Expression analysis of genes involved in teliospores germination of Tilletia indica inciting Karnal bunt of wheat.

BACKGROUND: Karnal bunt of wheat is an important quarantine disease, incited by Tilletia indica. It limits India's trade in wheat export. The teliospores are major source of inoculum to initiate and spread the Karnal bunt disease. The study aimed to identify the germination-related genes in the teliospores of T. indica. METHODS AND RESULTS: The candidate genes in the teliospores germination were identified through the differential gene expression analysis with suitable bioinformatics analysis. Keeping in soil-borne nature of fungi, the teliospores of T. indica (2015 and 2018) were subjected to the qPCR analysis. 20 candidate genes were identified having role in germination of teliospores of T. indica. Twenty genes, viz. Ti9297 (9.31, 7.87-fold), Ti8696 (5.13, 6.54-fold), Ti7699 (8.9, 7.7-fold), Ti7858 (10.33, 6.21-fold), Ti7954 (7.46, 5.54-fold), Ti7739 (5.46, 6.46-fold), Ti9665 (10.74, 7.64-fold), Ti9335 (6.75, 4.36-fold), Ti8396 (9.35, 7.72-fold), Ti8126 (8.87, 11.31-fold), Ti7326 (6.04, 7.7-fold), Ti10208 (13.83, 5.81-fold), Ti12356 (7.83, 8.02-fold), Ti14271 (9.98, 6.32-fold), Ti9234 (11.2, 8.72-fold), Ti 8876 (6.47, 3.55-fold), Ti 10,606 (4.97, 2.35-fold), Ti7758 (10.33, 8.78-fold), Ti4692 (6.89, 9.88-fold), and Ti3932 (5.77, 4.5-fold) were found highly expressed in the germinating teliospores of 2015 and 2018, respectively. Eight genes (Ti508, Ti4152, Ti5346, Ti2375, Ti3739, Ti1134, Ti4399, and Ti4422) were downregulated in the germinating teliospores but these eight genes were showed higher expression in the dormant teliospores. CONCLUSIONS: Twenty candidate genes were upregulated in the germinating teliospores are supposed to be involved in the process of germination. Eight genes were downregulated which were related to the process of the dormancy of teliospores. The study will be helpful to devise the newer management strategies for Karnal bunt disease of wheat.

High-Quality Genome Resource of Ustilaginoidea virens (UV2_4G), Causal Agent of an Emerging False Smut Disease in Rice.

Ustilaginoidea virens is the fungal pathogen causing an emerging false smut disease that affects crop yield as well as deteriorates quality of the grains by producing mycotoxins. A high quality genome of U. virens isolate UV2_4G was sequenced using Nanopore and Illumina HiSeq 2,000 sequencing platforms. The total assembled genome of Indian isolate UV2_4G was 35.9 Mb, which comprised 89 scaffolds with N50 of 700,296 bp. A total of 358,697 variants were identified in the genome, out of which 355,173 were SNPs and 3,524 were INDELS. Further, 7,390 SSRs belonging to different repeat types were also identified in the genome. Out of 7,444 proteins predicted, 7,206 were functionally annotated. A total of 1,307 CAZymes, 501 signal peptides, 1,876 effectors, and 2,709 genes involved in host-pathogen interactions were identified. Comparative analysis revealed isolate UV2_4G is distinct with 31 unique clusters and placed distantly in phylogenetic analysis. Taken together, this high-quality genome assembly and sequence annotation resource can give an improved insight for characterizing the biological and pathogenic mechanisms of U. virens.

Transcriptomic analysis of bakanae disease resistant and susceptible rice genotypes in response to infection by Fusarium fujikuroi.

