Microbes for Drought Resistance in Zea mays L. (Maize) PDF

# Microbility: Exploring the Influence of Microbes in Enhancing Zea mays L. (Maize) Resistance to Drought Stress ## Senior High School Department, Leyte National High School ## Science, Technology, Engineering, and Mathematics Strand (STEM) ## Practical Research II ## Wayne David C. Padullon, MAT #...

# Microbility: Exploring the Influence of Microbes in Enhancing Zea mays L. (Maize) Resistance to Drought Stress ## Senior High School Department, Leyte National High School ## Science, Technology, Engineering, and Mathematics Strand (STEM) ## Practical Research II ## Wayne David C. Padullon, MAT ## October 16, 2024 ## Statement of the Problem The increasing occurrence of drought poses a global challenge, affecting the production and threatening food security by directly impacting both plants and the interaction of microorganisms. More systems are needed to combat the effect of drought stress on plants. This study aims to investigate the effects of diverse microbial communities on the drought tolerance of *Zea mays L.* (Maize). This study specifically aims to: 1. **To determine the classification of the selected various microbes that can enhance the drought resistance of *Zea mays L.* (Maize).** * Fungi * Algae * Combination of Fungi and Algae 2. **To investigate whether manipulating two classifications of microbes - combination of algae and fungi - have a negative impact or can significantly enhance the drought tolerance of *Zea mays L.* (Maize) through examining morphological properties and the survival capability within 40 days. In determining whether the application of microbes are effective, the following parameters will be monitored, measured, analyzed, and interpreted:** * Leaf structure * Stem structure * Root System 3. **To determine the significant difference in drought tolerance of *Zea mays L.* (Maize) when grown in three (3) different soil types.** * Loam sandy soil * Sandy clay loam soil * Sterilized soil ## Scope and Delimitations The general intent of this study is to investigate the effects of fungi and algae in mitigating plant drought stress. Specifically, it will explore how fungi, algae, and combinations of fungi and algae interact with the plant's soil in increasing nutrient utilization through examining the morphological changes of *Zea mays L.* (Maize). In terms of classifying the microbes, this study will primarily focus on the general classifications of the microbes rather on the specific types of each microbe that will be used in the study. The scope of this study will encompass fungi, algae, and combination of fungi and algae. To maintain the focus and feasibility of the study, the study will be restricted to a specific set of plant species known for their agricultural importance and sensitivity to drought, namely *Zea mays L.* (Maize) focusing on the V6 phase of the plant, the stalk, and nodal root growth. ## Research Hypothesis This study will focus on the effect of fungi, algae, and combinations of fungi and algae, on the drought resistance of *Zea mays L.* (Maize). The expected outcomes of the researchers are listed below. ## Null Hypothesis (HO) Various microbes do not significantly affect the drought resistance of *Zea mays L.* (maize). ## Alternative Hypothesis (H1) Various microbes have a significant effect on enhancing the drought resistance of *Zea mays L.* (Maize). ## Conceptual Framework This conceptual framework provides a structured overview of the key elements and relationships within the study on the effects of microbial communities on plant resistance to drought stress. ## Figure 1 [Diagram description] Relation between variables illustrated using the IV-DV model: * **Independent Variable** * Algae * Fungi * Mixture of Algae and Fungi * **Moderating Variable** * Microbes Ratio * Soil type * **Mediator Variable** * Soil enrichment * **Dependent Variable** * Resistance of Maize (*Zea mays L.*) to Drought stress * **Control Variable** * Water application * Environmental conditions ## Independent Variables This variable refers to the various types of microbes to be used in the study, including fungi, algae, and a combination of fungi and algae. The researchers aim to examine the effectiveness of different microbial communities in plant growth, impact of soil health, and response to environmental change to enhance plant resistance to drought stress. ## Dependent Variable The dependent variable is plant resistance to drought stress, which is adaptive characteristics that allow it to withstand drought stress. The specific plant that the researchers will use is *Zea mays L.* (Maize). Researchers are investigating the impact of microbial communities on plant resistance to drought stress, selecting a plant species with known drought sensitivity. Additionally, exploring the long-term effects of microbial communities on drought resistance over multiple growing seasons could provide deeper insights into their impact. ## Moderating Variables The moderating variables in this study are the ratio of microbial communities to be used and soil type. These factors might influence the interaction between the microbes and the plant. The ratio of the soil community to be used can lead to different levels of effectiveness in enhancing the plant's drought resistance. This is the reason why the ratio of each microbe in each treatment must be equal to ensure the fairness of the results. The soil type must also be the same, as different soil types can affect microbial activity, which could significantly affect the factors that the researchers will be basing their decision to determine whether the experiment succeeded. ## Control Variables The control variables in this study are water application, environmental conditions, and soil type. For this study, the researchers must control the amount of water provided to the plants. This is important for establishing drought stress. In order for this study to succeed, environmental conditions such as temperature, light, and water availability must be maintained throughout the experiment. Controlling these factors ensures that any changes in *Zea mays L.