Rot Symptoms Research Articles

Disease prevalence, incidence, morphological and molecular characterisation of Lasiodiplodia pseudotheobromae causing collar rot disease on peanut plants in Turkey

AbstractPeanut (Arachis hypogaea L.) holds significant commercial and dietary importance as a major source of edible oil and protein in Turkey. Stem, collar or root rot, caused by several fungal disease agent, are serious soil-borne diseases of peanut. Accurate and precise identification of the disease agent provides fundamental and precise information for integrated plant management. During the period from June to September 2021, symptoms consistent with collar rot disease, including dark-brown stem rot, chlorotic leaves, wilting, and eventual whole plant death, were observed on peanut plants cultivated in the different districts of Osmaniye Province of Turkey. The disease incidence ranged from 8.0 to 45.0% in the inspected fields with an average of 3.4% overall. Twenty-four single-spore representative isolates were obtained from surface-disinfected symptomatic tissues. Morphological characteristics of fungal mycelium, conidial and pycnidial structures on potato sucrose agar (PSA) and water agar (WA) closely resembled those described for Lasiodiplodia spp. All isolates caused typical collar rot symptoms upon artificial inoculation of peanut seedlings. Morphological identification of Lasiodiplodia spp. isolates was corroborated by MALDI-TOF and molecular analyses utilizing sequences from the internal transcribed spacer (ITS), β-tubulin 2 (tub2) and translation elongation factor-1 alpha (TEF1-α) loci. Phylogenetic analysis confirmed that the representative fungal isolates (MKUBK-B1 and MKUBK-K22) belong to Lasiodiplodia pseudotheobromae. To the best of our knowledge, this is the first report of L. pseudotheobromae infecting peanut plants in Turkey. This work is expected to contribute to previously limited knowledge regarding the host range, incidence and prevalence of L. pseudotheobromae as a soilborne pathogen of peanuts. Due to the potential destructiveness and broad host range of this pathogen, it is essential to develop new strategies to establish more reliable, environmentally sustainable, and cost-effective management approaches for this disease.

First Report of Alternaria gossypina Causing Stem Base Rot on Passilora edulis in Guangxi, China.

Passion fruit (Passilora edulis), known as the "king of fruit juices", is popular in southern China (Yuan et al. 2019). Stem base rot is a devastating disease of passion fruit commonly caused by several Fusarium spp. (Zakaria, 2022). In July 2022, typical symptoms of stem base rot were observed in a poorly managed "Qinmi No. 9" Golden passion fruit orchard in Jingxi (23°13'10"N, 106°5'23"E). The disease incidence had reached 40% (n=200) in the survey. Symptoms included ulceration and mutilation at the stem base, making the plants prone to breakage when pulled, wilting and drooping leaves above ground, and severe cases leading to the entire plant withering and dying. Fourteen plants with obvious symptoms were collected. Thin sections of plant tissue were cut from junction of sickness and health of stem, sterilized with ethanol and sodium hypochlorite, and placed on PDA medium at 28°C. Sixty fungal strains were obtained, 90% of which was identified as Fusarium based on morphology. 80% of Fusarium were F. oxysporum species complex (FOSC), but pathogenicity experiment showed all FOSC were weakly pathogenic. However, two severely pathogenic fungi with similar morphology but distinct from Fusarium were identified from the same plant. The representative strain C11 was selected for further study. C11 demonstrated a rapid growth rate, reaching a 90 mm diameter colony on PDA cultured at 25°C for 7 days. The colony displaced a round, flat shape with an overall light brown front, and cottony gray or light brown mycelium, while the reverse side was dark brown. Conidia production was observed as typically occurred in multiple chains after 14 days culture on OA medium, with round, oval or straight rod-like brown conidia ranging in size from 5.74-23.42×14.67-67.22, featuring 1-8 transverse septa and 0-3 mediastinum (Figure S1). For molecular identification, the internal transcribed spacer (ITS, OR616614), translation elongation factor 1-alpha (TEF, OR633298), alternaria major allergen (Alt a1, OR633294) gene, glyceraldehyde 3-phosphate dehydrogenase (GAPDH, OR633295), RNA polymerase subunit II gene (RPB2, OR633297), 18S Small subunit rDNA (SSU, OR616608) , 28S Large Subunit rDNA (LSU, OR616615), the KOG1058 gene regions (OR633296) and an approximately segment of the anonymous noncoding region (OPA10-2, OR633299) were amplified from C11 (Liu et al. 1999, Li et al. 2023, Andrew et al. 2009), and deposited in GenBank with accession number shown in the brackets. Phylogenetic trees were constructed in MEGA11 after splicing by BioEdit (Figure S3). Combining morphology and molecular analyses, C11 was identified as Alternaria gossypina (Woudenberg et al. 2015). To test the pathogenicity, the base of the seedling stem (20cm in height) of 50 healthy "Tainong No. 1" variety of purple passion fruit, which was more susceptible to stem-base rot, was puncture wound with needles, inoculated with 6 mm diameter colonies of fungi, and then wrapped in wet cotton (Ángel et al. 2018). Ten healthy seedlings inoculated with PDA were used as controls. These plants were cultured in an artificial greenhouse at 30±5℃with 80±5% humidity. After 15 days, the plants inoculated with C11 exhibited symptoms similar to those in the field, whereas the controls remained healthy. A. gossypina was reisolated from the diseased plant stems, with the morphology and GAPDH sequence consistent with the inoculated (Figure S1, S2). This is the first report of passion fruit stem rot caused by A. gossypina. This finding will aid in the prevention and control of stem rot in passion fruit.