BACKGROUND: Fusarium fujikuroi causing bakanae is one of the most significant pathogens of rice and much responsible for yield losses thereby emerging as a major risk to food security. METHODS: In the present study transcriptomic analysis was conducted between two contrasting resistant (C101A51) and susceptible (Rasi) genotypes of rice with the combinations of C101A51 control (CC) vs. C101A51 inoculated (CI); Rasi control (RC) vs. Rasi inoculated (RI) and C101A51 inoculated (CI) vs. Rasi inoculated (RI). RESULTS: In CC vs. CI commonly expressed genes were 12,764. Out of them 567 (4%) were significantly upregulated and 1399 (9%) genes were downregulated. For the RC vs. RI 14, 333 (79%) genes were commonly expressed. For CI vs. RI 13,662 (72%) genes were commonly expressed. Genes related to cysteine proteinase inhibitor 10, disease resistance protein TAO1-like, oleosin 16 kDa-like, pathogenesis-related protein (PR1), (PR4), BTB/POZ and MATH domain-containing protein 5-like, alpha-amylase isozyme were upregulated in resistant genotype C101A51. Whereas, genes related to GDSL esterase/lipase, serine glyoxylate aminotransferase, CASP-like protein 2C1, WAT1-related protein, Cytoplasmic linker associated proteins, xyloglucan endotransglucosylase/hydrolase protein and ß-D xylosidase 7 were upregulated in susceptible genotype Rasi. Gene ontology analysis showed functions related to defence response (GO:0006952), regulation of plant hypersensitive type response (GO:0010363), Potassium ion transmembrane activity (GO:0015079), chloroplast (GO:0009507), response to wounding (GO:0009611), xylan biosynthetic process (GO:0045492) were upregulated in resistant genotype C101A51 under inoculated conditions. CONCLUSION: Real time PCR based validation of the selected DEGs showed that the qRT-PCR was consistent with the RNA-Seq results. This is the first transcriptomic study against bakanae disease of rice in Indian genotypes. Further, functional studies on identified genes and their utilization through different methodology will be helpful for the development of bakanae disease management strategies.

Metabolomic analysis of sheath blight disease of rice (Oryza sativa L.) induced by Rhizoctonia solani phytotoxin.

AIM: To understand the mechanism of necrosis incited by a host-selective phytotoxin designated as Rhizoctonia solani toxin (RST) identified to be a potential pathogenic factor of R. solani AG1 IA, causing sheath blight (ShB) of rice. METHODS AND RESULTS: The metabolomic changes induced by the phytotoxic metabolite in a ShB susceptible rice cultivar were elucidated by gas chromatography-mass spectrometry analysis and compared with that of the pathogen to identify rice metabolites targeted by the phytotoxin. The profiles of about 29 metabolites with various physiological roles in rice plants have been identified worldwide. Unsupervised and supervised multivariate chemometrics (principal component analysis and partial least squares-discriminant analysis) and cluster (Heat maps) analyses were used to compare the metabolites obtained from chemical profiles of the treatments with sterile distilled water (SDW) control. The results indicated that the rice plant expressed more metabolites in response to the pathogen than the phytotoxin and was lowest in SDW control. The key metabolites expressed in rice in response to the treatments were investigated by the variable importance in projection (VIP) analysis using p < 0.05 VIP >15. The analysis identified 7 and 11 upregulating metabolites in the phytotoxin and the pathogen treatments, respectively, compared to the untreated control. Among the phytotoxin-treated and the pathogen inoculated samples, the phytotoxin-treated sample recorded upregulation of six metabolites, whereas nine metabolites were upregulated in the pathogen-inoculated samples. These upregulating metabolites are speculated for the necrotic symptoms characteristic to both the phytotoxin and pathogen. In this analysis, hexadecanoic acid and dotriacontane were highly expressed metabolites specific to the phytotoxin and pathogen-treated samples, respectively. Besides upregulation, the metabolites also have a VIP score of >1.5 and hence fulfilled the criteria of classifying them as reliable potential biomarkers. In the pathway analysis, hexadecanoic acid and dotriacontane were identified to be involved in several important biosynthetic pathways of rice, such as the biosynthesis of saturated fatty acid and unsaturated fatty acids cutin, suberin and wax. CONCLUSIONS: The study concludes that though certain metabolites induced by the phytotoxin in the susceptible variety during necrosis shares with that of the pathogen, the identification of metabolites specific to the phytotoxin in comparison to the pathogenic and SDW controls indicated that the phytotoxin modulates the host metabolism differently and hence can be a potential pathogenicity factor of the ShB fungus. SIGNIFICANCE AND IMPACT OF THE STUDY: Due to lack of knowledge on the pathway genes of RST and in the absence of an ShB-resistant variety, understanding differentially expressed metabolic changes induced in the susceptible variety by the phytotoxin in comparison to that of the pathogenic and uninoculated controls enables us to identify the key metabolite changes during the ShB infection. Such metabolomic changes can further be used to infer gene functions for exploitation in ShB control.

