Powdery Mildew of Xanthium strumarium Caused by Podosphaera xanthii in central China.

Xanthium strumarium, known as cocklebur, is an annual herb and has been used in traditional Chinese medicine. In October 2020, powdery mildew-like disease signs and symptoms were observed on X. strumarium grown in a crop field, Xinxiang city, Henan Province, China (35.36076° N, 113.93467° E). The specimen (PX-XS2023) was stored in Xinxiang Key Laboratory of Plant Stress Biology. White colonies in irregular or coalesced circular shaped-lesions were abundant on both ad- and abaxial surfaces of leaves and covered up to 99 % of the leaf area. Some of the infected leaves were senesced. More than 70 % of plants (n = 130) exhibited these signs and symptoms. Conidiophores were straight or slightly curved, 55 to 160 × 11 to 13 µm composed of foot-cells, shorter cells and conidia. Conidia were ellipsoid to oval, 29 to 40 × 14 to 20 µm (n = 50), with a length/width ration of 2.0 to 2.5, containing fibrosin bodies. Dark brown to black chasmothecia were found on infected leaves. The appendages were mycelium-shaped and at the base of scattered or gregarious chasmothecia (n = 50, 70 to 120 µm in diameter). Asci were 55 to 80 × 50 to 65 µm (n=30). These morphological characteristics were consistent with those of Podosphaera xanthii (Braun and Cook 2012). The internal transcribed spacer (ITS) region and Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) region of the fungus (PX-XS2023) were amplified and sequenced with primers ITS1/ITS4 (White et al. 1990) and GAPDH1/GAPDH3R (Bradshaw et al. 2022) according to a previously reported method (Zhu et al. 2022). The resulting sequences were respectively deposited into GenBank (Accession No. MW300956 and PP236083). BLASTn analysis indicated that the sequences were respectively 99.82 % (564/565) and 100% (272/272) identical to P. xanthii (MT260063 and ON075658). The phylogenetic analysis indicated that the strain PX-XS2023 and P. xanthii were clustered into a same branch. Therefore, the causal agent of powdery mildew on X. strumarium was P. xanthii. To conduct pathogenicity assays, mature leaves of five healthy X. strumarium (height in 50 centimeters) were inoculated with fungal conidia by gently pressing surfaces of infested leaves onto leaves of healthy plants (Zhu et al. 2020). Five untreated plants served as controls. The controls and inoculated plants were separately maintained in greenhouses (humidity, 60%; light/dark, 16 h/8 h; temperature, 18°C). Eight days post-inoculation, signs of powdery mildew were detectable on inoculated plants, however, the controls were asymptomatic. Thus, the fungal pathogen was morphologically and molecularly identified and confirmed as P. xanthii. This powdery mildew caused by P. xanthii was previously reported on X. strumarium in Korea, Russia and India (Farr and Rossman, 2021). In addition, P. xanthii was recorded on X. strumarium in Xinjiang Province, China (Tai 1979). However, this is the first report of P. xanthii on X. strumarium in central China, where is around 3000 km away from Xinjiang Province with geographically differences. The sudden presence of powdery mildew caused by P. xanthii may adversely affect plant health and thus reduce medical value of X. strumarium. Therefore, the identification and confirmation of P. xanthii infecting X. strumarium enhance the knowledge on the hosts of this pathogen in China and will provide fundamental information for disease control in the future.

First Report of Leaf Spot Disease on Lonicera japonica Caused by Nigrospora oryzae in China.