Cultural, Morphological and Pathogenic Variability in the Colletotrichum capsici Isolates, Inciting Anthracnose and Fruit rot of Chilli (Capsicum annuum L.) in different Agro-Climatic Zones of Kerala, India

Aims: To study the cultural, morphological and pathogenic variability existing among the pathogen isolates of Colletotrichum capsici, inciting anthracnose and fruit rot of chilli in Kerala. Study Design: Completely randomized design (CRD). Place and Duration of Study: Department of Plant Pathology, College of Agriculture (Kerala Agricultural University), Vellayani, Thiruvananthapuram during 2018-2019. Methodology: A survey was conducted in the five agro-climatic zones (ACZs) of Kerala viz., Northern zone, Central zone, Special problem zone, High range zone and Southern zone to assess the incidence and severity of anthracnose disease. The various symptoms in the surveyed locations were also noted. Pathogen isolates were grown in PDA medium in the laboratory and colony characters (colour and mycelial growth rate of pathogen colonies) were observed. Also, the morphological characters viz., mycelial width, shape and size of conidia, acervular diameter, number and size of setae, appressorial size were recorded. Results: Maximum anthracnose incidence of 90 per cent and severity of 52.60 per cent were noticed at Regional Agricultural Research Station (RARS), Pilicode (Northern zone). Characteristic leaf spot, fruit rot and die-back symptoms were observed in the survey locations. Pathogen colonies showed different shades of white, off-white to grey turning brown or black with regular or irregular margins and concentric rings of black acervuli in the potato dextrose agar (PDA) medium. The diameter of growth of C. capsici isolates ranged from 7.20 to 8.60 cm at 7 days after incubation. All the C. capsici isolates initiated hyaline mycelium having a width of 1.73 - 2.36 µm and produced numerous sickle shaped conidia with a size range of 19.42 - 20.46 µm × 2.16 - 3.09 µm. Black coloured, circular or ellipsoidal acervuli (122.14 - 189.08 µm dia.) had 20 – 46 setae of length 74.13 to 107.30 µm. Also, brown or black appressoria of size 8.64 - 12.64 µm × 5.54 - 7.84 µm were formed in slide culture. Cc3 from College of Agriculture, Vellanikkara was obtained as the most virulent isolate producing a lesion size of 1.13 cm and PDI of 45.33 in the artificially inoculated chilli fruits (var. Vellayani Athulya) at 5 days after inoculation. Conclusion: All the nine C. capsici isolates exhibited variability in their cultural, morphological and pathogenic characteristics. Variability in phytopathogenic fungi helps in identifying novel and sustainable control methods to tackle the diseases incited by them. Understanding pathogen variability provides new insights for developing effective disease management methods. This will help in promoting sustainable agriculture and preventing severe crop losses.

Role of Volatile Organic Compounds Produced by Kosakonia cowanii Cp1 during Competitive Colonization Interaction against Pectobacterium aroidearum SM2

The competitive colonization of bacteria on similar ecological niches has a significant impact during their establishment. The synthesis speeds of different chemical classes of molecules during early competitive colonization can reduce the number of competitors through metabolic effects. In this work, we demonstrate for the first time that Kosakonia cowanii Cp1 previously isolated from the seeds of Capsicum pubescens R. P. produced volatile organic compounds (VOCs) during competitive colonization against Pectobacterium aroidearum SM2, affecting soft rot symptoms in serrano chili (Capsicum annuum L.). The pathogen P. aroidearum SM2 was isolated from the fruits of C. annuum var. Serrano with soft rot symptoms. The genome of the SM2 strain carries a 5,037,920 bp chromosome with 51.46% G + C content and 4925 predicted protein-coding genes. It presents 12 genes encoding plant-cell-wall-degrading enzymes (PCDEWs), 139 genes involved in five types of secretion systems, and 16 genes related to invasion motility. Pathogenic essays showed soft rot symptoms in the fruits of C. annuum L., Solanum lycopersicum, and Physalis philadelphica and the tubers of Solanum tuberosum. During the growth phases of K. cowanii Cp1, a mix of VOCs was identified by means of HS-SPME-GC-MS. Of these compounds, 2,5-dimethyl-pyrazine showed bactericidal effects and synergy with acetoin during the competitive colonization of K. cowanii Cp1 to completely reduce soft rot symptoms. This work provides novel evidence grounding a better understanding of bacterial interactions during competitive colonization on plant tissue, where VOC synthesis is essential and has a high potential capacity to control pathogenic microorganisms in agricultural systems.

First Report of Enterobacter mori Causing Bacterial Wilt on Tomato in China.

Tomato (Solanum lycopersicum L.) is one of the most important vegetable crops in China. In October 2023, a new bacterial disease was discovered on tomato plants in a 0.3-acre farm's greenhouse (35.514806N, 118.996106E) in Longshan Town, Shandong Province, China. Over 50% of the tomato plants showed symptoms of stem rot, leaf wilt, or plant death. Three diseased tomato plants were collected for pathogen isolation and purification. Two leaf samples, each about 1 cm2, were cut from the junction area of healthy and diseased parts and disinfected with 75% alcohol for 60 s, followed by 0.1% HgCl2 for 90 s, and then washed three times with sterilized H2O. The samples were subsequently ground with 1.0 mL sterilized H2O. The ground samples were diluted to 10-4, 10-5, and 10-6 and then were plated on a potato dextrose agar (PDA) plate, respectively. White mucous bacterial colonies appeared at 28℃ for 24~48 h, no fungal colony was observed. Six bacterial colonies were randomly selected for gram staining and found to be gram-negative. To further determine their species classification, fragments of the 16SrDNA, hsp60, gyrB, and rpoB genes were separately amplified using previously reported PCR conditions and with primer pairs, including 27F/1492R (Wu et al., 2023), HSP60-F /HSP60-R (Gül et al., 2023), gyrB UP-1 / gyrB UP-2r (Yamamoto et al., 1995) and rpoB CM81-F / rpoB CM32b-R (Brady et al., 2008). Sequence analysis showed that the obtained sequences of the 16SrDNA, hsp60, gyrB, and rpoB genes among these six colonies were identical and 100%, 100%, 99.31%, and 99.36% similar to those of Enterobacter mori accessions OP601841 (with a coverage of 100%), MT199160 (83%), OP676246 (100%), and MN594495 (100%), at nucleotide level, respectively. Sequences of the above four genes of 23LSFQ were submitted to GenBank under the accession numbers PP461247, PP474090, PP136037, and PP136038, respectively. We selected one of these six colonies, 23LSFQ, for further analysis. The phylogenetic tree based on the concatenated sequences of the above four genes using the maximum likelihood method with MEGAX software showed that 23LSFQ is grouped with E. mori LMG25706 (NCBI: txid980518). To determine the pathogenicity of 23LSFQ , we sprayed 23LSFQ (OD600=0.8) onto five 30-day-old healthy plants of the tomato cultivars Alisa Craig, Jinpeng NO.1, and Chaobei, respectively. These seedlings were incubated in a chamber at 28°C with a 16 h light/ 8h dark photoperiod and 60% relative humidity. The leaves of the inoculated plants became curled and wilted at two days post inoculation (dpi) and appeared necrotic at 10 dpi. The symptoms were similar to those observed in field-infected tomato plants. No symptoms were observed on the plants inoculated with water. We further sequenced the re-isolated bacteria from the symptomatic inoculated seedlings. Results showed that they belong to E. mori. The experiment was repeated three times. E. mori has been found to cause diseases on peaches (Ahmad et al., 2021), watermelons (Wu et al., 2023), Canna indica, (Zhang et al., 2023), and strawberries (Ji et al., 2023). E. cloacae has been found to cause diseases on tomatoes in Heilongjiang province (Jin et al., 2023). This is the first report of E. mori causing leaf yellowing and wilting on tomatoes in China. These results are significant for the safe production and disease control of greenhouse tomatoes.