Physiological race characterisation of Fusarium oxysporum strains infecting tomato employing candidate pathogenicity genes and host resistance.

Physiological races of 14 strains of Fusarium oxysporum f.sp. lycopersici were established by PCR profiling SIX gene expressions. No amplification of the SIX4 (Avr1) gene was observed in any of the 14 strains. Based on amplification of the SIX3 (Avr2) gene, 6 strains were distinguished as race 2. Race 2 strains are known to contain identical SIX3 sequences and differ from race 3 strains by single point mutations. Hence, based on polymorphic amplicons of the SIX3 gene detected by stringent PCR conditions, 8 strains were identified as race 3. The identity of the physiological races of the strains was validated by inoculating on three germplasm lines, EC-814916, FEB-2 and Pusa Rohini carrying I-2, I-3 and no I gene, respectively. The race 2 and race 3 strains were avirulent on EC-814916 and FEB-2 lines, respectively. All the 14 fungal strains were pathogenic on Pusa Rohini, the Fusarium wilt susceptible cultivar lacking R genes and exhibited different levels of virulence. In evaluating two other potential pathogenicity genes, Fow1 and Fow2 as markers for virulence, their expressions were observed among both the races of the Fol strains, and hence are not potential candidates for physiological race discrimination. However, strong expressions of the genes in the root tissues inoculated with the highly virulent strain, TOFU-IHBT in comparison to the uninoculated control indicated their roles in fungal pathogenicity. To understand the role of these pathogenicity genes in countering the host defence mechanisms, their expressions in response to ROS and phenolics, the earliest known defence mechanisms of host plants were assessed. In H2O2, the Fow2 gene expressed 1.4-fold greater than that of the control. On the contrary, in relation to the control, the expressions of Fow1 were strongly repressed exhibiting 0.7-to 0.8-fold lesser at 0.1 mM through 3 mM concentrations than that of the control indicating that the gene is modulated by the phenolic acid indicating the roles of Fow2 and Fow1 in alleviating oxidative stress and targeted by the phenolic acid, respectively.

First Report of Sclerotium hydrophilum Sace Causing Stem Rot Disease of Rice in North Eastern Plain Zone of India.

High incidence of stem rot in rice (Oryza sativa L.) caused by Sclerotium hydrophilum was observed in Eastern Gangetic plains of India including eastern Uttar Pradesh and Bihar states in rice-wheat irrigated ecosystem in the months of August and September of the years 2016-17 at the maximum tillering stage of the crop plants. A survey was conducted for the prevalence of rice diseases in Eastern Uttar Pradesh and Bihar. Stem rot incidence (12-14%) was observed in widely cultivated rice varieties viz., Swarna, Sabour Surbhit, Samba Mahsuri and Bhagalpur katarani. Brownish to black water-soaked lesions without distinct margins were visually observed on rice leaf sheaths in Pusa, Bihar and Allahabad, Uttar Pradesh (India). To isolate the causal pathogen, pieces of symptomatic leaf sheaths were treated with 1% sodium hypochlorite solution for 1 min, rinsed with sterile distilled water for 2 min, and then transferred to potato dextrose agar (PDA) medium for incubation at 280C. The cultivated mycelium was transferred to fresh PDA medium. Colonies of pure cultures on PDA medium were initially white and turned brown about 2 weeks later. The hyphal width was measured with a range of 4.0 to 6.0 µm. Large numbers of small globose sclerotia were observed on surface of the colonies at 5 days after sub-culturing. The sclerotia were white at first and then turned black over time with maturity. The diameters of sclerotia ranged from 0.32 to 0.51 mm with an average of 0.41 mm (n = 50). DNA of a representative isolate named SH1 was extracted, and the ITS region was amplified by PCR with universal primer pair ITS1/ITS4 (White et al. 1990). Sequence analysis showed 99.66% identity with Sclerotium hydrophilum isolate VC228 (accession no. KT362098) and accession no. obtained (KX181457). Phylogenetic analysis based onneighbour-joining method grouped the isolatesalongwith other isolates from Asia. The pathogen was identified as Sclerotium hydrophilum on the basis of cultural, morphological, pathogenic and molecular characteristics. Further, culture was deposited to Indian Type Culture Collection (ITCC) at the Indian Agricultural Research Institute, New Delhi for the identification where it was identified as Sclerotium hydrophilum with identification number 10039.15. Another culture specimen of the pathogen was also deposited at the ITCC-IARI, New Delhi with accession no. 19160.25. Koch's postulates were completed by inoculating rice plants with PDA disks of5 mm in size bearing both mycelium and sclerotia of S. hydrophilum in 45 days old plants of rice genotype 'Samba Mahsuri' under glass house conditions. After 4th days leaf sheath necrosis was observed around the inoculation plugsunder field conditions.The same fungus was re isolated from the symptomatic lesions of inoculated plants, fulfilling Koch's postulates. Pathogen was reported with high disease incidence in Northern Karnataka (Pramesh et al. 2017). However, based on literature and fungus-host distribution records this is the first report of S. hydrophilum on leaf sheath of rice from North-Eastern plain zone of India.