Lonicera japonica Thunb. is a traditional Chinese medicinal plant, which widely cultivated in China, Japan and Korea. From August to October in 2021 and 2022, severe leaf spots symptoms were observed on L. japonica in medicinal botanical garden of Shandong University of Traditional Chinese Medicine (36°55'89"N, 116°79'91"E), Jinan, Shandong Province, China. The disease incidence was above 80% in the 25 acre cultivation area. Early symptoms were small brown spots on the leaves. Then the number of small spots gradually increased and spread over the entire leaves. The small brown spots seldom merge together to form larger lesions. Leaves with typical symptoms were collected from twenty individual plants, and cut into small 5×5 mm fragments in the junction of infected and healthy tissues. The fragments were sterilized in 75% ethanol for 30 s and 1% NaClO for 60 s, rinsed three times in sterile water, and then placed on potato dextrose agar (PDA). After 3 days of incubation at 25°C, fungal plugs along the edge of the colony were cut and transferred to new PDA for purification. A total number of 23 colonies with similar morphological characteristics were obtained, and three representative strains (Lj14, Lj18 and Lj20) were selected for subsequent study. The colonies grew rapidly on PDA and covered the entire petri dish in 4 days. Colonies had abundant aerial hyphae, initially white, round, later turning gray and black. Conidia were oblate or nearly spherical, single-celled, black, and measured in size from 9.6 to 13.2 µm × 7.9 to 16.1 µm in diameter (n=150) (Figure S1). The observed characteristics were close to those of Nigrospora spp. ( Wang et al. 2017). The genomic DNA was extracted, and PCR amplification of the rDNA internal transcribed spacer (ITS), ß-tubulin gene (TUB), and translation elongation factor 1-alpha gene (TEF1) were completed by primers ITS1/ITS4, Bt2a/Bt2b and EF1-728F/EF1-986R (Carbone and Kohn, 1999). Sequences were deposited in GenBank (accession nos. OR936661, OR936662, OR936671 for ITS, OR947626, OR947627, OR947628 for TUB, and OR947629, OR947630, OR947631 for TEF1 sequences, respectively). BLAST analyses of ITS (OR936661), TUB(OR947626) and TEF1 (OR947629) sequences exhibited 100% (487 bp out of 487 bp), 99.48% (380 bp out of 382 bp), and 99.6% (248 bp out of 249 bp) similarity to the sequences of N. oryzae strains KoLRI_053384 (MZ855426), LC2991 (KY019496) and LC7307 (KY019409), respectively. Lj14, Lj18 and Lj20 formed a clade with N. oryzae LC6763 and LC2991 in phylogenetic tree (Figure S2). Based on morphological and molecular evidence, the pathogen was identified as N. oryzae (Berk. &Broome) Petch. To fulfill Koch's postulates, the pathogenicity was tested in vivo experiments. Thirty non-wounded healthy leaves of ten intact plants were inoculated with 10 µl spore suspension (106 spores/ml) of three strains, respectively. As negative control, thirty leaves of ten healthy plants were inoculated with sterile water. The inoculated plants were placed at 28°C in the growth chamber with high relative humidity. The pathogenicity tests were repeated three times. Distinct symptoms similar to that of natural conditions were observed on the leaves of inoculated plants after 4 to 7 days. The strain was reisolated from the lesions and identified as N. oryzae by morphological features and ITS sequence. The pathogen has been reported to cause leaf spot disease on tobacco (Wang et al. 2022) and asiatic dayflower (Qiu et al. 2022). To our knowledge, this is the first report of leaf spot caused by N. oryzae on Lonicera japonica in China. The research will be helpful for leaf spot disease control.

First Report of White Mold Caused by Sclerotinia sclerotiorum on Hymenocallis glauca in Mexico.

