Biology subjects

Vetukuri, R. R.

Publications and source records attributed to Vetukuri, R. R..

8 recordsLinked to original sources

bioRxiv2022

Characterization of environmental effects on flowering and plant architecture in an everbearing strawberry F1-hybrid by meristem dissection and gene expression analysis

Floral transition in the cultivated everbearing strawberry is a hot topic because these genotypes flower perpetually and are difficult to maintain in a non-flowering state. However, it has rarely been studied using morphogenetic and molecular analyses simultaneously. We therefore examined morphogenetic effects and the activation of genes involved in floral induction and initiation in seedlings of an everbearing F1-hybrid. Seedlings were grown at 12, 19, and 26{degrees}C under 10-h SD and 20-h LD conditions. We observed a strong environmental influence on meristem development and a FLOWERING LOCUS T1 (FaFT1)-SUPPRESSOR OF OVEREXPRESSION OF CONSTANS1 (FaSOC1) pathway similar to that in the everbearing woodland strawberry. The everbearing cultivar showed typical features of a quantitative LD plant, flowering earlier under LD than SD conditions at all temperatures. We also found that floral induction is facilitated by FaFT1 upregulation under LD conditions, while FaSOC1 upregulation in the apex leads to photoperiod-independent floral initiation. Moreover, we confirmed the strawberry meristem identity gene FRUITFULL (FaFUL) can also be used as an early indicator of floral initiation in EB cultivars. This study also highlights the advantages of using seed-propagated F1-hybrids for genetic studies because are genetically identical, and not biased by a previous flowering history.

plant biology↗

bioRxiv2022

Bioassay evaluation, GC-MS Analysis and Acute Toxicity Assessment of Methanolic Extracts of Trichoderma species in an Animal Model

Fungi of the genus Trichoderma have been marketed for the control of crop pests, weeds, and diseases. However, some Trichoderma species may produce toxic secondary metabolites; the existence of these compounds in bioformulated products, along with their relative risk, should receive due attention to ensure human safety. In this study, we investigated the in vitro antagonistic potential of T. asperellum AU131 and T. longibrachiatum AU158 as a biocontrol agent against F. xylarioides and the associated antagonistic mechanism with bioactive substances. Swiss albino mice were used to evaluate the in vivo toxicity and pathogenicity of T. asperellum AU131 and T. longibrachiatum AU158 methanolic crude extracts and spore suspensions, respectively, in a preliminary safety assessment for use as biofungicides. Metabolite profiling of the crude extracts were performed by Gas Chromatography-Mass Spectrometry (GC-MS). The agar diffusion assay of the crude extracts from both T. asperellum AU131 and T. longibrachiatum AU158 were the most effective at a concentration of 200g/mL, causing 62.5%, and 74.3% inhibition, respectively. A GC-MS analysis of crude extracts from both bioagents identified 23 volatile secondary metabolites classified as alcohols, acids, sesquiterpenes, ketones and aromatic compounds. The oral administration of crude extracts (doses of 600, 1200, 2000 and 5000 mg/kg body weight) and spore suspensions (doses of 1 x 106, 1 x 107, 1 x 108 and 1 x 109 spores/mL) to female Swiss albino mice over 14 days did not show any significant signs of toxicity, mortality or changes to body weight. The highest dose, 5000 mg/kg, can be used to determine the no observed effect level of crude extracts and spore suspensions of T. asperellum AU131 and T. longibrachiatum AU158 when calculating safety margins. It can be concluded that these two Trichoderma species can serve as a biocontrol agents against F. xylarioides and the mechanism for this function was due to the secondary metabolites with effective bioactive substance. Moreover, the tested spore suspensions and crude extracts were not pathogenic or toxic, respectively, when administered to Swiss albino mice at various doses.

microbiology↗

bioRxiv2022

Comparative small RNA and degradome sequencing provide new insights into antagonistic interactions in the biocontrol fungus Clonostachys rosea