First report of stem smut caused by Tranzscheliella hypodytes (Schltdl.) Vanky & McKenzie on Leymus secalinus (Georgi) Tzvelev in India.

Leymus secalinus (Blue wild rye) is a perennial grass species distributed in Leh-Ladakh region of India. Culms are usually solitary, 20-100 cm tall, 2-5-noded, smooth and glabrous. It is found on mountain slopes, rocky, stony and pebbled soils, grassy places, river banks, sandy and alkaline soils. It is one of the dominant species of the region and is mostly used for forage and grazing. L. secalinus plants with blackish-brown powdery spore mass/sori on the culm was observed in Leh region of Jammu and Kashmir, India during a wheat germplasm exploration (to collect wild relatives, land races, cultivars etc. of cultivated wheat) in September, 2018. Initially, sori were covered by the leaf sheath and at later stage more or less exposed with the absence of peridium. Infected culms and leaves are stunted, while inflorescences are abortive. Spores are globose, sub-globose to ovoid, blackish-brown in color, 3-5 x 4-4.5 µm in size, wall 0.5 µm thick and smooth. The fungus was identified as Tranzscheliella hypodytes (Schltdl.) Vanky & McKenzie based on morphological characteristics (Li et al. 2017; Vanky 2012, 2003; Vanky and McKenzie 2002). The ustilospore germination was not observed on potato dextrose agar during the lab experiment. In some cases, ustilospore germination is recorded in other smut fungi which shows considerable variation according to media and conditions used (Ingold, 1983). Moreover, there is no report available for the artificial culture of T. hypodytes. Identity was confirmed using the internal transcribed spacer (ITS) region of ribosomal DNA, amplified with universal primers ITS1 which was then sequenced. NCBI-BLAST showed 99.28% similarity with the ITS sequences of Tranzscheliella hypodytes (GenBank accession no. MH855460.1). The sequence was deposited in NCBI GenBank with accession number (MN108147). Pathogenecity test was conducted in glasshouse wherein smut infected samples of L. secalinus collected from Leh were made into small pieces and mixed with the sterilized soil filled in three pots before sowing. In addition, seeds were also mixed with smut spores before sowing. Inoculated pots after sowing were kept at 16±20C temperature and a relative humidity of 70% in a glass house. Control pots were maintained without inoculum. Inoculated and control pots were placed in separate chambers of the same glasshouse. Seventy days after sowing, inoculated plants were symptomatic, matching those described above and produced no seed. Pathogen reisolated from infected plants was identical in all respects to the isolate used to inoculate the plants, which confirmed the Koch's postulates. Control plants remained healthy and produced seed. Leh region is covered by sparse vegetation due to its topography and cold desert climate (cold, dry winter and warm, dry summer with very low precipitation, occuring in temperate region). Stem smut in L. secalinus can have significant negative impact on the fodder and grazing situation of the region. To our knowledge, this is the first report of stem smut caused by T. hypodytes on L. secalinus in India. A voucher specimen of the fungus was deposited at Herbarium Cryptogamae Indiae Orientialis (HCIO) (52182), ICAR-Indian Agricultural Research Institute, New Delhi.