First Report of Kiwifruit Root Rot Caused by Globisporangium spinosum in China.

Kiwifruit is widely cultivated for its high vitamin C content and nutritional value. In January 2022, root rot symptoms were found in about 30% of Actinidia chinensis cv. Jinyan plants grafted on A. deliciosa rootstocks in an orchard located in Sanming (26.32°N, 117.23°E), Fujian Province of China. The affected plants appeared stunted, with brown and decaying roots, some of which were covered with white hyphae. To isolate the pathogen, the surfaces of typical symptomatic roots were sterilized for 30 s using 75% ethanol, followed by four rinses in sterile water, placing on potato dextrose agar (PDA), and incubating away from light at 25°C for 7 days. 16 Globisporangium-like isolates were obtained through hyphal tip isolation, displaying a milky-white appearance with irregular protuberances on the surface, and yellow-white backs with radial fold lines. The isolates were then cultured on corn meal agar for 5 days at 25°C in dark for morphological characteristics. Under microscope, the hyphae appeared as long strips without septa and 4.1 to 8.2 µm wide (average 6.7 µm), containing irregularly sized spherical droplets. Both terminal and intercalary hyphae swellings were observed; these appeared either spherical or subspherical, with some having projections. Their dimensions were 12.3 to 27.6 µm (average 17.3 µm). The oospores were mostly spherical, either plerotic or aplerotic, 11.8 to 22.3 µm wide (average 18.9 µm), with occasional projections. The antheridia were rod-shaped and curved, with one end attached to the oogonia. Amplification of the sequences of internal transcribed spacer (ITS) regions and cytochrome c oxidase subunit I (COI) were conducted using the primers ITS1/ITS4 (White et al. 1990) and OomCoxI-Levlo/OomCoxI-Levup (Robideau et al. 2011), respectively. The sequencing results revealed identical ITS and COI sequences in all 16 isolates. BLASTn analysis of the 969-bp ITS sequence ON202808 showed 99.38-99.59% similarity (965/971bp, 967/971bp) with the KJ162353 and AY598701 sequences from Globisporangium spinosum isolates, while the 700-bp COI sequence ON075783 showed 100% and 99.41% identity (680/680bp, 676/680bp) with the GenBank sequences HQ708835 and HQ708832, respectively, from G. spinosum. Phylogenetic analysis also showed that the obtained isolate (termed MA16) clustered with isolates from G. spinosum on the same evolutionary branch. For pathogenicity testing, four-month-old healthy Jinyan (A. chinensis) plants grown in sterilized media were transferred to sterile petri dishes covered with wet filter paper, and their roots were inoculated with a 5-mm-wide disk of MA16 when cultivated on PDA medium for 5 days. Miliang-1 (A. deliciosa) and Hongyang (A. chinensis) plants were treated similarly. The control groups each included three plants that were inoculated with non-colonized PDA. The plants were kept at 25 °C with a 12-/12-h light/dark cycle for 10 days when the inoculated plants exhibited root rot symptoms similar to those seen in the field, together with rotting and browning of the leaves. The control plants appeared healthy with no symptoms. After re-isolated from infected tissues, the pathogen was verified to be G. spinosum according to its ITS sequence, thus fulfilling the Koch's postulates. Recently, Pythium spinosum has been classified as G. spinosum according to whole-genome sequencing and phylogenomic analysis (Nguyen et al. 2022). Based on the morphological features and pathogenicity results, MA16 was identified as G. spinosum (van der Plaats-Niterink 1981; Huo et al. 2023). This report appears to be the first description of kiwifruit root rots caused by G. spinosum in China, and its identification will assist the development of strategies to counteract the disease.

First Report of Pectobacterium polaris Causing Soft Rot on Broccoli in China.

In April 2023, soft rot symptoms were observed in broccoli (Brassica oleracea L. var. italica) commercial fields in Songming County, Yunnan province, China (103°12'E, 25°31'N). The disease incidence in these fields (6 ha in size) was high, exceeding 50%, and it caused significant yield loss. The affected plants displayed characteristic symptoms, with the roots and stems of broccoli becoming soft, yellowish-brown, rotten, and emitting a foul odor. To identify the causal agent, soft rot symptomatic stems were surface sterilized by dipping them in 75% ethanol for 30 seconds, followed by three successive rinses with sterile distilled water. Tissue specimens were then plated onto nutrient agar (NA) plates and incubated at 28°C for 24 hours. (Wang et al. 2022). Three representative bacterial isolates HYC22041801-HYC22041803 from broccoli were selected for further analysis. The colonies on NA plates appeared as white, small, round, and translucent with smooth edges. Physiological and biochemical tests were performed, along with 96 phenotypic screenings using the BIOLOG GENIII microplate system (Biolog, Hayward, CA, USA). Three isolates were negative for D-arabitol, maltose, and sorbitol, but were positive for cellobiose, α-D-glucose, sucrose, glycerol and gentiobiose tests, which are consistent with the reported type strain P. polaris NIBIO1006T (Chen et al. 2021). Total genomic DNA was extracted from three bacterial isolates using the QIAamp DNA Mini Kit (QIAGEN, USA). The 16S rRNA region and nine housekeeping genes (gapA, icdA, mdh, mtlD, pel, pgi, pmrA, proA and rpoS) were amplified with universal primers 27F/1492R (Monciardini et al., 2006) and designed specific primers (Xie et al., 2018), respectively. All amplicons were sequenced and deposited in GenBank with accession numbers ON723841-ON723843 and ON723846-ON723872. The BLASTn analysis of the 16S rRNA amplicons confirmed that the isolates HYC22041801-HYC22041803 belonged to the genus Pectobacterium. Phylogenetic trees based on 16S rRNA gene sequences and multilocus sequence analysis of other nine housekeeping genes of the three isolates were constructed and the results revealed that three isolates clustered with P. polaris type strain NIBIO1006T, which was previously isolated from potato (Dees et al., 2017). To confirm the pathogenicity, nine broccoli seedlings were stab inoculated with a bacterial suspension (108 CFU·ml-1), while sterile distilled liquid LB medium was used as a negative control. The seedlings were kept at 80% relative humidity and 28°C in a growth chamber. Three trials were conducted per isolate (HYC22041801-HYC22041803). After 3 days, the inoculated petioles showed soft rot symptoms similar to those observed initially in the field, while control plants remained asymptomatic. All three isolates were re-isolated successfully from symptomatic tissues to complete Koch's postulates. P. polaris has been previously reported as the causative agent of blackleg in potato in several countries, including Norway, Poland, Russia, and China (Handique et al. 2022; Wang et al. 2022). Additionally, it was reported to cause soft rot in Chinese cabbage in China (Chen et al. 2021). However, this is the first report of P. polaris causing soft rot disease in broccoli in China. This discovery is of great importance for vegetable growers because this bacterium is well established on Cruciferous vegetables in the local area, and effective measures are needed to manage this disease.

