Defense Pathways of Wheat Plants Inoculated with Zymoseptoria tritici under NaCl Stress Conditions: An Overview.

Due to being sessile, plants develop a broad range of defense pathways when they face abiotic or biotic stress factors. Although plants are subjected to more than one type of stress at a time in nature, the combined effects of either multiple stresses of one kind (abiotic or biotic) or more kinds (abiotic and biotic) have now been realized in agricultural lands due to increases in global warming and environmental pollution, along with population increases. Soil-borne pathogens, or pathogens infecting aerial parts, can have devastating effects on plants when combined with other stressors. Obtaining yields or crops from sensitive or moderately resistant plants could be impossible, and it could be very difficult from resistant plants. The mechanisms of combined stress in many plants have previously been studied and elucidated. Recent studies proposed new defense pathways and mechanisms through signaling cascades. In light of these mechanisms, it is now time to develop appropriate strategies for crop protection under multiple stress conditions. This may involve using disease-resistant or stress-tolerant plant varieties, implementing proper irrigation and drainage practices, and improving soil quality. However, generation of both stress-tolerant and disease-resistant crop plants is of crucial importance. The establishment of a database and understanding of the defense mechanisms under combined stress conditions would be meaningful for the development of resistant and tolerant plants. It is clear that leaf pathogens show great tolerance to salinity stress and result in pathogenicity in crop plants. We noticed that regulation of the stomata through biochemical applications and some effort with the upregulation of the minor gene expressions indirectly involved with the defense mechanisms could be a great way to increase the defense metabolites without interfering with quality parameters. In this review, we selected wheat as a model plant and Zymoseptoria tritici as a model leaf pathogen to evaluate the defense mechanisms under saline conditions through physiological, biochemical, and molecular pathways and suggested various ways to generate tolerant and resistant cereal plants.

Unveiling the Genetic Tapestry: Exploring Rhizoctonia solani AG-3 Anastomosis Groups in Potato Crops across Borders.

The current study was carried out to screen 10 isolates (ARS-01-ARS-10) of Rhizoctonia. solani from potato tubers cv. Kuroda, which were collected from various potato fields in Multan, Pakistan. The isolates were found to be morphologically identical, as the hyphae exhibit the production of branches at right angles and acute angles often accompanied by septum near the emerging branches. Anastomosis grouping showed that these isolates belonged to AG-3. A pathogenicity test was performed against the susceptible Kuroda variety and among the isolates, ARS-05 exhibited the highest mean severity score of approximately 5.43, followed by ARS-09, which showed a mean severity score of about 3.67, indicating a moderate level of severity. On the lower end of the severity scale, isolates ARS-06 and ARS-07 displayed mean severity scores of approximately 0.53 and 0.57, respectively, suggesting minimal symptom severity. These mean severity scores offer insights into the varying degrees of symptom expression among the different isolates of R. solani under examination. PCoA indicates that the severe isolate causing black scurf on the Kuroda variety was AG-3. A comprehensive analysis of the distribution, genetic variability, and phylogenetic relationships of R. solani anastomosis groups (AGs) related to potato crops across diverse geographic regions was also performed to examine AG prevalence in various countries. AG-3 was identified as the most widespread group, prevalent in Sweden, China, and the USA. AG-5 showed prominence in Sweden and the USA, while AG-2-1 exhibited prevalence in China and Japan. The phylogenetic analysis unveiled two different clades: Clade I comprising AG-3 and Clade II encompassing AG-2, AG-4, and AG-5, further subdivided into three subclades. Although AGs clustered together regardless of origin, their genetic diversity revealed complex evolutionary patterns. The findings pave the way for region-specific disease management strategies to combat R. solani's impact on potato crops.

Nitrogen sources alter ligninase and cellulase activities of thermophilic fungi isolated from compost and vermicompost.