Molecular characterization of Indian pathotypes of Puccinia striiformis f. sp. tritici and multigene phylogenetic analysis to establish inter- and intraspecific relationships.

Stripe rust caused by Puccinia striiformis f. sp. tritici (Pst) is one of the most devastating diseases of wheat (Triticum spp.) worldwide. Indian isolates were characterised based on their phenotypic reaction on differential hosts carrying different Yr genes. Based on virulence/avirulence structure, isolates were characterised into ten different pathotypes viz. 70S0-2, 67S64, 70S4, 66S0, 70S64, 66S64-1, 38S102, 47S102, 46S119, and 78S84. These Indian pathotypes of P. striiformis f. sp. tritici and 38 pathotypes of other rust species (P. graminis tritici and P. triticina) were used in this study to analyze their molecular phylogenetic relationship. The nucleotides of rDNA-ITS, partial ß-tubulin and ketopantoate reductase genes of all the pathotypes were sequenced directly after PCR. Based on sequence data of rDNA-ITS and ß-tubulin, three phylogenetic groups corresponding to three different species of Puccinia were obtained. Asian isolates formed a distinct evolutionary lineage than from those derived from USA. The sequence similarity of Indian pathotypes with other Asian (China and Iran) isolates indicated the same origin of pathotypes. The results will allow rapid identification of Indian P.striiformis f. sp. tritici pathotypes causing stripe rust in wheat, assist in making predictions regarding potential rust pathotypes, and identifying sources of resistance to the disease in advance.

Corrigendum: Unraveling the dynamics of wheat leaf blight complex: isolation, characterization, and insights into pathogen population under Indian conditions.

[This corrects the article DOI: 10.3389/fmicb.2024.1287721.].

Unraveling the dynamics of wheat leaf blight complex: isolation, characterization, and insights into pathogen population under Indian conditions.

Wheat, a staple food crop for 35% of the global population, faces a threat from Helminthosporium leaf blight (HLB), a complex of spot blotch (Bipolaris sorokiniana) and tan spot (Pyrenophora-tritici-repentis) diseases under warm and humid conditions. However, in Indian conditions, the knowledge of existing pathogen populations associated with the HLB complex is limited and largely dominated by only B. sorokiniana (spot blotch). To address this, diseased samples were collected from all six wheat growing zones during 2020-2022. The pathogenic species were identified through in-depth morphological characterization, supplemented with ITS-rDNA and GAPDH sequence analysis, a diagnostic SCAR marker, and pathogenicity studies on two wheat varieties: Sonalika and HD2733. The 32 isolates collected from 10 different states consist of B. spicifera (12.5% of all isolates), Exserohilum rostratum (9.3%), Bipolaris oryzae (3.1%), and B. sorokiniana (75%). B. sorokiniana exhibited the highest disease severity on both varieties. Other lesser-known pathogenic species also produced comparable disease severity as B. sorokiniana isolates and, therefore are economically important. Unraveling pathogen composition and biology aids in disease control and resistance breeding. Our study highlights economically impactful and lesser-known pathogenic species causing wheat leaf blight/spot blotch in India, guiding both current management and future resistance breeding strategies in plant pathology.

Identification and Characterization of Fusarium fujikuroi Pathotypes Responsible for an Emerging Bakanae Disease of Rice in India.

The bakanae disease of rice, or foolish seedling disease, is a well-known pathogen infecting rice hosts. Several studies have characterized Fusarium fujikuroi isolates collected from distant geographical regions and within similar geographical areas for secondary metabolite production, population structure, and diversity analysis, but none have attempted to characterize the isolates for virulence in a differential set of rice genotypes. Based on the disease response, a set of five rice genotypes with differing resistance levels were selected as a differential set for further characterization of the pathogen. Ninety-seven Fusarium fujikuroi isolates collected from different rice-growing areas of the country during the years 2011 to 2020 were characterized and evaluated for bakanae disease. Rice genotypes PB1509 and C101A51 were found to be highly susceptible and highly resistant, respectively. Further, based on the disease response, the isolates were grouped into 15 pathotypes. Pathotype 1, with the maximum isolates (19), was observed to be most prevalent, followed by pathotypes 2 and 3. Pathotype 8 was classified as highly virulent, as all the genotypes were susceptible, except for C101A51. When we compared the pathotype distribution in different states, pathotypes 11 and 15 were found to have originated from the state of Punjab. A positive correlation could be established between six pathotype groups and the gene expression of virulence-related genes such as acetylxylan (FFAC), exopolygalacturanase (FFEX), and pisatin demethylase (FFPD). The present study provides the distribution profiles of different pathotypes in Basmati-growing states of India, which will be further helpful for the deployment of breeding strategies and bakanae disease management.

