In five resistant CYP51A mutants, a single nucleotide substitution, I463V, was observed. The homologous I463V mutation, surprisingly, has not been found in other plant pathogens. Resistant mutants, when exposed to difenoconazole, showed a subtle increase in CYP51A and CYP51B expression levels compared to the wild-type strains; however, this elevation was not evident in the CtR61-2-3f and CtR61-2-4a mutants. Low resistance to difenoconazole in *C. truncatum* could potentially be associated with the emergence of the I463V point mutation in the CYP51A gene. The effectiveness of difenoconazole, tested in a greenhouse assay, increased with escalating doses, impacting both parental isolates and their mutant counterparts. genetic mapping The low to moderate resistance of *C. truncatum* to difenoconazole allows for its continued and responsible use in controlling soybean anthracnose.
The cultivar, Vitis vinifera cv. BRS Vitoria, a seedless black table grape, boasts a remarkably enjoyable flavor, readily cultivating throughout Brazil's diverse regions. In Petrolina, Pernambuco, Brazil, three vineyards observed grape berries displaying typical ripe rot symptoms throughout the period of November and December 2021. Ripe berries display initial symptoms as small, depressed lesions, showcasing tiny black acervuli. Lesions, expanding as the disease progresses, cover the entire fruit, displaying abundant orange conidia masses. Eventually, the berries are entirely transformed into mummies. Disease incidence, exceeding 90%, was observed alongside symptoms in the three inspected vineyards. Producers are contemplating eliminating their plantations, a drastic measure triggered by losses from the disease. Unfortunately, the current control methods are not only costly but also demonstrably ineffective. To isolate fungi, conidial masses were meticulously transferred from 10 diseased fruits to plates containing potato dextrose agar medium. joint genetic evaluation Cultures were incubated in an environment of continuous light and 25 degrees Celsius. Following inoculation for seven days, three fungal isolates (LM1543-1545) were harvested and cultured separately for species identification and subsequent pathogenicity assessments. The isolates presented cottony mycelial growth, ranging in color from white to gray, and hyaline conidia, cylindrical in form with rounded extremities, consistent with the characteristics of the Colletotrichum genus as described in Sutton (1980). The partial APN2-MAT/IGS, CAL, and GAPDH gene sequences were amplified, sequenced, and archived in GenBank (accession numbers OP643865-OP643872). Isolates from V. vinifera were positioned, within the clade, along with the ex-type and representative isolates from the C. siamense species. The maximum likelihood multilocus tree generated from the three combined loci exhibited substantial support (998% bootstrap support) for the clade, thus providing a certain and confident assignment of the isolates to the specified species. check details To establish the pathogen's capability to cause disease, grape bunches were inoculated. The grape bunches were sterilized on their surface by first soaking them in 70% ethanol for 30 seconds, then in 15% NaOCl for a minute, rinsing twice with sterile distilled water, and finally allowing them to air dry. Spraying fungal conidial suspensions (106 conidia per milliliter) was performed until complete run-off. A negative control was established by spraying grape bunches with sterile distilled water. Within a humid chamber, grapes' bunches were held at a temperature of 25 degrees Celsius, experiencing a 12-hour photoperiod for 48 hours. A single repetition of the experiment involved four replicates, each consisting of four inoculated bunches per isolate. The grape berries showed evidence of ripe rot, a typical symptom appearing seven days after the inoculation process. The negative control exhibited no observable symptoms. Identical to the C. siamense isolates from symptomatic field berries, the fungal isolates recovered from the inoculated berries displayed identical morphology, demonstrating compliance with Koch's postulates. In the United States, grape leaves were found to be associated with Colletotrichum siamense, as reported by Weir et al. (2012). Furthermore, this fungus was implicated in causing grape ripe rot across North America, as detailed by Cosseboom and Hu (2022). According to Echeverrigaray et al. (2020), C. fructicola, C. kahawae, C. karsti, C. limetticola, C. nymphaeae, and C. viniferum were the sole reported agents causing grape ripe rot in Brazil. In our records, this represents the first documented case of C. siamense being responsible for grape ripe rot in Brazil. Because C. siamense possesses a broad host range and is widely distributed, its considerable phytopathogenic potential necessitates the importance of this finding for disease management.