Fungi harboring lignocellulolytic activity accelerate the composting process of agricultural wastes; however, using thermophilic fungal isolates for this process has been paid little attention. Moreover, exogenous nitrogen sources may differently affect fungal lignocellulolytic activity. A total of 250 thermophilic fungi were isolated from local compost and vermicompost samples. First, the isolates were qualitative assayed for ligninase and cellulase activities using Congo red (CR) and carboxymethyl cellulose (CMC) as substrates, respectively. Then, twenty superior isolates harboring higher ligninase and cellulase activities were selected and quantitatively assayed for both enzymes in basic mineral (BM) liquid medium supplemented with the relevant substrates and nitrogen sources including (NH4)2SO4 (AS), NH4NO3 (AN), urea (U), AS + U (1:1), or AN + U (1:1) with final nitrogen concentration of 0.3 g/L. The highest ligninase activities of 99.94, 89.82, 95.42, 96.25, and 98.34% of CR decolorization were recorded in isolates VC85, VC94, VC85, C145, and VC85 in the presence of AS, U, AS + U, AN, and AN + U, respectively. Mean ligninase activity of 63.75% in superior isolates was achieved in the presence of AS and ranked the highest among other N compounds. The isolates C200 and C184 exhibited the highest cellulolytic activity in the presence of AS and AN + U by 8.8 and 6.5 U/ml, respectively. Mean cellulase activity of 3.90 U/mL was achieved in AN + U and ranked the highest among other N compounds. Molecular identification of twenty superior isolates confirmed that all of them are belonging to Aspergillus fumigatus group. Focusing on the highest ligninase activity of the isolate VC85 in the presence of AS, the combination can be recommended as a potential bio-accelerator for compost production.

Assessment of Genetic Variability and Evolutionary Relationships of Rhizoctonia solani Inherent in Legume Crops.

Rhizoctonia solani is one of the most common soil-borne fungal pathogens of legume crops worldwide. We collected rDNA-ITS sequences from NCBI GenBank, and the aim of this study was to examine the genetic diversity and phylogenetic relationships of various R. solani anastomosis groups (AGs) that are commonly associated with grain legumes (such as soybean, common bean, pea, peanut, cowpea, and chickpea) and forage legumes (including alfalfa and clover). Soybean is recognized as a host for multiple AGs, with AG-1 and AG-2 being extensively investigated. This is evidenced by the higher representation of sequences associated with these AGs in the NCBI GenBank. Other AGs documented in soybean include AG-4, AG-7, AG-11, AG-5, AG-6, and AG-9. Moreover, AG-4 has been extensively studied concerning its occurrence in chickpea, pea, peanut, and alfalfa. Research on the common bean has been primarily focused on AG-2, AG-4, and AG-1. Similarly, AG-1 has been the subject of extensive investigation in clover and cowpea. Collectively, AG-1, AG-2, and AG-4 have consistently been identified and studied across these diverse legume crops. The phylogenetic analysis of R. solani isolates across different legumes indicates that the distinct clades or subclades formed by the isolates correspond to their specific anastomosis groups (AGs) and subgroups, rather than being determined by their host legume crop. Additionally, there is a high degree of sequence similarity among isolates within the same clade or subclade. Principal coordinate analysis (PCoA) further supports this finding, as isolates belonging to the same AGs and/or subgroups cluster together, irrespective of their host legume. Therefore, the observed clustering of R. solani AGs and subgroups without a direct association with the host legume crop provides additional support for the concept of AGs in understanding the genetic relationships and evolution of R. solani.

Inactivation of a candidate effector gene of Zymoseptoria tritici affects its sporulation.

BACKGROUND: Wheat is one of the most important staple crops produced worldwide. Its susceptibility to plant diseases reduces its production significantly. One of the most important diseases of wheat is septoria tritici blotch, a devastating disease observed in fields with wet and temperate conditions. Z. tritici secretes effector proteins to influence the host's defense mechanisms, as is typical of plant pathogens. In this investigation, we evaluated the pathogenicity of some Zymoseptoria tritici effector candidate genes having a signal peptide for secretion with no known function. METHODS AND RESULTS: Three genes named Mycgr3G104383, Mycgr3G104444 and Mycgr3G105826 were knocked out separately through homologous recombination, generating Z. tritici IPO323 mutants lacking the functional copy of the corresponding genes. While KO1 and KO3 mutants did not show any significant differences during phenotypic and virulence investigations, the KO2 mutant generated exclusively macropycnidiospores in artificial media, different from wild-type IPO323 which produce only micropycidiospores. The mycelial growth capability of KO2 was also severely attenuated in all of the investigated growth conditions. These changes were observed independent of growth media and growth temperatures, implying that changes were genetic and inherited through generations. Virulence of knockout mutants in wheat leaves was observed to be similar to the wild-type IPO323. CONCLUSION: Understanding the biology of Z. tritici and its interactions with wheat will reveal new strategies to fight septoria tritici blotch, enabling breeding wheat cultivars resistant to a broader spectrum of Z. tritici strains. Furthermore, gene knockout via homologous recombination proved to be a powerful tool for discovering novel gene functions.