Impact of Fusarium Infection on Potato Quality, Starch Digestibility, In Vitro Glycemic Response, and Resistant Starch Content.

Potato dry rot disease caused by multiple Fusarium species is a major global concern in potato production. In this investigation, the tubers of cultivars Kufri Jyoti and Kufri Frysona were artificially inoculated with an individual or combined inoculum of Fusarium sambucinum and Fusarium solani. Fusarium sambucinum caused a significantly higher lesion development (p < 0.01) than Fusarium solani, irrespective of cultivars. The combined inoculum of both the Fusarium species caused significantly higher rot development (p < 0.005) in inoculated tubers. Analyses of starch and amylose content revealed that individual or mixed infection of fungi caused a significant reduction (p < 0.005) in these parameters compared to healthy tubers. The increased starch digestibility due to fungal infection caused a higher glycemic index and glycemic load. The resistant starch also deteriorated in the infected potato tubers as compared to the control. Kufri Jyoti showed a higher starch and amylose content reduction in response to the treatments compared to Kufri Frysona. The correlation analysis demonstrated a negative correlation in lesion diameter and rot volume with starch and amylose content (p < -0.80). However, the glycemic index and resistant starch were positively correlated with lesion development. Altogether, these findings highlight the progressive deterioration of quality parameters, which will be a critical concern for processing industry stakeholders and consumers.

Corrigendum: Temporal transcriptome of tomato elucidates the signaling pathways of induced systemic resistance and systemic acquired resistance activated by Chaetomium globosum.

[This corrects the article DOI: 10.3389/fgene.2022.1048578.].

Characterization and development of transcriptome-derived novel EST-SSR markers to assess genetic diversity in Chaetomium globosum.

Chaetomium globosum Kunze, an internationally recognized biocontrol fungus. It mycoparasitizes various plant pathogens and produce antifungal metabolites to suppress the growth of pathogenic fungi. Lack of detailed genome level diversity studies has delimited the development and utilization of potential C. globosum strains. The present study was taken to reveal the distribution, identification, and characterization of expressed sequence tag-simple sequence repeats (EST-SSRs) in C. globosum. RNA-Seq experiment was performed for C. globosum potential isolate Cg2 (AY429049) using Illumina HiSeq 2500. Reference-guided de novo assembly yielded 45,582 transcripts containing 27,957 unigenes. We generated a new set of 8485 EST-SSR markers distributed in 5908 unigene sequences with one SSR locus distribution density per 6.1 kb. Six distinct classes of SSR repeat motifs were identified. The most abundant were mononucleotide repeats (51.67%), followed by tri-nucleotides (36.61%). Out of 5034 EST-SSR primers, 50 primer pairs were selected and validated for the polymorphic study of 15 C. globosum isolates. Twenty-two SSR markers showed average genetic polymorphism among C. globosum isolates. The number of alleles (Na) per marker ranges from 2 to 4, with a total of 74 alleles detected for 22 markers with a mean polymorphism information content (PIC) value of 0.4. UPGMA hierarchical clustering analysis generated three main clusters of C. globosum isolates and exhibited a lower similarity index range from 0.59 to 0.85. Thus, the newly developed EST-SSR markers could replace traditional methods for determining diversity. The study will also enhance the genomic research in C. globosum to explore its biocontrol potential against phytopathogens. Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-023-03794-7.

Draft genome sequencing of Tilletia caries inciting common bunt of wheat provides pathogenicity-related genes.