As a traditional fruit from Southern China, plum (Prunus salicina L.) is encountered globally. Water-soaked spots and light yellow-green halos affected more than 50% of plum tree leaves in the Babu district of Hezhou, Guangxi (N 23°49' to 24°48', E 111°12' to 112°03') in August 2021. For isolating the causal agent, three diseased leaves, procured from three different orchards, were sectioned into 5 mm x 5 mm pieces. These pieces were disinfected, first by immersing them in 75% ethanol for 10 seconds, then submerging them in 2% sodium hypochlorite for one minute, and subsequently rinsed three times in sterile water. To grind the diseased sections, sterile water was used, and subsequently they were held static for approximately ten minutes. Ten-fold serial dilutions in water were produced, and 100 liters of each dilution, ranging from 10⁻¹ to 10⁻⁶, were then plated onto Luria-Bertani (LB) Agar. Following incubation at 28 degrees Celsius for 48 hours, a 73% similarity in the morphology of isolates was observed. Three isolates, designated as GY11-1, GY12-1, and GY15-1, were selected for more extensive research. Round, opaque, and convex colonies were yellow, rod-shaped, non-spore-forming, featuring smooth, bright, and precisely delineated edges. From the results of biochemical tests, the colonies are known to require oxygen for growth and to have a gram-negative staining reaction. Isolates could thrive on LB agar containing 0-2% (w/v) NaCl, demonstrating the capacity to utilize glucose, lactose, galactose, mannose, sucrose, maltose, and rhamnose as their carbon source. Their response to H2S production, oxidase, catalase, and gelatin was positive, but starch evoked a negative reaction. The process of amplifying the 16S rDNA from the genomic DNA of the three isolates involved the utilization of primers 27F and 1492R. The amplified DNA fragments, known as amplicons, were sequenced. Furthermore, five housekeeping genes, atpD, dnaK, gap, recA, and rpoB, from the three isolates, were amplified using their respective primer pairs and sequenced. Within GenBank, the sequences were cataloged: 16S rDNA (OP861004-OP861006); atpD (OQ703328-OQ703330); dnaK (OQ703331-OQ703333); gap (OQ703334-OQ703336); recA (OQ703337-OQ703339); and rpoB (OQ703340-OQ703342). The isolates were determined to be Sphingomonas spermidinifaciens through phylogenetic analysis of the concatenated six sequences (multilocus sequence analysis, MLSA) using MegaX 70's maximum-likelihood method, following comparison against sequences from various Sphingomonas type strains. In a greenhouse setting, healthy leaves harvested from two-year-old plum plants were employed to assess the pathogenicity of the isolates. Sterile needles were used to pierce the leaves, after which, bacterial suspensions, prepared in phosphate buffer saline (PBS) at an optical density of 0.05 at 600 nm, were applied to the wounds. A negative control, PBS buffer solution, was employed in the experiment. For each isolate, 20 leaves per plum tree were subjected to inoculation. Plastic bags, strategically placed over the plants, maintained the high humidity. Incubation at 28 degrees Celsius under continuous light resulted in the appearance of dark brown to black lesions on the leaves 3 days later. The average diameter of lesions reached 1 cm after seven days; the negative controls, however, remained free of symptoms. The bacteria re-isolated from the diseased leaves, upon morphological and molecular analysis, proved to be identical to the inoculation bacteria, in accordance with Koch's postulates. A Sphingomonas species-induced plant disease has been documented in mango, pomelo, and Spanish melon. In China, this is the inaugural report detailing S. spermidinifaciens's association with plum leaf spot disease. Future development of effective disease control methodologies is significantly aided by this report.
Tianqi and Sanqi, also known as Panax notoginseng, are among the world's most prized medicinal perennial herbs (Wang et al., 2016). In the Lincang sanqi base (23°43'10″N, 100°7'32″E), covering 1333 hectares, leaf spot was observed on P. notoginseng leaves in the month of August 2021. The initial manifestation of the disease on leaves, as water-soaked areas, progressed to irregular, round or oval leaf spots. These spots presented transparent or grayish-brown centers containing black, granular material, with an observed incidence of 10% to 20%. A causal agent was sought by selecting ten symptomatic leaves from each of ten P. notoginseng plants, at random. Leaves exhibiting symptoms were meticulously dissected into small squares (5 mm2), ensuring asymptomatic tissue boundaries were preserved. The pieces were disinfected in 75% ethanol for 30 seconds, followed by a 3-minute immersion in 2% sodium hypochlorite, and finally rinsed three times with sterile distilled water. Tissue portions were set upon PDA plates and placed in an incubator at 20°C, maintaining a 12-hour light/dark cycle. From a top view, seven pure isolates showed a dark gray coloration, matching their taupe coloration when examined from the rear, and uniformly displaying flat and villous surfaces, with similar colony morphologies. Glabrous or sparsely mycelial pycnidia, ranging in form from globose to subglobose and in color from dark brown to black, showed sizes between 2246 and 15594 (average) microns. A recurring value of 'm' within the period 1305 to 1820 had an average of 6957.