Necrotrophic mycoparasitism is an intricate process involving recognition, physical mycelial contact and killing of host fungi (mycohosts). During such interactions, mycoparasites undergo a complex developmental process involving massive regulatory changes of gene expression to produce a range of chemical compounds and proteins that contribute to the parasitism of the mycohosts. Small-RNAs (sRNAs) are vital components of post-transcriptional gene regulation, although their role in gene expression regulation during mycoparasitism remain understudied. Here, we investigated the role of sRNA-mediated gene regulation in mycoparasitism by performing sRNA and degradome tags sequencing of the mycoparasitic fungus Clonostachys rosea interacting with the plant pathogenic mycohosts Botrytis cinerea and Fusarium graminearum at two time points. The majority of differentially expressed sRNAs were down-regulated during the interactions with the mycohosts compared to a C. rosea self-interaction control, thus allowing de-suppression (up-regulation) of mycohost-responsive genes. Degradome analysis showed a positive correlation between high degradome counts and antisense sRNA mapping and led to the identification of 201 sRNA-mediated gene targets for 282 differentially expressed sRNAs. Analysis of sRNA gene targets revealed that the regulation of genes coding for membrane proteins was a common response against both mycohosts. While the regulation of genes involved in oxidative stress tolerance and cellular metabolic and biosynthetic processes was exclusive against F. graminearum highlighting common and mycohosts-specific gene regulation of C. rosea. By combining these results with transcriptome data collected in similar experimental conditions during a previous study, we expand the understanding of the role of sRNA in regulating interspecific fungal interactions and mycoparasitism. ImportanceSmall-RNAs (sRNAs) are emerging as key players in pathogenic and symbiotic fungus-plant interactions, however, their role in fungal-fungal interactions remains elusive. In this study, we employed the necrotrophic mycoparasite Clonostachys rosea and plant pathogenic mycohots Botrytis cinerea and Fusarium graminearum and investigated the sRNA-mediated gene regulation in mycoparasitic interactions. The combined approach of sRNA and degradome tag sequencing identified 201 sRNA-mediated putative gene targets for 282 differentially expressed sRNAs highlighting the role of sRNA-mediated regulation of mycoparasitism in C. rosea. We also identified 36 known and 13 novel miRNAs and their potential gene targets at endogenous level, and at a cross-species level in B. cinerea and F. graminearum indicating a role of cross-species RNAi in mycoparasitism, representing a novel mechanism in biocontrol interactions. Furthermore, we showed that C. rosea adapts its transcriptional response, and thereby its interaction mechanisms, based on the interaction stages and identity of the mycohost.

microbiology↗

bioRxiv2022

Biodiversity of the Genus Trichoderma in the Rhizosphere of Coffee (Coffea arabica) Plants in Ethiopia and their Potential Use in Biocontrol of Coffee Wilt Disease

The present study investigated the distribution status and biodiversity of Trichoderma species surveyed from coffee rhizosphere soil samples from Ethiopia and their potential for biocontrol of coffee wilt disease (CWD) caused by Fusarium xylarioides. Trichoderma isolates were identified based on molecular approaches and morphological characteristics followed by biodiversity analysis using different biodiversity indices. The antagonistic potential of Trichoderma isolates was evaluated against F. xylarioides using the dual confrontation technique and agar diffusion bioassays. A relatively high diversity of species was observed including 16 taxa and 11 undescribed isolates. Trichoderma asperellum, T. asperelloides and T. longibrachiatum were classified as abundant species, with dominance (Y) values of 0.062, 0.056 and 0.034, respectively. Trichoderma asperellum was the most abundant species (comprising 39.6% of all isolates) in all investigated coffee ecosystems. Shannons biodiversity index (H), the evenness (E), Simpsons biodiversity index (D), and the abundance index (J) were calculated for each coffee ecosystem, revealing that species diversity and evenness were highest in the Jimma zone (H =1.97, E = 0.76, D = 0.91, J = 2.73). The average diversity values for Trichoderma species originating from the coffee ecosystem were H = 1.77, D = 0.7, E = 0.75 and J = 2.4. In vitro confrontation experiments revealed that T. asperellum AU131 and T. longibrachiatum AU158 reduced the mycelial growth of F. xylarioides by over 80%. The potential use of these Trichoderma species for disease management of F. xylarioides and to reduce its impact on coffee cultivation is discussed in relation to Ethiopias ongoing coffee wilt disease crisis.