* (Maize) resistance to drought is due to the interaction of microbes and not from environmental factors. By isolating the impact of microbes, the study provides a clearer understanding of their role in reducing drought stress on maize. ## Mediator Variable The mediating variable in this study is soil enrichment. Microbes enrich the soil by improving its quality, nutrient content, and soil health, which could enhance the maize (*Zea Mays L.*) resistance to drought. ## Significance of the Study This study intends to investigate how **microbes**, specifically algae and fungi, significantly influence plant resistance to drought stress, with the possible improvement of land management and agricultural sustainability, and contribute valuable knowledge to the scientific community. ## Farmers The study will provide farmers with the information on how they can adopt practices that enhance the health of their soil. This, in turn, results in more resilient plants that deliver high yields, even when facing adverse weather conditions. ## Consumers The research will provide ways in terms of advancements in understanding microbial communities that can see tangible benefits in their daily lives from the application of microbial research to improve drought resilience in crops. ## Enhanced Crop Resilience The findings of the study can help provide new alternatives to make plants drought-friendly using microbes that could lead to lower crop losses during times of drought. # Chapter II: Review of Related Literature This chapter of the paper presents a diverse review of related studies from various sources such as books, articles, and research studies that were conducted previously by the researchers, experts, and specialists relevant to understanding the study of microbes on plant resistance to drought stress. ## Benefits of Microbes This section will provide a comprehensive review of related literature regarding the benefits of microbes. Key findings on the benefits of microbes by related studies from reliable sources will be presented and discussed in this part. By doing so, researchers can acquire important knowledge that can potentially be of use in this research study, and likewise come up with ideas on how to maximize these benefits. Without further ado, the next paragraphs will present an overview of the main microbes involved in this study and present and discuss the benefits of microbes. This study will focus on two classifications of microbes: **Algae and Fungi**. Algae are members of mostly aquatic photosynthetic organisms under the Protista kingdom. Algae have many types and range in size from microscopic to gigantic. These microbes are eukaryotic organisms that photosynthesize but lack the specialized multicellular structure of plants. On the other hand, fungi are also eukaryotic microorganisms under the Fungi kingdom. These microorganisms are ubiquitous and can occur as yeasts, molds, and mushrooms. Moreover, fungi are clearly distinguished from all other organisms by nutrient intake, vegetative growth, and physical characteristics. Additionally, fungi are heterotrophic and digest food externally by releasing hydrolytic enzymes before absorbing it into the mycelia. These microbes exist for a reason just like humans do. These microbes have a significant impact on ecosystems. According to Roles of Microbes in Ecosystem (2017) microbes significantly contributed to the production of oxygen in Earth's history; nutrient cycling-which would stop the primary productivity without this recycling of nutrients; nitrogen fixation of microbes from the atmosphere into a useable form; microbes allow herbivores to consume poor quality food; finally, give plant roots access to nutrients in the soil. Conforming to this, Singavarapu et al. (2023) and Hermans et al. (2023) claimed these microbes are fundamental components of the ecosystem responsible for maintaining soil quality and plant regulation. With this information, this study aims to maximize the benefits of microbes-specifically Fungi and Algae and the combination of both--to enhance the drought-stress resistance of *Zea mays L.* (Maize), and contribute to sustainable agriculture. The next part will provide a comprehensive review of related literature on the benefits of two main microbes involved in this study by breaking down the information from existing related studies. ## Benefits of Algae Algae are found almost everywhere, land or water. These organisms are beneficial. Many studies exist proving algae is beneficial, especially in the field of agriculture. According to the study of Mahapatra et al. (2018), algae can be beneficial to agriculture as these organisms can be used as biofertilizers and can improve soil health. The study shows algae can be a substitute for chemical fertilizers due to characteristics such as ubiquity, short time of reproduction, enhanced metabolic flux, and inherent capabilities of nitrogen fixation. The study revealed that algae are beneficial in agriculture as these organisms are rich in nutrients-especially cyanobacteria which are rich in nitrogen, phosphorus, and potassium which are essential to plants. As a biofertilizer, a study by Chatterjee et al. (2017) has proven algae to enhance rice productivity. The presence of algae, particularly the blue-green algae (BGA), in soil results in the formation of soil aggregates, and regulates soil aeration, and soil temperature, thus, improving the physical and chemical properties of the soil. Chatterjee et # Chapter III: Methodology This chapter offers information on the research methodology that was implemented, this includes an overview of the research design, research locale, the instruments that the researchers utilized, the procedures that was implemented, how the analysis was accomplished, and the ethical consideration throughout the study. ## Research Locale The study will be conducted at Barangay Rosal Sta. Rita, Samar. It will take place during the period from January 2024 to March 2024. Due to its spacious grounds, the place will allow the researchers to build a greenhouse wherein the experiment will take place. ## Research Sample(s) * **Zea mays L. (Maize)** * *Zea mays L.