Identification and primary distribution of Citrus viroid V in citrus in Punjab, Pakistan.

BACKGROUND: Citrus plants are prone to infection by different viroids which deteriorate their vigor and production. Citrus viroid V (CVd-V) is among the six citrus viroids, belongs to genus Apscaviroid (family Pospiviroidae) which induces symptoms of mild necrotic lesions on branches and cracks on trunk portion. METHODS AND RESULTS: A survey was conducted to evaluate the prevalence of CVd-V in core and non-core citrus cultivated areas of Punjab, Pakistan. A total of 154 samples from different citrus cultivars were tested for CVd-V infection by RT-PCR. The results revealed 66.66% disease incidence of CVd-V. Citrus cultivars Palestinia Sweet lime, Roy Ruby, Olinda Valencia, Kaghzi lime, and Dancy were identified as new citrus hosts of CVd-V for the first time from Pakistan. The viroid infection was confirmed by biological indexing on indicator host Etrog citron. The reported primers used for the detection of CVd-V did not amplify, rather showed non-specific amplification, which led to the designing of new primers. Whereas, new back-to-back designed primers (CVd-V AF1/CVd-V AR1) detected CVd-V successfully and obtained an expected amplified product of CVd-V with 294 bp. Sequencing analysis confirmed the new host of CVd-V showing 98-100% nucleotide sequence homology with those reported previously from other countries while 100% sequence homology to the isolates reported from Pakistan. Based on phylogenetic analysis using all CVd-V sequences in GenBank, two main CVd-V groups (I and II) were identified, and newly identified isolates during this study fall in the group I. CONCLUSION: The study revealed that there are some changes in the nucleotide sequences of CVd-V which made difficult for their detection using reported primers. All isolates of Pakistan showed high sequence homology with other isolates of CVd-V from Iran and USA whereas; the isolates from China, Japan, Tunisia, and Africa are distantly related. It is evident that CVd-V is spreading in all citrus cultivars in Pakistan.

Prevalence, transmission and molecular characterization of Cotton leaf curl Multan virus infecting hollyhock plants in Pakistan.

BACKGROUND: Hollyhock (Alcea rosea) is an ornamental plant belonging to the Malvaceae family and has a remarkable aesthetic and medicinal value. A number of distinct infectious entity including fungi, nematode, bacteria and most importantly both single and double stranded DNA and RNA viruses are reported from infected hollyhock plant. Begomoviruses, the well reputed member of the family Geminiviridae infected the hollyhock recently with a new hollyhock vein yellowing virus and in the present study it infected the hollyhock plant with Cotton leaf curl Multan virus (CLCuMV) which cause the disease of leaf curling. METHODS AND RESULTS: The symptomatic leaves of the hollyhock plants were collected based on the characteristic symptoms of leaf curling, puckering as well as vein thickening. DNA was extracted by using the recommended 2× CTAB protocol and PCR technique was optimized for the detection of begomovirus followed by sequencing. The data of disease incidence of infection location wise was collected based on the positive results of PCR amplification. Virus free whitefly collected from cotton field and feed on infected hollyhock plant in cage for few days then used for the transmission study of begomovirus on healthy hollyhock plants. Results of PCR amplification indicated that the primers Av/Ac core, Begomo 01/02, and CLCV 01/02 showed the bands of 579 bp, 2.8 kb and 1.1 kb respectively. The betasatellite was amplified by using beta01/02 and CLCuMuBF11/R33, which showed the band of 1400 bp and 481 bp. Disease incidence and Transmission study confirmed the begomovirus in hollyhock plants at molecular level. The sequence obtained with Av/Ac core primers showed the 99% identity with Cotton leaf curl Multan virus-Rajasthan strain and betasatellite primers showed 98% identity with Cotton leaf curl Multan betasatellite. CONCLUSION: Hollyhock plants infected by CLCuMV and associated betasatellite has been reported as a possible source of virus inoculum from Pakistan. These findings extend the range of Begomoviruses and betasatellites known to infect A. rosea and highlight this hollyhock species as an important reservoir of agriculturally important Begomoviruses and betasatellites.

Molecular characterization of divergent isolates of Citrus bent leaf viroid (CBLVd) from citrus cultivars of Punjab, Pakistan.