microbiology↗

bioRxiv2021

A family of cell wall transglutaminases is essential for appressorium development and pathogenicity in Phytophthora infestans

Transglutaminases (TGases) are enzymes highly conserved among prokaryotic and eukaryotic organisms, where their role is to catalyse protein cross-linking. One of the putative TGases of Phytophthora infestans has previously been shown to be localised to the cell wall. Based on sequence similarity we were able to identify six more genes annotated as putative TGases and show that these seven genes group together in phylogenetic analysis. All of the seven proteins are predicted to contain transmembrane helices and both a TGase domain and a MANSC domain, the latter of which was previously shown to play a role in protein stability. Chemical inhibition of transglutaminase activity and silencing of the entire family of the putative cell wall TGases are both lethal to P. infestans indicating the importance of these proteins in cell wall formation and stability. The intermediate phenotype obtained with lower drug concentrations and less efficient silencing displays a number of deformations to germ tubes and appressoria. Both chemically treated and silenced lines show lower pathogenicity than the wild type in leaf infection assays. Finally, we show that appressoria of P. infestans possess the ability to build up turgor pressure and that this ability is decreased by chemical inhibition of TGases.

molecular biology↗

bioRxiv2021

Mycorrhizal inoculation and treatment with biochar and compost from pruning waste improve the qualitative properties of a calcareous soil under wheat cultivation

Most calcareous soils have relatively low levels of organic matter. To address this issue and improve the qualitative properties of calcareous soils, soils can be treated with mycorrhizal fungi and/or exogenous organic material such as biochar or compost derived from tree pruning waste. To evaluate the effect of pruning waste biochar (PWB) and pruning waste compost (PWC) derived from apple and grape trees combined with arbuscular mycorrhizal fungi (AMF) on the biological indices of calcareous soils, a rhizobox study on wheat plants using a completely randomized design was conducted under greenhouse conditions. The studied factors included the source of the type of organic material applied (PWB, PWC, and control), the nature of the microbial inoculation (inoculation with AMF or no inoculation), and the zone to which the treatments were applied (rhizosphere and non-rhizosphere soil). At the end of the plant growth period, organic carbon (OC), microbial biomass carbon (MBC), microbial biomass phosphorous (MBP), microbial respiration (BR), substrate-induced respiration (SIR), alkaline (ALP), acid (ACP) phosphatase enzyme activities in the rhizosphere and non-rhizosphere soils, and root mycorrhizal colonization were determined. Simultaneous application of a source of organic matter and AMF inoculation significantly increased the OC and biological indices of soil relative to those observed when applying organic matter without AMF inoculation. Additionally, MBC, MBP, ACP and ALP - enzymes activities in the rhizosphere zone were significantly higher than in the non-rhizosphere. AMF increased BR and SIR levels in the rhizosphere by 13.06% and 7.95% compared to non-rhizosphere, respectively. It can be concluded that in calcareous soils with low organic carbon contents, organic amendments such as PWC and PWB can improve soil biological properties by increasing microbial activity and changing the properties of the rhizosphere.

microbiology↗

bioRxiv2021

The role of Dicer-dependent RNA interference in regulating cross-species communication during fungus-fungus interactions