Common bunt of wheat caused by Tilletia caries is an important disease worldwide. The T. caries TC1_MSG genome was sequenced using the Illumina HiSeq 2500 and Nanopore ONT platforms. The Nanopore library was prepared using the ligation sequencing kit SQK-LSK110 to generate approximately 24 GB for sequencing. The assembly size of 38.18 Mb was generated with a GC content of 56.10%. The whole genome shotgun project was deposited at DDBJ/ENA/GenBank under the accession number JALUTQ000000000. Forty-six contigs were obtained with N50 of 1,798,756 bp. In total, 10,698 genes were predicted in the assembled genome. Out of 10,698 genes, 10,255 genes were predicted significantly in the genome. The repeat sequences made up approximately 1.57% of the genome. Molecular function, cellular components, and biological processes for predicted genes were mapped into the genome. In addition, repeat elements in the genome were assessed. In all, 0.89% of retroelements were observed, followed by long terminal repeat elements (0.86%) in the genome. In simple sequence repeat (SSR) analysis, 8,582 SSRs were found in the genome assembly. The trinucleotide SSR type (3,703) was the most abundant. Few putative secretory signal peptides and pathogenicity-related genes were predicted. The genomic information of T. caries will be valuable in understanding the pathogenesis mechanism as well as developing new methods for the management of the common bunt disease of wheat.

Cloning and expression of a small heat and salt tolerant protein (Hsp22) from Chaetomium globosum.

The present study reports molecular characterization of small heat shock protein gene in Indian isolates of Chaetomium globosum, C. perlucidum, C. reflexum, C. cochlioides and C. cupreum. Six isolates of C. globosum and other species showed a band of 630bp using specific primers. Amplified cDNA product of C. globosum (Cg 1) cloned and sequenced showed 603bp open reading frame encoding 200 amino-acids. The protein sequence had a molecular mass of 22 kDa and was therefore, named Hsp22. BlastX analysis revealed that the gene codes for a protein homologous to previously characterized Hsp22.4 gene from C. globosum (AAR36902.1, XP 001229241.1) and shared 95% identity in amino acid sequence. It also showed varying degree of similarities with small Hsp protein from Neurospora spp. (60%), Myceliophthora sp. (59%), Glomerella sp. (50%), Hypocrea sp. (52%), and Fusarium spp. (51%). This gene was further cloned into pET28a (+) and transformed E. coli BL21 cells were induced by IPTG, and the expressed protein of 30 kDa was analyzed by SDS-PAGE. The IPTG induced transformants displayed significantly greater resistance to NaCl and Na2CO3 stresses.

Corrigendum: Characterization of Fusarium Spp. Inciting Vascular Wilt of Tomato and Its Management by a Chaetomium-Based Biocontrol Consortium.

[This corrects the article DOI: 10.3389/fpls.2021.748013.].

Transcriptome Analysis of Wheat-Tilletia indica Interaction Provides Defense and Pathogenesis-Related Genes.

Karnal bunt (Tilletia indica Mitra) is an internationally quarantined disease of wheat. Until now, very little information has been available on the molecular basis of resistance and pathogenicity of T. indica. To investigate the molecular basis of host−pathogen interaction, the transcriptome of T. indica inoculated resistant (HD29) and susceptible (WH542) genotypes of wheat were analyzed. Approximately 58 million reads were generated using RNA sequencing by the Illumina NextSeq500 platform. These sequence reads were aligned to a reference genome of wheat to compare the expression level of genes in resistant and susceptible genotypes. The high-quality reads were deposited in the NCBI SRA database (SRP159223). More than 80,000 genes were expressed in both the resistant and susceptible wheat genotypes. Of these, 76,088 were commonly expressed genes, including 3184 significantly upregulated and 1778 downregulated genes. Four thousand one hundred thirteen and 5604 genes were exclusively expressed in susceptible and resistant genotypes, respectively. Based on the significance, 503 genes were upregulated and 387 genes were downregulated. Using gene ontology, the majority of coding sequences were associated with response to stimuli, stress, carbohydrate metabolism, developmental process, and catalytic activity. Highly differentially expressed genes (integral component of membrane, exonuclease activity, nucleic acid binding, DNA binding, metal ion binding) were validated in resistant and susceptible genotypes using qPCR analysis and similar expression levels were found in RNA-Seq. Apart from the wheat, the mapping of T. indica was 7.07% and 7.63% of resistant and susceptible hosts, respectively, upon infection, which revealed significant pathogenesis-related genes. This first study provided in-depth information and new insights into wheat−T. indica interaction for managing Karnal bunt disease of wheat.

