Maize Seedlings Colonization with Serendipita indica and Its Colonization Efficiency Analysis.

Maize is one of the most important crops in the world, and ensuring its successful growth and productivity is crucial for global food security. One way to enhance maize growth and productivity is by improving the colonization of its roots by beneficial microorganisms. In this regard, Serendipita indica, a plant growth-promoting fungus, has gained attention for its ability to enhance plant growth and productivity, especially in cereal crops and medicinal plants. Previous studies have shown that S. indica can colonize various plant species, including maize, but the efficiency of the colonization process in maize seedlings has not been extensively characterized. This protocol outlines a method for efficient colonization of maize seedlings with the beneficial fungus S. indica. The protocol includes the preparation of stock solutions, maintenance and growth of S. indica, surface sterilization and germination of seeds, preparation of S. indica chlamydospores, and colonization of maize plants with S. indica. The advantages of this protocol include the use of surface sterilization techniques that minimize contamination, the production of a large number of viable chlamydospores, and efficient colonization of maize seedlings with S. indica. This protocol may be useful for researchers studying the role of S. indica in promoting plant growth and combating biotic and abiotic stress. Additionally, this protocol may be used in the development of biofertilizers using S. indica as a means of increasing crop yields and reducing dependence on synthetic fertilizers. Overall, this protocol offers a reliable and efficient method for colonizing maize seedlings with S. indica and may have potential applications in the agricultural industry. This study also provides a valuable tool for researchers interested in studying plant-microbe interactions in maize and highlights the potential of S. indica as a biocontrol agent to enhance maize productivity under adverse conditions. Key features • This protocol builds upon the method developed by Narayan et al. (2022), and its application optimized for the root endophytic symbiotic fungus S. indica. • This protocol also allows for histochemical analysis to visualize the colonized fungal spores in the root cells of host plant species. • This protocol helps in mathematical calculation of the percent colonization or efficiency of colonization. • This protocol utilizes readily available laboratory equipment, including a light microscope, autoclave, and laminar flow hood, ensuring ease of reproducibility in other research laboratories.

Integrated omics approaches for flax improvement under abiotic and biotic stress: Current status and future prospects.

Flax (Linum usitatissimum L.) or linseed is one of the important industrial crops grown all over the world for seed oil and fiber. Besides oil and fiber, flax offers a wide range of nutritional and therapeutic applications as a feed and food source owing to high amount of α-linolenic acid (omega-3 fatty acid), lignans, protein, minerals, and vitamins. Periodic losses caused by unpredictable environmental stresses such as drought, heat, salinity-alkalinity, and diseases pose a threat to meet the rising market demand. Furthermore, these abiotic and biotic stressors have a negative impact on biological diversity and quality of oil/fiber. Therefore, understanding the interaction of genetic and environmental factors in stress tolerance mechanism and identification of underlying genes for economically important traits is critical for flax improvement and sustainability. In recent technological era, numerous omics techniques such as genomics, transcriptomics, metabolomics, proteomics, phenomics, and ionomics have evolved. The advancements in sequencing technologies accelerated development of genomic resources which facilitated finer genetic mapping, quantitative trait loci (QTL) mapping, genome-wide association studies (GWAS), and genomic selection in major cereal and oilseed crops including flax. Extensive studies in the area of genomics and transcriptomics have been conducted post flax genome sequencing. Interestingly, research has been focused more for abiotic stresses tolerance compared to disease resistance in flax through transcriptomics, while the other areas of omics such as metabolomics, proteomics, ionomics, and phenomics are in the initial stages in flax and several key questions remain unanswered. Little has been explored in the integration of omic-scale data to explain complex genetic, physiological and biochemical basis of stress tolerance in flax. In this review, the current status of various omics approaches for elucidation of molecular pathways underlying abiotic and biotic stress tolerance in flax have been presented and the importance of integrated omics technologies in future research and breeding have been emphasized to ensure sustainable yield in challenging environments.

Sulfur nutrition and its role in plant growth and development.