Bisifusarium lunatum causing cladode soft rot in pear cactus (Nopalea cochenillifera L. Salm-Dyck) in Brazil.

Cactus pear var. miúda (Nopalea cochenillifera L. Salm-Dyck) is an important crop for the Northeast region of Brazil, composing one of the main sources of animal feed. By April 2021, cladode rot caused death of several cactus pear plants in a production area located in Itaporanga, Paraíba state, Brazil (7°21'55.35" S and 38°11'38.68" W). The infected cladodes showed brown circular necrotic spots, and soft rot with perforations that extended throughout the cladode, followed by tipping over and death of the infected plants. The incidence of the disease ranged from 10 to 30% of the plants. Bisifusarium strains were isolated and cultured on potato dextrose agar (PDA) and syntetic-nutrient-poor-agar (SNA). The colonies showed purple color on PDA. On SNA, macroconidia (n = 100) were abundant, hyaline, slightly falcate, three-septate, measuring 11.0-23.1 x 2.3-4.1 μm. Microconidia (n = 100) were oval, generally aseptate, measuring 4.1-8.7 x 2.3-3.0 μm. Conidiogenic cells formed into short monophialides. Chlamydospores were not observed. According to these morphological features, the pathogen was initially identified as Bisifusarium lunatum (Gryzenhoutm et al. 2017). For further confirmation of the identification, the partial sequences of translation elongation factor 1-alpha (TEF1-α) and the second largest subunit of RNA polymerase II (RPB2) genes were sequenced for a representative isolate (CMA 34: GenBank accession no: TEF1-α: OR536502; and RPB2: OR553509) and compared to other Bisifusarium species from GenBank database. Subsequently, it was subjected to a phylogenetic analysis of maximum likelihood including previously published sequences. According to BLAST searches, the TEF1-α and RPB2 sequences were 99% (637/640 nt) and 100% (312/312 nt) similar to B. lunatum (COUFAL0213: TEF1-α (MK640219), and RPB2 (MK301291)), respectively. The isolate was also clustered in a clade containing the ex-type of B. lunatum with 100% support (SH-aLRT and UFboot), being confidently assigned to this species. The pathogenicity test was performed after Medeiros et al. (2015), by using healthy two months old cactus pear seedlings (n = 10) cultivated in a greenhouse. Sterile toothpicks were distributed over colonies of the representative isolate grown on PDA at 25 ± 2 °C for seven days. Seedling cladodes were stuck with the toothpicks, moistened with sterile water and covered with transparent plastic bags for 24h, thus simulating a humid chamber. Following three months, all control plants (stuck with sterile toothpicks) remained healthy, while those inoculated with the representative isolate exhibited rot symptoms. This test was performed twice. B. lunatum was reisolated from symptomatic cladodes and identified as previously described, thus fulfilling the Koch's postulates. To our best knowledge, this is the first report of B. lunatum causing soft rot on N. cochenillifera in Brazil. Besides N. cochenillifera, this species was also reported on Opuntia ficus-indica in India (Gryzenhoutm et al., 2017), which raises concern regarding its ability to infect other forage sources for cattle feed in Brazilian semiarid regions. The present study highlights that the precise identification of B. lunatum is a key factor to adjust control strategies and management of the disease to prevent the spread of this disease to prevent its spread to other crops.

The first report of rot disease of onion occurred by Pseudomonas kitaguniensis, causing severe rot symptoms at lower temperatures

The first report of rot disease of onion occurred by Pseudomonas kitaguniensis, causing severe rot symptoms at lower temperatures

Surviving the Potato Stems: Differences in Genes Required for Fitness by Dickeya dadantii and Dickeya dianthicola.

Bacteria belonging to the genus Dickeya cause blackleg and soft rot symptoms on many plant hosts, including potato. Although there is considerable knowledge about the genetic determinants that allow Dickeya to colonize host plants, as well as the genes that contribute to virulence, much is still unknown. To identify the genes important for fitness in potato stems, we constructed and evaluated randomly barcoded transposon mutant (RB-TnSeq) libraries of Dickeya dadantii and Dickeya dianthicola. We identified 169 and 157 genes important for growth in D. dadantii and D. dianthicola in stems, respectively. This included genes related to metabolic pathways, chemotaxis and motility, transcriptional regulation, transport across membranes, membrane biogenesis, detoxification mechanisms, and virulence-related genes, including a potential virulence cluster srfABC, c-di-GMP modulating genes, and pectin degradation genes. When we compared the results of the stem assay with other datasets, we identified genes important for growth in stems versus tubers and in vitro conditions. Additionally, our data showed differences in fitness determinants for D. dadantii and D. dianthicola. These data provide important insights into the mechanisms used by Dickeya when interacting with and colonizing plants and thus might provide targets for management.