* (Maize) was selected as a model organism due to its importance as a staple crop and its sensitivity to drought stress. The V6 growth stage, a critical period for water availability and plant development, makes maize particularly suitable for evaluating strategies to enhance drought resistance. * **Algae** * Algae were chosen as one of the microbes to enhance maize drought tolerance due to their ability to improve soil moisture retention and supply essential nutrients, potentially promoting plant growth and survival under water-limited conditions. * **Fungi** * Fungi will be used because of its ability to improve water and nutrient uptake in plants, which can increase drought tolerance by enhancing the plant's resilience to water stress. * **Mixture of Algae and Fungi** * Researchers will be investigating the potential synergistic effects of an algae-fungal mixture on plant drought resistance. It is hypothesized that the combined application of both organisms will provide a faster improvement in drought tolerance compared to using either microbe alone, through complementary mechanisms that enhance water retention and nutrient availability. ## Soil Samples The researchers will be using three (3) types of soil: a) loam sandy soil, b) sandy clay loam soil, and c) sterilized soil. These soil will be used to establish the experimental plots. ## Research Design This section outlines the research design that will be employed in this study. A quantitative approach, specifically true experimental research, will be used to investigate the role of algae and fungi in enhancing drought stress resistance in Maize (*Zea Mays L.*). The Randomized Complete Block Design (RCBD) will be adopted for this study. According to Grant (2010) RCBD is the standard design for agricultural experiments where similar experimental units are grouped into blocks or replicates. This is used to assess variation in an experiment by accounting for spatial effects in the field or greenhouse. Treatments are randomly assigned to the subjects in different blocks, wherein treatments are grouped in different blocks with different intervening variables that may influence the relationship between the independent variables and dependent variables. This will be done to assess any variation and to ensure that the observed results are solely attributed to the interaction between the microbes and the plant. This study will employ two classifications of microbes: algae and fungi. These will be arranged following the Randomized Complete Block Design (RCBD). The *Zea Mays L.* (Maize) will undergo six (6) different treatments: Algae treatments, Fungi treatments, Algae-dominant treatments, Fungi-dominant treatments, treatment of equal concentration of algae and fungi, and a controlled treatment wherein no microbes will be applied to assess the effects of the microbes. These treatments-except for controlled treatments--will have four (4) with nine (9) replicates each in varying amounts of microbes: 20 milliliters, 40 milliliters, and 60 milliliters of microbial application; the controlled treatments will have three (3) replications, this will be a total of 48 samples. Following the RCBD design, the forty-eight (48) samples will be divided into three (3) blocks. Each three (3) block differs in the type of soil to be used-sterilized soil, clay soil, and sandy loam soil. This will be done to monitor and analyze the variations in the results from different treatments in different soil types, and ensure that the results are mainly due to the interaction of the microbes and the plant. ## Figure 2 [Diagram description] Map of randomized designation of treatments in different blocks. ## Research Set-up In conducting this experiment, the researchers will build a greenhouse as it provides an ideal setting to recreate and control the variables that may affect the relationship between microbes and *Zea mays L.* (Maize.) The greenhouse will serve as the facility where the experiment will take place. The measurements of this greenhouse will be based on the size and the number of pots that will be used. Furthermore, the greenhouse will be constructed primarily from sustainable bamboo and covered with polycarbonate to ensure that the samples receive sunlight without being exposed to external precipitation, such as rain. This design allows exposure to light while protecting the experimental setup from the influence of unpredictable weather conditions. To know the area for the greenhouse that will be built, the diameter of a 10L plant pot will be used as the basis which is 28cm. The experiment will use six (6) pots in a block with controlled treatment and the distance from one treatment to another is 7.62cm Hence, to compute for the length needed for the pots to be plotted is: (28cm)(6 pots) + (7.62cm)(5pots) = 168cm + 38.1cm = 206.1cm or 2.06m On the other hand, the pots will be plotted in 9 different blocks with a distance of 1m from the other. To compute for the width needed for the pots to be plotted is: (9 blocks)(1 meter) = 9m With these measurements, the area of the land where the pots will be plotted will be computed using the formula for finding the area of a rectangle which is A=LxW. Hence, the area is equal to (2.06m)(9m) which will result to 18.54m². So, for the researchers to be able to mobilize freely in the facility, 0.5m is added to the length and width of the land needed for the pots to be plotted. Therefore, the area of the greenhouse will be calculated with this equation: A=(L+0.5m)(W+0.5m) ## Sample Preparation In the implementation of this study, various materials will be cultivated by the researchers. Additionally, certain materials will be procured from local sources to meet the specific requirements of this study. * **Collection of *Zea mays L.