Sulfur is one of the essential nutrients that is required for the adequate growth and development of plants. Sulfur is a structural component of protein disulfide bonds, amino acids, vitamins, and cofactors. Most of the sulfur in soil is present in organic matter and hence not accessible to the plants. Anionic form of sulfur (SO42-) is the primary source of sulfur for plants that are generally present in minimal amounts in the soil. It is water-soluble, so readily leaches out of the soil. Sulfur and sulfur-containing compounds act as signaling molecules in stress management as well as normal metabolic processes. They also take part in crosstalk of complex signaling network as a mediator molecule. Plants uptake sulfate directly from the soil by using their dedicated sulfate transporters. In addition, plants also use the sulfur transporter of a symbiotically associated organism like bacteria and fungi to uptake sulfur from the soil especially under sulfur depleted conditions. So, sulfur is a very important component of plant metabolism and its analysis with different dimensions is highly required to improve the overall well-being of plants, and dependent animals as well as human beings. The deficiency of sulfur leads to stunted growth of plants and ultimately loss of yield. In this review, we have focused on sulfur nutrition, uptake, transport, and inter-organismic transfer to host plants. Given the strong potential for agricultural use of sulfur sources and their applications, we cover what is known about sulfur impact on the plant health. We identify opportunities to expand our understanding of how the application of soil microbes like AMF or other root endophytic fungi affects plant sulfur uptake and in turn plant growth and development.

Functional characterization of a high-affinity iron transporter (PiFTR) from the endophytic fungus Piriformospora indica and its role in plant growth and development.

Iron (Fe) is a micronutrient required for plant growth and development; however, most Fe forms in soil are not readily available to plants, resulting in low Fe contents in plants and, thereby, causing Fe deficiency in humans. Biofortification through plant-fungal co-cultivation might be a sustainable approach to increase crop Fe contents. Therefore, we aimed to examine the role of a Piriformospora indica Fe transporter on rice Fe uptake under low Fe conditions. A high-affinity Fe transporter (PiFTR) from P. indica was identified and functionally characterized. PiFTR fulfilled all criteria expected of a functional Fe transporter under Fe-limited conditions. Additionally, PiFTR expression was induced when P. indica was grown under low Fe conditions, and PiFTR complemented a yeast mutant lacking Fe transport. A knockdown (KD) P. indica strain was created via RNA interference to understand the physiological role of PiFTR. We observed that the KD-PiFTR-P. indica strain transported a significantly lower amount of Fe to colonized rice (Oryza sativa) than the wild type (WT) P. indica. WT P. indica-colonized rice plants were healthier and performed significantly better than KD-PiFTR-P. indica-colonized rice plants. Our study offers potential avenues for an agronomically sound amelioration of plant growth in low Fe environments.

ColGen: An end-to-end deep learning model to predict thermal stability of de novo collagen sequences.

Collagen is the most abundant structural protein in humans, with dozens of sequence variants accounting for over 30% of the protein in an animal body. The fibrillar and hierarchical arrangements of collagen are critical in providing mechanical properties with high strength and toughness. Due to this ubiquitous role in human tissues, collagen-based biomaterials are commonly used for tissue repairs and regeneration, requiring chemical and thermal stability over a range of temperatures during materials preparation ex vivo and subsequent utility in vivo. Collagen unfolds from a triple helix to a random coil structure during a temperature interval in which the midpoint or Tm is used as a measure to evaluate the thermal stability of the molecules. However, finding a robust framework to facilitate the design of a specific collagen sequence to yield a specific Tm remains a challenge, including using conventional molecular dynamics modeling. Here we propose a de novo framework to provide a model that outputs the Tm values of input collagen sequences by incorporating deep learning trained on a large data set of collagen sequences and corresponding Tm values. By using this framework, we are able to quickly evaluate how mutations and order in the primary sequence affect the stability of collagen triple helices. Specifically, we confirm that mutations to glycines, mutations in the middle of a sequence, and short sequence lengths cause the greatest drop in Tm values.

Molybdenum and hydrogen sulfide synergistically mitigate arsenic toxicity by modulating defense system, nitrogen and cysteine assimilation in faba bean (Vicia faba L.) seedlings.