Citrus viroid infection is emerging as a serious threat because of its efficient systemic movement within the host plant and its quick spread due to contaminated pruning tools. A survey was conducted to investigate the primary distribution and molecular characterization of Citrus bent leaf viroid (CBLVd) and its variants in different citrus cultivars. A total of 154 symptomatic citrus samples were collected and detected by RT‒PCR with newly designed specific primers with the incidence of 36.33%. During biological indexing study on Etrog citron, expressions of reduced leaf size, yellowing with a light green pattern, and bending were observed. Amplified products were sequenced and analyzed using a nucleotide BLAST search, which showed 98% homology with other CBLVd isolates. The results of the phylogenetic tree analysis showed the presence of two main groups (A and B), with the predominant variants of CBLVd, i.e., CVd-I-LSS (Citrus viroid Low Sequence Similarity) sequences, clustering in subgroup A1 along with newly detected CVd-I-LSS from Palestinian sweet lime (Citrus limettioides), which has been identified as a new host of CVd-I-LSS in Pakistan. Further analysis of the sequences in subgroup A1 showed that the variant of CVd-I-LSS infecting citrus cultivars had a close relationship with isolates reported from China, Japan, and Iran, which may have resulted from the exchange of planting material. This study also unveiled the variability in nucleotide sequences of CBLVd, which made it unable to be detected by old primers. The results of this study indicate that the widespread presence of divergent variants of CBLVd is a major concern for the citrus industry in Pakistan and other countries where virulent isolates of CBLVd are prevalent. These findings suggest the need for future research on effective management and quarantine measures to stop the spread of CBLVd.

Remarkable recent changes in the genetic diversity of the avirulence gene AvrStb6 in global populations of the wheat pathogen Zymoseptoria tritici.

Septoria tritici blotch (STB), caused by the fungus Zymoseptoria tritici, is one of the most economically important diseases of wheat. Recently, both factors of a gene-for-gene interaction between Z. tritici and wheat, the wheat receptor-like kinase Stb6 and the Z. tritici secreted effector protein AvrStb6, have been identified. Previous analyses revealed a high diversity of AvrStb6 haplotypes present in earlier Z. tritici isolate collections, with up to c.18% of analysed isolates possessing the avirulence isoform of AvrStb6 identical to that originally identified in the reference isolate IPO323. With Stb6 present in many commercial wheat cultivars globally, we aimed to assess potential changes in AvrStb6 genetic diversity and the incidence of haplotypes allowing evasion of Stb6-mediated resistance in more recent Z. tritici populations. Here we show, using targeted resequencing of AvrStb6, that this gene is universally present in field isolates sampled from major wheat-growing regions of the world in 2013-2017. However, in contrast to the data from previous AvrStb6 population studies, we report a complete absence of the originally described avirulence isoform of AvrStb6 amongst modern Z. tritici isolates. Moreover, a remarkably small number of haplotypes, each encoding AvrStb6 protein isoforms conditioning virulence on Stb6-containing wheat, were found to predominate among modern Z. tritici isolates. A single virulence isoform of AvrStb6 was found to be particularly abundant throughout the global population. These findings indicate that, despite the ability of Z. tritici to sexually reproduce on resistant hosts, AvrStb6 avirulence haplotypes tend to be eliminated in subsequent populations.

Comparison of three different treatment modalities in the management of cancer cachexia.

AIMS AND BACKGROUND: The optimal treatment of cancer cachexia remains unknown. In this study, we compared the efficacy of three different treatment modalities in the management of cancer cachexia. METHODS: Sixty-two assessable cachectic cancer patients were randomized to one of the following three arms: 1) megesterol acetate (MA) plus meloxicam (n = 23); 2) MA plus meloxicam plus oral eicosapentaenoic acid (EPA)-enriched nutritional supplement (n = 21); or 3) meloxicam plus oral EPA-enriched nutritional supplement (n = 18). Treatment duration was 3 months. RESULTS: The treatment arms were well balanced at baseline. The primary efficacy (body weight and lean body mass) and secondary efficacy (body mass index, quality of life, and serum levels of IL-6 and TNF-α) parameters improved after treatment in all three arms. There were no statistically significant differences between treatment groups in the mean percentage changes in all efficacy parameters from baseline to end of study. CONCLUSIONS: MA plus meloxicam or EPA supplement plus meloxicam may be effective treatment options in the management of cancer cachexia. The combined use of these agents does not provide further advantages.