Dicer-like (DCL) proteins play a vital role in transcriptional and post-transcriptional gene silencing, also known as RNA interference (RNAi), by cleaving double-stranded RNAs or single-stranded RNAs with stem-loop structures into small RNAs. Although DCL-mediated RNAi can regulate interspecific communication between pathogenic/mutualistic organisms and their hosts, its role in parasitic fungus-fungus interactions is yet to be investigated. In this study, we deleted dcl genes in the mycoparasitic fungus Clonostachys rosea and analyzed the transcriptome and secondary metabolome to characterize the regulatory functions of DCL-dependent RNAi in mycoparasitism. Deletion of dcl2 resulted in a mutant with reduced growth rate, pigment production and antagonism towards the plant pathogenic fungus Botrytis cinerea. Moreover, the {Delta}dcl2 mutant displayed a reduced ability to control fusarium foot rot disease on wheat, caused by Fusarium graminearum, and reduced production of 62 secondary metabolites (SM) including yellow-coloured sorbicillinoids. Transcriptome sequencing of the in vitro interaction between the C. rosea {Delta}dcl2 strain and B. cinerea or F. graminearum identified downregulation of genes coding for transcription factors, membrane transporters, hydrolytic enzymes and SM biosynthesis enzymes putatively involved in antagonistic interactions, in comparison with the C. rosea wild type interaction. Sixty-one putative novel microRNA-like RNAs (milRNAs) were identified in C. rosea, and 11 was upregulated in the {Delta}dcl2 mutant. In addition to putative endogenous gene targets, these DCL2-dependent milRNAs were predicted to target B. cinerea and F. graminearum virulence factor genes, which showed an increased expression during interaction with the {Delta}dcl2 mutant incapable of producing the targeting milRNAs. This paper constitutes the first step in elucidating the role of RNAi in mycoparasitism, with important implications for biological control of plant diseases. This study further indicates a possible cross-species regulatory activity of fungal milRNAs, emphasizing a novel role of RNAi in fungal interactions and ecology. Author summaryRNA interference (RNAi) is a conserved cellular mechanism mediated by small RNAs (sRNAs) regulating biological processes through the targeted destruction or modulation of RNA filaments necessary for protein synthesis. Dicer-like endoribonucleases (DCL) play a vital role in the RNAi pathway by generating sRNAs. In this study, we identified two DCL-encoding genes in the mycoparasitic fungus Clonostachys rosea and investigated a role of DCL-mediated RNAi in interference interactions between Clonostachys rosea and the two important fungal pathogens Botrytis cinerea and Fusarium graminearum (here called mycohost). Using transcriptome (sRNA and mRNA) sequencing and secondary metabolome analysis approach, we found that the dcl mutants were not able to produce 11 sRNAs predicted to finetune the regulatory network of genes known to be involved in production of hydrolytic enzymes, antifungal compounds, and membrane transporters needed for antagonistic action of C. rosea. We also found C. rosea sRNAs putatively targeting known virulence factors in the mycohost, indicating RNAi-mediated cross-species communication. Our study expanded the understanding of underlying mechanisms of cross-species communication during interference interactions and showed that DCL-mediated RNAi is an important regulator of parasitic fungus-fungus interactions. The results pose the base for future works studying the role of DCL-based cross-species RNAi in fungal interactions.

genomics↗

bioRxiv2021

Spray-induced gene silencing as a potential tool to control potato late blight disease

Phytophthora infestans causes late blight disease on potato and tomato and is currently controlled by resistant cultivars or intensive fungicide spraying. Here, we investigated an alternative means for late blight control by spraying potato leaves with double-stranded RNAs (dsRNA) that target P. infestans genes that are essential for infection. Through confocal microscopy, we show that the sporangia of P. infestans expressing Green Fluorescent Protein (GFP) can take up in vitro synthesized dsRNAs homologous to GFP directly from their surroundings, including leaves, which leads to the reduced relative expression of GFP. We further demonstrate the potential of spray induced gene silencing (SIGS) in controlling potato late blight disease by targeting developmentally important genes in P.infestans such as guanine-nucleotide binding (G) protein {beta}-subunit (PiGPB1), haustorial membrane protein (PiHmp1), cutinase (PiCut3), and endo-1,3(4)-{beta}-glucanase (PiEndo3). Our results demonstrate that SIGS can be potentially used to mitigate potato late blight; however, the degree of disease control is dependent on the selection of the target genes.

plant biology↗