Hydrogen sulfide (H2S) has emerged as a potential gasotransmitter in plants with a beneficial role in stress amelioration. Despite the various known functions of H2S in plants, not much information is available to explain the associative role of molybdenum (Mo) and hydrogen sulfide (H2S) signaling in plants under arsenic toxicity. In view to address such lacunae in our understanding of the integrative roles of these biomolecules, the present work attempts to decipher the roles of Mo and H2S in mitigation of arsenate (AsV) toxicity in faba bean (Vicia faba L.) seedlings. AsV-stressed seedlings supplemented with exogenous Mo and/or NaHS treatments (H2S donor) showed resilience to AsV toxicity manifested by reduction of apoptosis, reactive oxygen species (ROS) content, down-regulation of NADPH oxidase and GOase activity followed by upregulation of antioxidative enzymes in leaves. Fluorescent localization of ROS in roots reveals changes in its intensity and spatial distribution in response to MO and NaHS supplementation during AsV stress. Under AsV toxicity conditions, seedlings subjected to Mo + NaHS showed an increased rate of nitrogen metabolism evident by elevation in nitrate reductase, nitrite reductase and glutamine synthetase activity. Furthermore, the application of Mo and NaHS in combination positively upregulates cysteine and hydrogen sulfide biosynthesis in the absence and presence of AsV stress. Mo plus NaHS-supplemented seedlings exposed to AsV toxicity showed a substantial reduction in oxidative stress manifested by reduced ELKG, lowered MDA content and higher accumulation of proline in leaves. Taken together, the present findings provide substantial evidence on the synergetic role of Mo and H2S in mitigating AsV stress in faba bean seedlings. Thus, the application of Mo and NaHS reveals their agronomic importance to encounter heavy metal stress for management of various food crops.

Sulfur transfer from the endophytic fungus Serendipita indica improves maize growth and requires the sulfate transporter SiSulT.

A deficiency of the essential macronutrient sulfur leads to stunted plant growth and yield loss; however, an association with a symbiotic fungus can greatly improve nutrient uptake by the host plant. Here, we identified and functionally characterized a high-affinity sulfate transporter from the endophytic fungus Serendipita indica. SiSulT fulfills all the criteria expected of a functional sulfate transporter responding to sulfur limitation: SiSulT expression was induced when S. indica was grown under low-sulfate conditions, and heterologous expression of SiSulT complemented a yeast mutant lacking sulfate transport. We generated a knockdown strain of SiSulT by RNA interference to investigate the consequences of the partial loss of this transporter for the fungus and the host plant (maize, Zea mays) during colonization. Wild-type (WT) S. indica, but not the knockdown strain (kd-SiSulT), largely compensated for low-sulfate availability and supported plant growth. Colonization by WT S. indica also allowed maize roots to allocate precious resources away from sulfate assimilation under low-sulfur conditions, as evidenced by the reduction in expression of most sulfate assimilation genes. Our study illustrates the utility of the endophyte S. indica in sulfur nutrition research and offers potential avenues for agronomically sound amelioration of plant growth in low-sulfate environments.

Metal transporters in organelles and their roles in heavy metal transportation and sequestration mechanisms in plants.

Heavy metal toxicity is one of the major concerns for agriculture and health. Accumulation of toxic heavy metals at high concentrations in edible parts of crop plants is the primary cause of disease in humans and cattle. A dramatic increase in industrialization, urbanization, and other high anthropogenic activities has led to the accumulation of heavy metals in agricultural soil, which has consequently disrupted soil conditions and affected crop yield. By now, plants have developed several mechanisms to cope with heavy metal stress. However, not all plants are equally effective in dealing with the toxicity of high heavy metal concentrations. Plants have modified their anatomy, morphophysiology, and molecular networks to survive under changing environmental conditions. Heavy metal sequestration is one of the essential processes evolved by some plants to deal with heavy metals' toxic concentration. Some plants even have the ability to accumulate metals in high quantities in the shoots/organelles without toxic effects. For intercellular and interorganeller metal transport, plants harbor spatially distributed various transporters which mainly help in uptake, translocation, and redistribution of metals. This review discusses different heavy metal transporters in different organelles and their roles in metal sequestration and redistribution to help plants cope with heavy metal stress. A good understanding of the processes at stake helps in developing more tolerant crops without affecting their productivity.

Plant mineral transport systems and the potential for crop improvement.