In Mexico, there are 29 native species of the genus Hymenocallis, where H. glauca is one of the most cultivated bulbous plants. It holds economic importance as it is commercialized as a potted plant and cut flower (Leszczyñska and Borys, 2001). In October 2023, field sampling was conducted in the Research Center in Horticulture and Native Plants (18°55'55" N, 98°24'02.8"W) of UPAEP University. H. glauca diseased plants were found in an area of 0.4 ha, with an incidence of 35% and an estimated severity of 45% on infected plants in vegetative stage. The symptoms included chlorosis of foliage, necrosis at the base of the stem, and soft rot with abundant white to gray mycelium and abundant production of black, irregular sclerotia of approximately 3.5 mm diameter. Finally, the plants wilted and died. The fungus was isolated from 40 symptomatic plants. Sclerotia were collected, disinfested with 3% NaOCl for one minute, rinsed with sterile distilled water (SDW), and plated on Petri dishes containing potato dextrose agar (PDA) with sterile forceps. Subsequently, a sterile dissecting needle was used to place fragments of mycelium directly on Petri dishes with PDA. Plates were incubated at 23 °C in dark for 7 days. One isolate was obtained from each diseased plant by the hyphal-tip method (20 isolates from sclerotia and 20 from mycelium). After 7 days, colonies had fast-growing, dense, and cottony-white aerial mycelium forming irregular sclerotia of 3.57 ± 0.59 mm (mean ± standard deviation, n=100). In each Petri dish there were produced 21.5 ± 7.9 sclerotia (mean ± standard deviation, n=40), after 11 days; these were initially white and gradually turned black. The isolates were tentatively identified as Sclerotinia sclerotiorum based on morphological characteristics (Saharan and Mehta 2008). Two representative isolates were chosen for molecular identification and genomic DNA was extracted by the CTAB protocol. The ITS region and the glyceraldehyde 3-phosphate dehydrogenase (G3PDH) gene were amplified and sequenced (Staats et al. 2005; White et al. 1990). The sequences of a representative isolate (SsHg3) were deposited in GenBank (ITS- PP094578; G3PDH- PP101843). BLAST analysis of the partial sequences ITS (519 bp), and G3PDH (950 bp) showed 100% similarity to S. sclerotiorum isolates (GenBank: MG249967, MW082601). Pathogenicity was confirmed by inoculating 30 H. glauca plants in vegetative stage grown in pots with sterile soil. Ten sclerotia were deposited at the base of the stem, 10 mm below the soil surface. As control treatment, SDW was applied to 10 plants. The plants were placed in a greenhouse at 23 °C and 90% relative humidity. After 17 days, all inoculated plants displayed symptoms similar to those observed in the field, while no symptoms were observed on the controls. The fungus was re-isolated from the inoculated plants as described above, fulfilling Koch's postulates. The pathogenicity tests were repeated three times. S. sclerotiorum has been reported causing white mold on other bulbous plants, like fennel (Foeniculum vulgare) in Korea (Choi et al. 2015). To our knowledge, this is the first report of S. sclerotiorum causing white mold on H. glauca in Mexico. Information about diseases affecting this plant is very limited, so this research is essential for developing integrated management strategies and preventing spread to other production areas.

Colletotrichum caladii sp. nov. causing anthracnose leaf spot of Caladium × hortulanum (Araceae) in Florida, USA.

Caladium (Caladium × hortulanum) is an ornamental plant popular for its variable and colorful foliage. In 2020, plants showing leaf spots and blight, typical of anthracnose, were found in a field trial at the University of Florida's Gulf Coast Research and Education Center (UF/GCREC) in Wimauma, FL, USA. Leaf samples consistently yielded a Colletotrichum-like species with curved conidia and abundant setae production in the acervuli. The internal transcribed spacer (ITS), partial sequences of the glyceraldehyde-3-phosphate dehydrogenase gene (gapdh), actin gene (act), chitin synthase 1 gene (chs-1), beta-tubulin gene (tub2), and histone3 gene (his3) were amplified and sequenced. Blastn searches in the NCBI GenBank database revealed similarities to species of the Colletotrichum truncatum species complex. Phylogenetic analyses using multi-locus sequence data supports a distinct species within this complex, with the closest related species being C. curcumae. Based on morphological and phylogenetic analyses, a new species of Colletotrichum, named C. caladii, is reported. Pathogenicity assays and subsequent isolation confirmed that this species was the causal agent of the disease.

Phytophthora pseudocryptogea Is Associated with Root and Crown Rot of Nursery-Grown Common Sage in Hungary.