Genome Sequence Resource of a Taro Bacterial Soft Rot Pathogen, Dickeya fangzhongdai ZXC1

Dickeya fangzhongdai ZXC1 is a newly identified highly virulent causal agent of taro bacterial soft rot disease isolated from a taro sample showing typical soft rot symptoms in Shaoguan, Guangdong Province, China. The pathogen produces plant cell wall–degrading enzymes that macerate the taro tube tissues. In this study, we report the whole-genome sequencing analysis of strain ZXC1. The results showed that strain ZXC1 has one circular DNA chromosome of 5,129,951 bp with a 56.59% G+C content. In addition to sharing a conserved zms gene cluster that encodes the genes for biosynthesis of phytotoxin zeamines and 11 copies of predicted pectate lyase genes, the strain ZXC1 genome contains more prophage loci and a higher number of type IV secretion system and type IV secretion system gene clusters than the D. fangzhongdai strains isolated from pears or phalaenopsis, which may account for its strong virulence phenotype. The data from this study present a valuable resource for elucidation of the pathogenic mechanisms of D. fangzhongdai and may aid in developing new disease control approaches to safeguard taro production. [Formula: see text] Copyright © 2024 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license .

First report of Fusarium falciforme causing root rot and wilt on strawberry in Sinaloa, Mexico.

In Mexico strawberry production has great economic importance for the local and export markets as the country is the main strawberry supplier to the United States (SIAP, 2020). In 2022, strawberry plants with yellowing and wilting leaves, root rot and wilting, necrosis of vascular bundles and small fruits symptoms were observed in different commercial fields in the north-central Mexican state of Sinaloa, causing yield losses of about 10%. Typical Fusarium spp. colonies were recovered from all samples. They produced abundant white aerial mycelium with cream to orange pigment and branched septate hyphae (Fig. 1) (Leslie and Summerell, 2006). A total of 18 monosporic isolates were obtained by serial dilutions. The 18 isolates grown for 10 days on carnation leaf agar (CLA) produced hyaline microconidia with 0-2 septa, measuring 9.2 - 15.4 by 4.5 - 6.5 μm (n = 40) and hyaline macroconidia with three septa that measured 28.4 - 53.5 by 4.5 - 9 μm (n = 40). Chlamydospores were not observed. A fragment of the translation elongation factor 1-alpha (EF1-alpha) gene was amplified by polymerase chain reaction (PCR) using the primer pair EF-1/EF-2 (O'Donnell et al. 1998) from two monosporic isolates. The sequences were registered in the NCBI GenBank under accession numbers OR878541 and OR878543 (FRESIN178 and FRESIN194). BLASTn queries of NCBI GenBank identified the sequences as F. falciforme with 98% and 100% similarity to accession numbers OQ262968 and DQ246941 respectively. Fusarium ID database also identified the sequences as F. falciforme, is a member of the F. solani species complex (FSSC). Phylogenetic analysis revealed the partial EF1 sequences grouped with F. falciforme (Fig. 2). A pathogenicity test was performed on thirty strawberry plants (cv. Cabrillo) grown in sterile vermiculite. The plants were inoculated by immersing roots in 20 mL of a conidial suspension (1 × 105 conidia/mL) of isolate FRESIN194. Twelve uninoculated plants served as the control. All plants were grown for 60 days under greenhouse conditions (28 to 35°C). The assay was repeated twice. After 50 days, symptoms of root rot and wilting leaves like those observed in the field were observed. Uninoculated control plants did not develop symptoms. The fungus was reisolated from necrotic tissues of the inoculated plants and identified as F. falciforme by sequencing the EF1-alpha gene and morphological characteristics, completing Koch's postulates. Fusarium falciforme has been reported as the causal agent of root rot, stem rot, and wilt of tomato, papaya, chickpea, onion, common bean, and maize in Mexico (Díaz-Najera et al. 2021, Douriet-Angulo et al. 2021, Felix et al 2022, Tirado-Ramírez et al 2018, Vega-Gutiérrez et al. 2019a, Vega-Gutiérrez et al. 2019b). To our knowledge this is the first report of F. falciforme causing root rot and wilt on strawberry in Sinaloa, Mexico. This result provides useful information for the development and implementation of disease control strategies to mitigate damage caused by F. falciforme.

First report of Phytophthora infestans genotype US17 causing late blight on potato and tomato in British Columbia, Canada

Potato and tomato are grown in all Canadian provinces and contribute billions of dollars in exports and sales. Late blight, caused by Phytophthora infestans, is a destructive disease of potato and tomato worldwide and is responsible for millions of dollars in losses annually. Changes in pathogen strains were reported to be associated with late blight epidemics in several regions and pose challenges to late blight management. In British Columbia (BC), diverse strains of P. infestans were reported. Using multiplex microsatellite or simple sequence repeat (SSR) markers, we identified the new genotype US17 of P. infestans from samples of potato and tomato with late blight in BC. All isolates were of the A1 mating type. One, five, and one isolates were sensitive, moderately resistant, and highly resistant to metalaxyl-m, respectively. This is the first report of the occurrence of the US17 genotype in BC and Canada. The US17 strain was also able to cause tuber rot and foliar blight symptoms in potato in pathogen-inoculated assays.

First Report of Bacterial Leaf Streak of Rice Caused by Pantoea ananatis in Guangdong Province, China.