MAIN CONCLUSION: Nutrient transporter genes could be a potential candidate for improving crop plants, with enhanced nutrient uptake leading to increased crop yield by providing tolerance against different biotic and abiotic stresses. The world's food supply is nearing a crisis in meeting the demands of an ever-growing global population, and an increase in both yield and nutrient value of major crops is vitally necessary to meet the increased population demand. Nutrients play an important role in plant metabolism as well as growth and development, and nutrient deficiency results in retarded plant growth and leads to reduced crop yield. A variety of cellular processes govern crop plant nutrient absorption from the soil. Among these, nutrient membrane transporters play an important role in the acquisition of nutrients from soil and transport of these nutrients to their target sites. In addition, as excess nutrient delivery has toxic effects on plant growth, these membrane transporters also play a significant role in the removal of excess nutrients in the crop plant. The key function provided by membrane transporters is the ability to supply the crop plant with an adequate level of tolerance against environmental stresses, such as soil acidity, alkalinity, salinity, drought, and pathogen attack. Membrane transporter genes have been utilized for the improvement of crop plants, with enhanced nutrient uptake leading to increased crop yield by providing tolerance against different biotic and abiotic stresses. Further understanding of the basic mechanisms of nutrient transport in crop plants could facilitate the advanced design of engineered plant crops to achieve increased yield and improve nutrient quality through the use of genetic technologies as well as molecular breeding. This review is focused on nutrient toxicity and tolerance mechanisms in crop plants to aid in understanding and addressing the anticipated global food demand.

Crosstalk between phytohormones and secondary metabolites in the drought stress tolerance of crop plants: A review.

Drought stress negatively affects crop performance and weakens global food security. It triggers the activation of downstream pathways, mainly through phytohormones homeostasis and their signaling networks, which further initiate the biosynthesis of secondary metabolites (SMs). Roots sense drought stress, the signal travels to the above-ground tissues to induce systemic phytohormones signaling. The systemic signals further trigger the biosynthesis of SMs and stomatal closure to prevent water loss. SMs primarily scavenge reactive oxygen species (ROS) to protect plants from lipid peroxidation and also perform additional defense-related functions. Moreover, drought-induced volatile SMs can alert the plant tissues to perform drought stress mitigating functions in plants. Other phytohormone-induced stress responses include cell wall and cuticle thickening, root and leaf morphology alteration, and anatomical changes of roots, stems, and leaves, which in turn minimize the oxidative stress, water loss, and other adverse effects of drought. Exogenous applications of phytohormones and genetic engineering of phytohormones signaling and biosynthesis pathways mitigate the drought stress effects. Direct modulation of the SMs biosynthetic pathway genes or indirect via phytohormones' regulation provides drought tolerance. Thus, phytohormones and SMs play key roles in plant development under the drought stress environment in crop plants.

Dynamically tunable light responsive silk-elastin-like proteins.

Dynamically tunable biomaterials are of particular interest in the field of biomedical engineering because of the potential utility for shape-change materials, drug and cell delivery and tissue regeneration. Stimuli-responsive proteins formed into hydrogels are potential candidates for such systems, due to the genetic tailorability and control over structure-function relationships. Here we report the synthesis of genetically engineered Silk-Elastin-Like Protein (SELP) photoresponsive hydrogels. Polymerization of the SELPs and monomeric adenosylcobalamin (AdoB12)-dependent photoreceptor C-terminal adenosylcobalamin binding domain (CarHC) was achieved using genetically encoded SpyTag-SpyCatcher peptide-protein pairs under mild physiological conditions. The hydrogels exhibited a partial collapse of the crosslinked molecular network with both decreased loss and storage moduli upon exposure to visible light. The materials were also evaluated for cytotoxicity and the encapsulation and release of L929 murine fibroblasts from 3D cultures. The design of these photo-responsible proteins provides new stimuli-responsive SELP-CarHC hydrogels for dynamically tunable protein-based materials.

Functional Characterization of a Magnesium Transporter of Root Endophytic Fungus Piriformospora indica.

Magnesium (Mg) is a crucial macronutrient required for the regular growth of plants. Here we report the identification, isolation and functional characterization of Mg-transporter PiMgT1 in root endophytic fungus Piriformospora indica. We also report the role of P. indica in the improvement of the Mg nutrition of the plant particularly under Mg deficiency condition. Protein BLAST (BLASTp) for conserved domains analysis showed that PiMgT1 belong to CorA like protein family of bacteria. We have also observed the presence of conserved 'GMN' signature sequence which suggests that PiMgT1 belongs to Mg transporter family. Phylogenetic analysis revealed that PiMgT1 clustered among fungal CorA family members nearer to basidiomycetes. Functionality of PiMgT1 was confirmed by complementation of a yeast magnesium transporter mutant CM66. We have observed that PiMgT1 restored the growth of mutant and showed comparable growth with that of WT. We found statistically significant (p < 0.05) two fold increase in the total intracellular Mg content of mutant complemented with PiMgT1 as compared to the mutant. These observations suggest that PiMgT1 is actively involved in Mg uptake by the fungus and may be helping in the nutritional status of the host plant.