In October 2009, necrotic bark lesions at the root collar and lower stem associated with root rot, reduced growth, and wilting were observed on container-grown 2-year-old common sage (Salvia officinalis L. 'Icterina') in two ornamental nurseries in Somogy and Zala counties in Hungary. The disease occurred at a frequency of 15-20% (100 to 150 symptomatic plants in each nursery). A P. cryptogea-like species was isolated consistently from necrotic root collars of many plants on carrot (CA) PARPB agar. Six isolates from the nursery in Zala county and three isolates from the nursery in Somogy county were deposited in the culture collection of Plant Protection Institute (Budapest, Hungary). All developed slightly petaloid colonies on CA agar. Chlamydospores and gametangia were not present in single and dual culture combinations of isolates. Radial colony growth was the fastest at 25°C (6.8 to 7.4 mm/day) and no growth occurred above 34°C. On mycelial discs floating in nonsterile stream water, persistent, nonpapillate, mostly ovoid to obpyriform sporangia (37.4±3.5 to 47.8±4.6 µm long and 22.3±2.6 to 29.2±3.7 µm wide) and hyphal swellings were produced abundantly. Pathogenicity of one selected isolate from each nursery was tested on 3-month-old seedlings of S. officinalis 'Icterina' in 2010. Isolates were grown for 4 weeks at 20°C on autoclaved millet grains moistened with CA broth. Infested and uninfested grains were mixed with autoclaved soil (30 cm3 grain/liter), and the mixes were used as potting media for transplanting five treated and five control plants per isolate, respectively. Plants were kept in a growth room (20-25°C, 16/8 h dark/light). Pots were flooded for 24 hours on the 1st day and every 2 weeks. All and only treated plants showed symptoms of wilt associated with basal stem and root necrosis within three weeks. The trial was repeated with the same result. The pathogen could be reisolated only from the treated plants. Identity of isolates from nurseries and inoculated plants was confirmed recently by amplification and sequence analysis of the rDNA internal transcribed spacers (ITS) and gene regions of cytochrome c oxidase subunit I (coxI) and ß-tubulin (tub) according to Jung et al. (2017). BLASTn searches showed 100% identity and only 97.3-99.0% similarity to the corresponding sequences of authenthic P. pseudocryptogea and P. cryptogea strains, respectively (e.g., GenBank accession nos. KP288336-KP288342, KP288370-KP288372, KP288386-KP288392, MN872725, MN872776). Sequences of the 9 field isolates were deposited in GenBank under accession nos. OR771701-OR771709 (ITS), OR787508-OR787516 (coxI) and OR787517-OR787525 (tub). P. pseudocryptogea was delineated from P. cryptogea sensu lato (Safaiefarahani et al. 2015), which has been reported from S. officinalis in the United States (Koike 1997), and S. leucantha (Cacciola et al. 2002) and S. officinalis (Garibaldi et al. 2015) in Italy. The known natural hosts of P. pseudocryptogea includes plant species in families other than Lamiaceae (cf. Aloi et al. 2023), but it was pathogenic on the lamiaceous Plectranthus scutellarioides in artificial inoculations (Christova 2020). The pathogen is present in European nurseries (Antonelli et al. 2023). This is the first report of P. pseudocryptogea on S. officinalis in Hungary. The causal agent threatens the production of sages and other ornamentals, and its spread in Hungary should be prevented by proper disease management and phytosanitary actions.

Arthrocladiella mougeotii causing powdery mildew on goji berry plants (Lycium barbarum and L. chinense) and mixed infections with Phyllactinia chubutiana in California.

Goji berries (Lycium barbarum and L. chinense) have a rich historical significance in traditional Chinese medicine and have gained popularity as a superfood in Western cultures. From 2021 to 2023, powdery mildew was observed on goji plants of both species in community and residential gardens in Yolo County, California (USA). Disease severity varied from 20 to 100% of infected leaves per plant. Powdery mildew was characterized by the presence of white fungal colonies on both sides of leaves and fruit sepals. Additionally, a brownish discoloration was observed in infected mature leaves, resulting in further defoliation. Morphologically, the fungus matched the description of Arthrocladiella mougeotii. The pathogen identity was confirmed by phylogenetic analyses of the rDNA internal transcribed spacer and the 28S rDNA gene sequences. Pathogenicity was confirmed by inoculating healthy L. barbarum plants using infected leaves and successfully reproducing powdery mildew symptoms after 28 days (22°C, 60% RH), with A. mougeotii colonies confirmed by morphology. Control leaves remained symptomless. Co-infection with Phyllactinia chubutiana was detected on plants from two separate gardens, with A. mougeotii observed first in late spring (May to June) and P. chubutiana later in the summer (July to August). These results revealed that both A. mougeotii and P. chubutiana constitute causal agents of powdery mildew on goji berry plants, often infecting the same plant tissues simultaneously. To our knowledge, this is the first report of A. mougeotii causing powdery mildew on L. barbarum and L. chinense in California, which provides a better understanding of the etiology of powdery mildew of goji plants in California.

Occurrence of Neopestalotiopsis clavispora Causing Leaf Blight on Acer oblongum in China.