Rice (Oryza sativa L.) is a crucial staple crop worldwide, and bacterial diseases are among the primary factors affecting rice yield. In late October 2022, bacterial leaf streak disease was observed on the leaves of the rice variety Meixiangzhan 2 across multiple fields (approximately 130 hm2) in Leizhou City, Guangdong Province, China. The incidence rate was up to 30% in each field. Infected rice leaves exhibited distinctive symptoms at the boundary between diseased and healthy tissue, featuring dark green to yellow-brown streaks, while most of the leaf margin exhibited symptoms of either leaf edge or sheath rot. Disease progression from the leaf tip inwards revealed gray-white or dehydrated lesions with a bluish-gray color. Some leaves exhibited wrinkling at the edges, and severe symptoms at the leaf tip resembled those of bacterial leaf blight in rice. Ten leaves were collected from 10 infected rice plants in three distinct fields, and leaf pieces at the border of diseased and healthy areas were surface disinfected with 75% anhydrous ethanol for 60 seconds, rinsed three times with sterile water, and then soaked in sterile water for 8 hours. The obtained bacterial suspension was diluted at a ratio of 1: 106, and 100 μL of the diluted samples were plated on Potato Dextrose Agar (PDA) plates. After incubation at 28°C for 48 hours, the yellow bacterial colonies that appeared, were purified on PDA plates. To confirm the bacterial species, the amplification of genes gyrB, leuS, rpoB, and 16S rDNA was performed on six randomly selected isolates from the three different fields using the primers 27F/1492R, gyrB-F/R, leuS-F/R and rpoB-F/R, as reported by Yu et al (2022), respectively. PCR products were sequenced. All six isolates had identical sequences for all genes sequenced.The gene sequences of 16S rDNA (960 bp), gyrB (953 bp), leuS (733 bp), and rpoB (877 bp) for LZ1, were deposited in the NCBI database under accession numbers PP048830 , PP068625 , PP068626, and PP068627, respectively. These sequences were subsequently compared using BLASTn tool against the NCBI nr/nt database. The 16S rDNA, gyrB, leuS, and rpoB of LZ1 showed similarities of 99.90%, 99.16%, 99.73%, and 99.89%, with the corresponding sequences of P. ananatis TZ39 (GenBank accession numbers MZ800600.1 for 16S rDNA, and CP081342.1 for gyrB, leuS and rpoB ). MLSA analysis using concatenated sequences of gyrB, leuS, and rpoB genes indicated that the isolated strain LZ1 belongs to P. ananatis. In the tillering stage of rice varieties Meixiangzhan 2 and Huahangyuzhan, P. ananatis LZ1 was inoculated at a concentration of 108 CFU/mL using the leaf-cutting method, with sterile water used as a control (Toh et al., 2019). After 14 days of bacterial inoculation, the inoculated leaves gradually became necrotic, changing from light green to brown showing identical symptoms as those in the field, while the control plants remained symptom-free. Subsequent 16S rDNA, gyrB, leuS and rpoB gene sequencing results further confirmed the identity of the pathogen as P. ananatis, thereby fulfilling Koch's postulates. Previous reports have already identified P. ananatis as the pathogen causing rice bacterial leaf streak (Kini et al., 2017; Arayaskul et al., 2019; Yu et al., 2022; Lu et al., 2022; Luna et al., 2023; Yuan et al., 2023). This is the first report of rice bacterial leaf streak caused by P. ananatis in Guangdong Province, China, laying the foundation for future research to establish strategies for the prevention and control of this disease.

Insights into the Interaction between the Biocontrol Agent Bacillus amyloliquefaciens QST 713, the Pathogen Monilinia fructicola and Peach Fruit

Brown rot disease caused by Monilinia fructicola is one of the most important peach fruit threats in the world. The use of biological control agents (BCAs), instead of synthetic fungicides, to successfully inhibit postharvest disease development is a challenge in sustainable and efficient crop management. The commercially available BCA Bacillus amyloliquefaciens QST 713 (formerly Bacillus subtilis QST713) is able to inhibit a variety of fungal pathogens and suppress several plant diseases. Our results showed that this BCA inhibited mycelial growth in vitro, and was able to suppress the disease’s severity in peach fruits via delaying and reducing brown rot symptoms. A transcriptomic analysis of fruits during their pre-treatment with this biocontrol agent following M. fructicola challenge revealed a significant upregulation of specific differentially expressed genes (DEGs) at 48 h after inoculation (HAI). These genes are related to the activation of several transcriptional factors, such as members of the WRKY and NAC families, and receptors that are involved in pathogen recognition and signaling transduction (e.g., LRR-RLKs). Furthermore, the inhibition of M. fructicola by this biocontrol agent was confirmed by analyzing the expression profiles of specific fungal genes, which highlighted the direct antimicrobial impact of this bacterial strain against the fungus. Hence, these findings clearly suggest that B. amyloliquefaciens QST 713 is an efficient BCA against brown rot disease, which can directly inhibit M. fructicola and improve peach fruit tolerance.

First report of Macrophomina phaseolina causing сharcoal rot in soybean (Glycine max (L.) Merr.) plants in Uzbekistan.

The soybean production area is expanding in Uzbekistan. Soybeans were planted on an area of 10 thsd ha and the harvest amounted to 30 thsd metric tons in 2023 (IPAD, https://ipad.fas.usda.gov/countrysummary). Macrophomina phaseolina (Mp) is a soil- and seed-borne fungal pathogen causing economically important diseases of legume crops (Pennerman et al. 2024). Drought stress and a warm climate are favorable to this pathogen (Irulappan et al. 2022). Under these conditions, its microsclerotia survive for a longer period and become more virulent (Chamorro et al. 2015). In August 2022, typical symptoms of charcoal rot were observed in about 25% of "Orzu" soybean cultivar affecting 6 ha located on the experimental base "Durmon" of our institute. Diseased plants displayed the following charcoal rot symptoms: leaves turn yellow, then wilt, die, and remain attached to the plant; the lower portion of the stem and tap root have a light gray or ashy black discoloration; tiny black specks on the lower stem and root; after splitting the stem, it has the appearance of fine charcoal powder. In order to determine the causal agent of these symptoms, a total of 17 diseased plants were collected from focal lesions in soybean plantings. From each plant, twelve sections of stem and root tissue were selected, cut into small 5-mm pieces, and surface sterilized with 1% sodium hypochlorite for four minutes, then rinsed three times with sterile distilled water. The disinfected tissues were dried on sterile filter paper for 5 min and placed on PDA Petri plates, which were incubated in an incubation chamber for 3 days (16 h light (26oC) and 8 h dark (18oC)). Fungi were subsequently subcultured on PDA and incubated for 7 days to obtain pure cultures. Six monohyphal colonies were purified. The colonies showed dense growth, with a gray initial mycelium becoming darker with aging. After 8 days on PDA, black-colored microsclerotia with spherical to oblong shapes were observed. On average, they measured 60 µm in width and 130 µm in length (n = 30). From six isolated monohyphal colonies, one has been chosen for molecular-genetic identification. Molecular-genetic analysis was conducted by amplification and sequencing of the ITS region with the ITS1 and ITS4 primers (White et al. 1990). The resulting sequence was deposited in the NCBI database under accession number OQ073450. After BLAST analysis (Altschul et al. 1990) it was 100% identical with the reference sequences of Mp (accession MT039671, MT039663 and MH496040) isolated in sugar beet, maize and sunflower, respectively, from Serbia. In order to verify the pathogenicity, soybean seedlings (cv. Orzu) were dipped into spore suspension (1 × 107 spores/ml) of sequenced strain R-17 for 1 minute and transferred to a 15 cm diameter plastic pot with 350 g of sterilized soil mix. After 25 days, the inoculated plants showed classic charcoal rot symptoms, while the control plants remained healthy. The pathogen was successfully reisolated from the infected seedlings onto PDA, fulfilling Koch's postulate. The identity of the re-isolated strain was confirmed by morphological features and sequencing of the ITS region. It should be noted that in Uzbekistan, Mp has not been documented in any plants. Therefore, according to our knowledge, this is the first report of this fungus affecting soybean plants in Uzbekistan. Since molecular-genetic analysis of the R-17 strain showed clustering with strains from Serbia, we speculate that there may have been a recent introduction of Mp from Serbia into Uzbekistan. This assumption is additionally confirmed by the fact that Serbia is the largest seed exporter in Uzbekistan. The increase in charcoal rot disease poses a major challenge to soybean production in Uzbekistan. Understanding the genetic diversity of Mp can be utilized to manage this disease, improve soybean yield, and help soybean breeding programs in Uzbekistan.