Antioxidant enzymes in chickpea colonized by Piriformospora indica participate in defense against the pathogen Botrytis cinerea.

Piriformospora indica, a root endophytic fungus, promotes growth of the economically important chickpea plant (Cicer arietinum Linn.) and protects it against the pathogenic fungus Botrytis cinerea. Biomass and root development were found to be significantly improved in chickpea plants colonized with P. indica as compared to the plants grown without P. indica as well as from the plants infected with the B. cinerea. Our PCR analyses showed that gradual increase in the colonization of P. indica in the plants result in the inhibition of the colonization of B. cinerea. P. indica colonized plants showed increased antioxidant enzyme activities. Interestingly, there were pronounced decrease in the antioxidant enzyme activities in shoots infected with B. cinerea and colonized with P. indica in alternate and simultaneous mode as compared to plants infected with B. cinerea alone. We conclude that P. indica helps plants to overcome the disease load by enhancing antioxidant enzyme defense system. Our data suggest that, bio-protective action of P. indica might be mediated via systemic induction of antioxidant defense in the host plants.

A single gene all3940 (Dps) overexpression in Anabaena sp. PCC 7120 confers multiple abiotic stress tolerance via proteomic alterations.

DNA-binding proteins (Dps) induced during starvation play an important role in gene regulation and maintaining homeostasis in bacteria. The nitrogen-fixing cyanobacterium, Anabaena PCC7120, has four genes annotated as coding for Dps; however, the information on their physiological roles is limiting. One of the genes coding for Dps, 'all3940' was found to be induced under different abiotic stresses in Anabaena and upon overexpression enhanced the tolerance of Anabaena to a multitude of stresses, which included salinity, heat, heavy metals, pesticide, and nutrient starvation. On the other hand, mutation in the gene resulted in decreased growth of Anabaena. The modulation in the levels of All3940 in Anabaena, achieved either by overexpression of the protein or mutation of the gene, resulted in changes in the proteome, which correlated well with the physiological changes observed. Proteins required for varied physiological activities, such as photosynthesis, carbon-metabolism, oxidative stress alleviation, exhibited change in protein profile upon modulation of All3940 levels in Anabaena. This suggested a direct or an indirect effect of All3940 on the expression of the above stress-responsive proteins, thereby enhancing tolerance in Anabaena PCC7120. Thus, All3940, though categorized as a Dps, is possibly a general stress protein having a global role in regulating tolerance to multitude of stresses in Anabaena.

MazF ribonucleases promote Mycobacterium tuberculosis drug tolerance and virulence in guinea pigs.

Toxin-antitoxin (TA) systems are highly conserved in members of the Mycobacterium tuberculosis (Mtb) complex and have been proposed to play an important role in physiology and virulence. Nine of these TA systems belong to the mazEF family, encoding the intracellular MazF toxin and its antitoxin, MazE. By overexpressing each of the nine putative MazF homologues in Mycobacterium bovis BCG, here we show that Rv1102c (MazF3), Rv1991c (MazF6) and Rv2801c (MazF9) induce bacteriostasis. The construction of various single-, double- and triple-mutant Mtb strains reveals that these MazF ribonucleases contribute synergistically to the ability of Mtb to adapt to conditions such as oxidative stress, nutrient depletion and drug exposure. Moreover, guinea pigs infected with the triple-mutant strain exhibits significantly reduced bacterial loads and pathological damage in infected tissues in comparison with parental strain-infected guinea pigs. The present study highlights the importance of MazF ribonucleases in Mtb stress adaptation, drug tolerance and virulence.

Differential response of oyster shell powder on enzyme profile and nutritional value of oyster mushroom Pleurotus florida PF05.