Genome-Wide Identification and Characterisation of Cytokinin-O-Glucosyltransferase (CGT) Genes of Rice Specific to Potential Pathogens.

Cytokinin glucosyltransferases (CGTs) are key enzymes of plants for regulating the level and function of cytokinins. In a genomic identification of rice CGTs, 41 genes with the plant secondary product glycosyltransferases (PSPG) motif of 44-amino-acid consensus sequence characteristic of plant uridine diphosphate (UDP)-glycosyltransferases (UGTs) were identified. In-silico physicochemical characterisation revealed that, though the CGTs belong to the same subfamily, they display varying molecular weights, ranging from 19.6 kDa to 59.7 kDa. The proteins were primarily acidic (87.8%) and hydrophilic (58.6%) and were observed to be distributed in the plastids (16), plasma membrane (13), mitochondria (5), and cytosol (4). Phylogenetic analysis of the CGTs revealed that their evolutionary relatedness ranged from 70-100%, and they aligned themselves into two major clusters. In a comprehensive analysis of the available transcriptomics data of rice samples representing different growth stages only the CGT, Os04g25440.1 was significantly expressed at the vegetative stage, whereas 16 other genes were highly expressed only at the reproductive growth stage. On the contrary, six genes, LOC_Os07g30610.1, LOC_Os04g25440.1, LOC_Os07g30620.1, LOC_Os04g25490.1, LOC_Os04g37820.1, and LOC_Os04g25800.1, were significantly upregulated in rice plants inoculated with Rhizoctonia solani (RS), Xoo (Xanthomonas oryzae pv. oryzae) and Mor (Magnaporthe oryzae). In a qRT-PCR analysis of rice sheath tissue susceptible to Rhizoctonia solani, Mor, and Xoo pathogens, compared to the sterile distilled water control, at 24 h post-infection only two genes displayed significant upregulation in response to all the three pathogens: LOC_Os07g30620.1 and LOC_Os04g25820.1. On the other hand, the expression of genes LOC_Os07g30610.1, LOC_Os04g25440, LOC_Os04g25490, and LOC_Os04g25800 were observed to be pathogen-specific. These genes were identified as the candidate-responsive CGT genes and could serve as potential susceptibility genes for facilitating pathogen infection.

Whole-genome sequence analysis of Bipolaris sorokiniana infecting wheat in India and characterization of ToxA gene in different isolates as pathogenicity determinants.

Spot blotch disease of wheat caused by Bipolaris sorokiniana Boerma (Sacc.) is an emerging problem in South Asian countries. Whole genome of a highly virulent isolate of B. sorokiniana BS112 (BHU, Uttar Pradesh; Accession no. GCA_004329375.1) was sequenced using a hybrid assembly approach. Secreted proteins, virulence gene(s), pathogenicity-related gene(s) were identified and the role of ToxA gene present in this genome, in the development of disease was recognized. ToxA gene (535 bp) was analyzed and identified in the genome of B. sorokiniana (BS112) which revealed 100% homology with the ToxA gene of Pyrenophora tritici repentis (Accession no. MH017419). Furthermore, ToxA gene was amplified, sequenced and validated in 39 isolates of B. sorokiniana which confirmed the presence of ToxA gene in all the isolates taken for this study. All ToxA sequences were submitted in NCBI database (MN601358-MN601396). As ToxA gene interacts with Tsn1 gene of host, 13 wheat genotypes were evaluated out of which 5 genotypes (38.4%) were found to be Tsn1 positive with more severe necrotic lesions compared to Tsn1-negative wheat genotypes. In vitro expression analysis of ToxA gene in the pathogen B. sorokiniana using qPCR revealed maximum upregulation (14.67 fold) at 1st day after inoculation (DAI) in the medium. Furthermore, in planta expression analysis of ToxA gene in Tsn1-positive and Tsn1-negative genotypes, revealed maximum expression (7.89-fold) in Tsn1-positive genotype, Agra local at 5th DAI compared to Tsn1-negative genotype Chiriya 7 showing minimum expression (0.048-fold) at 5th DAI. In planta ToxA-Tsn1 interaction studies suggested that spot blotch disease is more severe in Tsn1-positive genotypes, which will be helpful in better understanding and management of spot blotch disease of wheat. Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-022-03213-3.