Isolation and identification of Podospora flexuosa (syn. Cladorrhinum flexuosum), a potential biocontrol agent detected in sugar beet cultivation areas in Türkiye

Sugar beet (Beta vulgaris L.) is widely cultivated in Konya province of Türkiye and about one third of Türkiye’s sugar beet production is obtained from this province. As a result of the isolations made from plants showing severe root rot and desiccation symptoms in sugar beet fields in the region in 2015-2017, a new endophytic fungal isolate was obtained from leaves along with soil and foliar disease agents belonging to different fungal species. The fungal isolate has dull yellow, light olive to light brown, fast growing colony-like, flexible conidiophores and septate branched hyphae. The representative fungal isolate did not cause disease symptoms on host plant leaves. Molecular identification of the fungal isolate was carried out using primers specific to the ITS gene region, and it was identified as Podospora flexuosa (syn. Cladorrhinum flexuosum) based on morphological and molecular characteristics. Since P. flexuosa is reported to be a biological control agent living as a saprophyte in soil and plant materials, it is thought that the fungal isolate obtained in our study can be used as a potential biocontrol agent in the biological control of plant disease agents in the future. To the best of our knowledge, this fungal species is reported for the first time for Türkiye.

Elaborate Report of Allium sativum Clove Brown Rot Caused by Fusarium solani in China.

In September 2023, brown rot disease was observed on cloves of garlic (Allium sativum) variety "Zipi-1" (purple skin) collected from Jinxiang County, China during scientific research at the Beijing Academy of Agricultural and Forestry Sciences after being planted in nutrient soil for approximately 2 weeks in a growth chamber maintaining 22℃, 60% relative humidity, and 16 hours of light. Out of the 90 garlic cloves investigated, 18 showed signs of decay, characterized by a brown color and rot, with the surface covered in blue and white mold layers. Six symptomatic cloves were collected for isolating the pathogen using the method described in a previous study (Wu et al. 2021). After 2 d of incubation, individual spores were harvested from the fungal colonies and recultured. Single-spore cultures growing on PDA medium appeared white and flocculent when viewed from the top, and yellowish-brown when viewed from the bottom. After 5 d of cultivation, the colonies had a diameter of approximately 5.8 cm and microscopic examination revealed that the mycelium had a diameter of about 9-13 µm (Fig. S1a, b and c). Isolate As1 produced three types of spores: oval-shaped chlamydospores with a diameter of approximately 6 µm, while spindle-shaped microconidia and sickle-shaped macroconidia measuring approximately 6-7 × 20-30 µm and 8 × 50 µm, respectively (Fig. S1d and e). The mycelial characteristics and reproductive structures of the isolates fit the morphological description of Fusarium solani (Xie et al. 2022). To confirm the identification, TEF1, RPB1 and RPB2 regions of the genome were amplified from three separate isolates (As1, As2, and As3) using EF1/EF2, RPB1-Fa/G2R, RPB2-5F2/7cR, and RPB2-7cF/11aR primer pairs (O'Donnell et al. 2022). The results indicated that the sequences of the three isolates were completely identical. Furthermore, the BLASTn comparison results of the TEF1 (OR916018, 710bp, 100%), RPB1 (OR916019, 1797bp, 99.8%), and RPB2 (OR916020, 1874bp, 100%) sequences in the FUSARIUM-ID v.3.0 database revealed that As1 was identified as F. solani species complex 5 (O'Donnell et al. 2022). To assess the pathogenicity of As1, the surface of healthy garlic cloves (n = 30) was spread with 106 microconidia/mL As1 suspension, while a control group (n = 30) was inoculated with sterile water. All inoculated cloves were placed in an artificial climate chamber under same conditions described above. After 10 d, all inoculated cloves exhibited rot symptoms consistent with those of the initially infected cloves identified in September 2023, while the control plant cloves remained asymptomatic (Fig. S2). Based on morphological and molecular characters (TEF1, RPB1, and RPB2), the reisolated pathogen from diseased plants was identical to the As1 isolate used for inoculation, and the disease assays were repeated twice. Fusarium spp. has been reported as the causal agent of garlic rot disease in several countries such as Mexico, America, and Russia (Gálvez and Palmero 2022). Tai (1979) previously published a report on the presence of F. solani in garlic; however, the content in the book is rather basic, lacking detailed information on the isolation, identification, and the potential for causing garlic diseases, whether postharvest or during growth. Our research can be considered a supplement and improvement to the study by Tai (1979) and lays the groundwork for future studies on management strategies to combat plant diseases caused by F. solani.

First Report of Botryosphaeria dothidea Causing Postharvest Fruit Rot of Plum in China.

