016) assert that over 2,500 endophytic fungi taxa were isolated from leaves of one tree. The roots of one tree have the capability of harboring over 100species of ectomycorrhizal fungi as well as numerous other endophytic species. The fungi mostly found on leaves are majorly of the phylum Ascomycota.
Studies have also illustrated that fungal diversity in marine and animal habitats is quite low compared to soil and plants (Wagg et al. 2014). Nonetheless, this conclusion may also be underestimated because of the inadequacy in large-scale researches in these habitats. The marine and animal environments are home to very significant and novel lineages of fungi; thus suggesting that fungal diversity in such environments may be higher than initially reported (Zeglin 2015). It is also important to note that the fungal diversity in relation to the marine environment is often less explored; this is because the organisms are less abundant compared to the bacteria in the habitats. For example, investigations have revealed that a new proposed fungal phylum- the Cryptomycota was found in freshwater habitats. The fungi are single-celled and have the ability to form flagella, and are also parasites of algae and other fungi. Fascinatingly, the distinct marine sediments and human skin environments can harbor just a few numbers of yeast species particularly those from the Basidiomycota and Ascomycota phyla. In particular, from Ascomycota, some Candida species are known to cause thrush, while from Basidiomycota, the Malassezia species cause dandruff. On the other hand, other species from these two genera are known to be found in the fungal communities in marine sediments and corals.
The major difference attained in the fungal communities’ composition in various environments is therefore associated with the benefits of the hyphal growth, which is essential for osmotrophy especially in solid substrates like solid as well as woody plant tissues. The morphological innovation, as well as the ability to develop symbioses with the living plants, is an ecological success particularly of the hyphal fungi in most of the novel niches that were developed during terrestrial ecosystem generation.
Problem Statement
The diversity of fungi keeps changing profoundly across different environments. The fact remains that fungi exist in almost all environments, the question is, which environment harbors the most number of fungi? Several studies have been performed and different conclusions have been made, with some studies concluding that the soil has the greatest fungi diversity, while others vouching for the marine environments and others plants. Therefore, this research purposes to undertake an experiment to determine the diversity of fungi in dry and wet habitats as detailed in the methodology section.
Hypothesis
The research aims at proving the below null hypothesis.
H0: Fungal diversity is greater in dry soil and plant habitats compared to aquatic or wet and animal habitats.
The variables to be tested in the experiment include moisture content in the soil samples and fungi diversity (Shannon index, richness, evenness, as well as the Chaol indices richness). indicators
Methods
The materials required for the experiment include pH paper, soil samples from different sites, kit for DNA isolation, microscope, burner, and safety glasses. The study samples from varied locations that are pre-identified and collected across the range of soil samples to represent the different environmental conditions such as dry sand, dry wood, and dry moist sand. The samples are well labeled and prepared for the fungi diversity tests.
The soil samples from different environmental conditions to be used for the experiment are prepared to ensure that they are suitable to give the required results. Different procedures are implemented in sample preparation. The initial process of sample preparation involves serving the soil samples using 2- mm wire mesh to remove the larger grains and to create uniformity in the samples. Same grain samples tend to have distinct properties that will be suitable in identification if fungi diversity. The samples are them kept in clean, well-labeled containers and stored in a cool place, about 40C before subjection to further experiments and chemical analysis.
After cooling the samples for a sufficient period of time, the samples are put through soil temperature and physiochemical classification. Each soil sample is classified with regards to the pH, humus content, and organic carbon, availability of potassium, phosphorus, and magnesium. The electrical conductivity of each soil sample is also identified to be used in the determination of electrolyte concentration. The ratio of total organic carbon to total nitrogen is also determined for each soil sample. It is important that all these properties are determined for dry soil samples at about 1050C.
The next sample preparation process is DNA extraction, where every soil sample is subjected to the 250 mg soil mass DNA Isolation Kit. The researcher has to follow the manufacturer’s instructions manual that comes with the DNA isolation kit that assists in the standardization of soil DNA concentration to about 20 ng µL-1.
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The procedure is used to identify the fungi present in each soil sample. The stain method classifies fungi and bacteria into different clusters depending on the reactions of the different fungi to the stain. The reaction of the fungi and bacteria may test gram-negative, gram-variable, or gram-positive. Despite the limited explanations on the mechanisms used in the gram stain, it is understandable that the bacteria and fungi cells to stain wall vary because of the chemistry and complexity differences of the fungi cells such as the peptidoglycan and polymer. For instance, the gram-negative fungi contain less polymer or peptidoglycan while the gram-positive have more. The procedure for fungi classification is as follows:
The water droplet is placed in the middle of a sterilized microscope slide.
Bacteria of Fungi colony is selected by flaming and cooling the agar in the loop.
The fungi are evenly spread in the slide with water droplet and left to dry.
The dry fungi smear is gently heated by passing the slide over low heat flames a couple of times than allowed to cool.
Crystal violet mixture is added to the smear in few drops. The slide is then rinsed using deionized water after some time. The excess water is removed and the slide covered with iodine to set the stain.
The slide is then rinsed using decolorizer to classify the fungi.
The process is repeated for each soil sample.
After classification of the fungi in each soil sample, the samples are subjected to further ecological succession procedure to determine the fungi diversity. The process for fungi diversity calculation is as follows:
The fungi colonies for each soil sample in the agar plate are observed and described.
The records from the Gram stains for all soil samples in the earlier procedure are used to calculate the fungi diversity at this stage using the Shannon-Weiner Index.
H’= -∑i=1Spi ln pi =
Where S- the number of types
The pHs for each soil sample is also recorded.
The fungal diversity in each soil sample is determined by calculating the Shannon index, richness, evenness, as well as the Chaol indices richness indicators. The structural differences of the fungal community in every soil sample are assessed through the T-test and non-metric multidimensional scaling. The statistical differences in the samples chemical properties, temperature, richness, and copy number of fungi are analyzed using T-test and ANOVA multiple correlation analysis.
- Cuadros-Orellana, Sara, et al. “Assessment of fungal diversity in the environment using metagenomics: a decade in review.” Fungal Genomics & Biology 3.2 (2013): 1-13.
- Peay, Kabir G., Peter G. Kennedy, and Jennifer M. Talbot. “Dimensions of biodiversity in the Earth mycobiome.” Nature Reviews Microbiology 14.7 (2016): 434-447.
- Wagg, Cameron, et al. “Soil biodiversity and soil community composition determine ecosystem multifunctionality.” Proceedings of the National Academy of Sciences 111.14 (2014): 5266-5270.
- Zeglin, Lydia H. “Stream microbial diversity in response to environmental changes: review and synthesis of existing research.” Frontiers in Microbiology 6 (2015): 454.