Oyster mushroom Pleurotus florida was cultivated on different combinations of wheat straw (WS) as basal substrate and oyster shell powder (OSP) supplement. The OSP supplementation considerably responded to different cultivation phases. The mycelium grew fast and showed rapid growth rate (8.91 mmd(-1)) in WS + OSP (97 + 3) combination while WS + OSP (92 + 8) showed maximum laccase (3.133 U/g) and Mn peroxidase (MnP) activities (0.091 U/g). The climax level of laccase (5.433 U/g) and MnP (0.097 U/g) was recorded during fruit body initiation in WS + OSP (97 + 3) and WS + OSP (98 + 2) combinations, respectively. The WS + OSP (97 + 3) combination represented the best condition for mushroom cultivation and produced the highest biological efficiency (147%). In addition, protein and lipid contents in fruit bodies were slightly improved in response to OSP. The carbohydrate was significantly increased by raising concentration of OSP. The highest values of protein, carbohydrate, and lipid noted were 31.3 µg/g, 0.0639 (g/g), and 0.373 (g/g) correspondingly. Conclusively it was evident that lower concentrations of OSP acted positively and relatively to higher concentrations and improved nutritional content which may suitably be used to enhance both yield and nutritional values of mushroom.

Cyanobacterial diversity shifts induced by butachlor in selected Indian rice fields in Eastern Uttar Pradesh and Western Bihar analyzed with PCR and DGGE.

The present study examines the effects of 30 mg/kg butachlor on the cyanobacterial diversity of rice fields in Eastern Uttar Pradesh and Western Bihar in India. A total of 40 samples were grouped into three classes [(i) acidic, (ii) neutral, and (iii) alkaline soils], based on physicochemical and principle component analyses. Acidic soils mainly harbored Westillopsis, Trichormus, Anabaenopsis, and unicellular cyanobacteria; whereas Nostoc, Anabaena, Calothrix, Tolypothrix, and Aulosira were found in neutral and alkaline soils. Molecular characterization using 16S rRNA PCR and DGGE revealed the presence of 13 different phylotypes of cyanobacteria in these samples. Butachlor treatment of the soil samples led to the disappearance of 5 and the emergence of 2 additional phylotypes. A total of 40 DGGE bands showed significant reproducible changes upon treatment with butachlor. Phylogenetic analyses divided the phylotypes into five major clusters exhibiting interesting links with soil pH. Aulosira, Anabaena, Trichormus, and Anabaenopsis were sensitive to butachlor treatment, whereas uncultured cyanobacteria, a chroococcalean member, Westillopsis, Nostoc, Calothrix, Tolypothrix, Rivularia, Gloeotrichia, Fischerella, Leptolyngbya, and Cylindrospermum, appeared to be tolerant against butachlor at their native soil pH. Butachlor-induced inhibition of nitrogen fixation was found to be 65% (maximum) and 33% (minimum) in the soil samples of pH 9.23 and 5.20, respectively. In conclusion, low butachlor doses may prove beneficial in paddy fields having a neutral to alkaline soil pH.

Iron starvation-induced proteomic changes in Anabaena (Nostoc) sp. PCC 7120: exploring survival strategy.

This study provides first-hand proteomic data on the survival strategy of Anabaena sp. PCC 7120 when subjected to long-term iron-starvation conditions. 2D-gel electrophoresis followed by MALDI-TOF/MS analysis of iron-deficient Anabaena revealed significant and reproducible alterations in ten proteins, of which six are associated with photosynthesis and respiration, three with the antioxidative defense system, and the last, hypothetical protein all1861, conceivably connected with iron homeostasis. Iron-starved Anabaena registered a reduction in growth, photosynthetic pigments, PSI, PSII, whole-chain electron transport, carbon and nitrogen fixation, and ATP and NADPH content. The kinetics of hypothetical protein all1861 expression, with no change in expression until day 3, maximum expression on the 7th day, and a decline in expression from the 15th day onward, coupled with in silico analysis, suggested its role in iron sequestration and homeostasis. Interestingly, the up-regulated FBP-aldolase, Mn/Fe-SOD, and all1861 all appear to assist the survival of Anabeana subjected to iron-starvation conditions. Furthermore, the N2-fixation capabilities of the iron-starved Anabaena encourage us to recommend its application as a biofertilizer, particularly in iron-limited paddy soils.