The Nai plum (Prunus salicina var. cordata cv. Younai) holds significance as an important deciduous fruit crop in China. In July 2023, symptoms of postharvest fruit rot were observed on Nai plum with a 10% disease incidence of harvested fruits in three supermarkets, located in Nanchang City, Jiangxi Province, China. Infected fruits displayed brown, circular lesions, accompanied by a transition in the surrounding peel color from cyan to red. To investigate the causal agent, small sections (3 to 4 mm2) from the periphery of ten infected fruits were subjected to surface sterilization using 75% ethanol for 30 seconds. Following sterilization, the samples were rinsed three times with sterilized distilled water, air-dried, and aseptically placed on potato dextrose agar (PDA) at 25 ℃ for 3 days. Isolated colonies were subcultured by hyphal tiptransfer. Ten of the resulting 12 fungal isolates showed similar morphological characteristics. The colonies exhibited an initial white hue, gradually transitioning to gray, and featured short and thick aerial hyphae with an irregular colony margin. Microscopic examination revealed conidiogenous cells that were hyaline, aseptate, and narrowly fusiform. The conidia were measured 11.0 to 15.6 × 3.2 to 4.9 µm (x̅ = 13.5 ± 1.4 × 4.0 ± 0.4 µm, n = 30), and were hyaline and subcylindrical. The morphological characteristics were in accordance with those of the Botryosphaeria species (Crous et al. 2006). To identify the strain, two representative isolates, JFRL03-1792 and JFRL03-1793, were selected for further characterization. Amplification of nucleotide sequences from three regions (ITS, TEF1-a and TUB2) was conducted using the primer sets ITS5/ITS4, EF1-728F/EF1-986R, and BT2A/BT2B, respectively (Guo et al. 2023). The resulting sequences were deposited in GenBank under the accession numbers: OR418373 and OR418374 for ITS; OR424405 and OR424405 for TEF1-a; OR424411 and OR424412 for TUB2. A BLASTN homology search of the obtained sequences revealed a high similarity of 99%-100% to the ITS (AY236949, 511/513 nucleotides), TEF1-a (AY236898, 282/282 nucleotides), and TUB2 (AY236927, 431/431 nucleotides) sequences of Botryosphaeria dothidea CWM8000 (ex-type). Maximum likelihood analyses were performed for the combined ITS, TEF1-a, and TUB2 dataset using Phylosuite V1.2.2 (Zhang et al. 2020). The resulting phylogenetic tree indicated that the two representative isolates were clustered together with Botryosphaeria dothidea in a clade with 95% bootstrap support. Based on the comprehensive assessment of morphological and molecular data, the two isolates were conclusively identified as B. dothidea. To confirm pathogenicity, six healthy Nai plum fruits were surface sterilized with 75% ethanol and were subsequently wounded with a sterile needle. A 5-mm-diameter mycelial plug of the isolate JFRL03-1792, cultured on PDA at 25 ℃ for three days, was applied to the wound. Another set of six fruits was inoculated with sterile agar plugs as control. Following incubation in a climatic chamber at 25 ℃ and 80% relative humidity, the fruits were examined after 5 days. The experiment was repeated twice. The fruits inoculated with B. dothidea displayed typical rot symptoms, while the control fruits remained asymptomatic. In adherence to Koch's postulates, the fungus was successfully re-isolated from the inoculated fruits and confirmed as B. dothidea through morphological and molecular analysis. B. dothidea has previously been reported causing fruit rot on kiwifruit, winter jujube, and apple (Tang et al. 2012; Zhou et al. 2015; Marsberg et al. 2017; Xu et al. 2023). In addition, B. dothidea also reported causing Botryosphaeria canker disease on plum (Lin et al. 1994). But to our knowledge, this is the first documentation of B. dothidea causing postharvest fruit rot on plum in China. This discovery imparts critical insights into the management of this high-risk disease affecting plum in China.

First Report of Monosporascus cannonballus causing Root Rot and Vine Decline in Watermelon (Citrullus lanatus) in China.

In June 2023, a sudden outbreak root rot and vine decline symptoms was observed during a watermelon (Citrullus lanatus T.) variety demonstration trial located in Taizhou City, Zhejiang Province, China, with an incidence rate ranging from 75% to 100% and an affected area of nearly 2,000 square meters. The disease initially appeared with a rapid and alarming invasion of root rot and vine decline symptoms within watermelon plants. Affected plants exhibited rapid deterioration, showing symptoms of wilting, yellowing and eventual demise, predominantly during the pre-harvest stage. Notably, numerous black, spherical, erumpent perithecia were clearly visible on the watermelon's root epidermis, a characteristic trait of the disease. Symptomatic plant samples were rigorously disinfected with 75% ethanol, and plated on potato dextrose agar medium for incubation at 25°C, successfully isolate two potential strains. These isolates were inoculated in oatmeal agar and incubated in a 25℃ light incubator. After 30 days, mature perithecia, the same as those found on the watermelon's root epidermis, reached a diameter of 500 μm. Each perithecium contained several pear-shaped asci, 56 to 108.5 μm in length and 30.5 to 46.4 μm in width, typically holding 1, rarely 2 ascospores. These characteristics align precisely with the typical strains of Monosporascus cannonballus Pollack and Uecker (1974). Additionally, sequencing the internal transcribed spacer region of ribosomal DNA (ITS) gene (White et al., 1990), large subunit ribosomal RNA (LSU) gene (Rehner and Samuels 1995), and beta-tubulin (TUB) gene (Glass and Donaldson, 1995) were performed. BLAST analysis indicated the highest nucleotide sequence identity with M. cannonballus CBS 586.93 reference sequence (ITS: 100%, JQ771930; TUB: 98.99%, JQ907292). Representative sequences of isolate ZJUP0990-2 from these regions were deposited in GenBank (Accession No.: OR357656 for ITS; OR474500 for LSU; OR365762 for TUB). A multigene phylogenomic analysis (ITS-LSU-TUB) was undertaken to ascertain the exact phylogenetic position of M. cannonballus within the genus Monosporascus. The amalgamation of both morphological and molecular insights consistently reaffirmed the accurate classification of the causative agent as M. cannonballus. To validate the pathogenicity of M. cannonballus, a controlled greenhouse experiment was conducted using watermelon (cv. Nabite) as the subject. Mycelium fragments, harvested from the edge of the colony ZJUP0990-2, were inoculated into oat liquid medium and cultivated under dark conditions at a consistent temperature of 30°C for 7 days. After 20 days, the inoculated plants exhibited root rot and wilting, mirroring the symptoms observed during the field outbreak. In contrast, the control plants did not exhibit any signs of disease. M. cannonballus was successfully re-isolated from the symptomatic roots of the inoculated plants, satisfying Koch's postulates. This experiment was repeated three times. This pathogenic fungus has previously been documented as a menace to melons in various regions including Mexico (Chew-Madinaveitia et al., 2012) and Brazil (Sales et al., 2004), as well as watermelons in Brazil (Sales et al., 2010), northern Mexico (Gaytan-Mascorro et al., 2012), and Saudi Arabia (Karlatti et al., 1997). To our knowledge, this is the first reported presence of M. cannonballus on watermelons in China. This new disease poses a serious threat to watermelon production, potentially leading to severe economic losses and impacting food security.