* (Maize).** *Zea mays L.* (Maize) seeds will be sourced from local agricultural stores. Forty-eight (48) seeds will be individually grown in loam soil and will be watered normally three (3) weeks until it reaches the V6 growth phase. * **Collection of Algae.** The researchers will be combining three (3) types of algae:water silk, cyanobacteria, and seagrapes. These algae will be obtained from commercial suppliers and / or natural resources. * **Collection of Fungi.** The researchers will be combining three (3) types of fungi: yeast, mushroom, and molds. These fungi will be obtained from commercial suppliers and/or natural resources. * **Collection of Mixture of Algae and Fungi.** For the treatment group receiving both algae and fungi, a mixture will be prepared by combining equal concentrations of algae and fungi. * **Collection of Soil.** The soil that will be used in this study are: a) loam sandy soil, b) sandy clay loam soil, and c) sterilized soil. Loam sandy soil and sandy clay loam soil will be collected in local agricultural stores. Sterilized soil will be obtained through undergoing loam soil in heat. ## Extraction In conducting this experiment, the study Microbial inoculants: A strategy to improve plant growth and productivity of Harish & Rai (2016) will be used as a guide for the extraction of algae and fungi. The algae and fungi will be extracted using a solvent-based method. To extract the fungi and algae , the selected three (3) types of fungi will be individually ground into a fine powder using a mortar and pestle The powder will then be soaked in water at 27-28°C for 10 minutes. Similarly, the three (3) selected types of algae will be soaked in water at 27-28°C for 10 minutes The extracted algae and fungi will then be prepared as a concentrated solution for soil inoculation This solution will be applied to the soil at specific concentrations to evaluate its impact on plant growth and drought resistance ## Treatment Preparations In this study, to investigate the effects of microbial inoculation on soil to enhance *Zea mays L.* (Maize) drought resistance, the researchers will ensure that *Zea mays L.* (Maize) plants had reached the V6 growth stage before initiating the experiment. Forty-eight (48) pots will be prepared, each containing a different microbial inoculant at three (3) varying concentrations: 20 milliliters, 40 milliliters and 60 milliliters, and one (1) pot for each soil type for the negative control. After inoculation with the respective microbial treatments, all experimental groups including the negative control will be subjected to a drought stress phase. Drought conditions will be simulated by withholding water for a predetermined period of eight (8) days. The researchers will be observing the effect of different microbes on the plant morphological aspects. The effects of each group will be compared to evaluate how the different microbial treatments (algae, fungi, and their mixture) affect maize drought resistance. * **Negative control.** The negative control group will consist of *Zea mays* (Maize) plants that will not receive any microbial treatment (no algae, no fungi, no mixture). These plants will be subjected to drought stress. This group is necessary to establish a baseline drought response in untreated maize. By comparing the performance of the negative control plants with those receiving microbial treatments, the researchers can assess the effectiveness of the treatments. ## Data Collection In collection of the data, to observe the effects of different microbes, researchers will use morphological analysis to assess the changes of the plant. According to the study by Smith & Brown (2020) that morphological analysis refers to examining the physical form of the plant through their leaf structure, root development, stem growth and thickness, and fruiting patterns. The plant will be monitored every three (3) days to examine the leaf changes, stem growth. ## Statistical Analysis Data analysis will be conducted using Microsoft Excel and the Statistical Package for the Social Sciences (SPSS). This study will focus on comparing different samples subjected to various microbial treatments to determine if specific microbial classifications or concentrations significantly improve drought-stress tolerance in *Zea mays L.* (Maize). A Two-way ANOVA will be employed to analyze the effects of algae or fungi treatments on drought resistance, examining if variations in microbial treatment yield significant differences in plant drought tolerance. To identify specific differences between sample concentrations, a post hoc test, specifically Tukey's Honest Significant Difference (HSD) test, will be applied following a significant ANOVA result. ANOVA is a statistically significant test often used with multiple comparison procedures to detect group differences (Sawyer, 2009). If the F-test from the Two-way ANOVA reveals significant differences among the groups, Tukey's HSD test will identify the specific group differences. The study aims to determine whether fungi or algae treatments alone significantly impact drought resistance in *Zea mays L.* (Maize) and to compare the effects of different concentration levels to assess their efficacy in