Acer oblongum is native to Southwest China and is also distributed in Nepal and Northern India. It is an excellent garden ornamental tree species, suitable for solitary planting in courtyards and parks. From June to August 2022, severe leaf blight occurred on A. oblongum in Baihe Wetland Park (32°5'42" N, 112°28'13" E) in Nanyang City, China. The foliar disease rate reached 59% (n=100). Early signs were yellow spots on the leaves, mainly on the middle and edge parts. Then, the lesions gradually expanded, became amorphous, and turned yellowish brown, eventually led to necrosis on leaves and branches. Twenty diseased leaves were collected and the junction areas between infected and healthy tissues were cut into 5 x 5 mm2 pieces. The collected plant materials were sterilized in 75% ethanol and 1% NaClO for 30 s and 1 minute, respectively, followed by rinsing in sterile water, and placing on a potato dextrose agar (PDA) plate supplemented with 50 µg ml-1 streptomycin at 25 °C for 3 days. Colony edges were cut and transferred to new PDA plates for purification culture. A total of 18 purified fungal strains were obtained, which showed similar phenotypes in morphological characteristics. All colonies had spread radially with wavy surfaces, and dense cream to white aerial hyphae. After 14 days in culture, black fruiting bodies appeared. Conidia were fusiform to slightly clavate, with five cells and two or three setae, 4.2 to 7.9 µm × 17.5 to 25.4 µm in diameter (n = 100). The apical and basal cells and setae were colorless, three median cells were brown, and the middle cell was dark brown. Morphological characteristics of all 18 strains were consistent with the genus description of Neopestalotiopsis spp. (Maharachchikumbura et al. 2014). Further molecular identification showed that the ITS region sequences of all strains have extremely high homology with Neopestalotiopsis spp. The ß-tubulin gene (TUB), and the translation elongation factor 1-alpha gene (TEF1) were amplified for molecular identification (Shu et al. 2020). The sequences of three representative strains (FE-05, 09, 16) from different regions were deposited in GenBank with accession Nos. OQ867279, OQ867288, OQ867289 (ITS), OQ870207, OQ870208, OQ870209 (TUB), and OQ870204, OQ870205, OQ870206 (TEF1). BLASTn analysis of these sequences showed 99 to 100% identity to Neopestalotiopsis clavispora strains (OK655673, MZ648263 for ITS, ON000362, MZ286974 fr TUB, MH423941, MK512481 for TEF1). These morphological features and molecular identification indicated that the pathogen has the same characteristics as N. clavispora. Pathogenicity was tested on ten healthy 3-month-old seedlings using the three representative strains through in vivo experiments. For each strain, the conidial suspension (106 conidia ml-1) in absorbent cotton balls (50 µl of inoculum) were inoculated onto the healthy leaves of two seedlings, while a total of two other plants were served with sterile water as a blank control. The plants were potted in a climate incubator at 28°C and a relative humidity of approximately 90%. Symptoms consistent with natural lesions were observed on the inoculated leaves after 5 days while the control plants remained healthy. The strains of N. clavispora were reisolated from the symptomatic inoculated leaves, fulfilling Koch's postulates. N. clavispora is known to cause disease in a variety of plants in China, such as Dendrobium officinale (Cao et al., 2022), Fragaria ananassa (Shi et al., 2022), and Garcinia mangostana (Qiu et al., 2019). To the best of our knowledge, this is the first report of N. clavispora causing leaf blight on A. oblongum in China. The yellowing and falling off of leaves would seriously affects the garden landscape. It is necessary to further clarify the host range of the pathogen to select appropriate landscape matching plants in future planning.

Hematological and plasma biochemical profile of two species of freshwater stingrays from the Amazon.

The evaluation of hematological and plasma biochemical parameters and the subsequent establishment of reference intervals facilitate the diagnosis of the health status of animals. This work aimed to determine the blood parameters of wild specimens of the stingrays Potamotrygon motoro and Potamotrygon orbignyi from the lower Solimões River region, Amazonas, Brazil. One hundred forty-one stingrays were captured, 92 specimens of P. motoro and 49 of P. orbignyi, of both sexes and at different stages of development. No effect of sex was observed on the blood parameters of juvenile animals for both species. P. motoro neonates presented a distinct hematological and biochemical profile, with significantly lower hematocrit values, hemoglobina, number of erythrocytes, mean corpuscular hemoglobin concentration, monocytes, plasma glucose, total proteins, albumin, and globulin. On the other hand, total cholesterol and urea levels were significantly higher in this same group compared to juveniles of the same species. Comparison between species revealed lower values of triglycerides and total cholesterol in P. orbignyi of both sexes. The results obtained are pioneering for these Amazonian species in white water environments and will serve as a basis for evaluating the health status of wild stingrays. Thus, from the analysis of the blood of the P. motoro and P. orbignyi stingrays, it was possible to observe good health conditions.

Podosphaera xanthii Causing Powdery Mildew on Salvia farinacea in Central China.

Salvia farinacea, commonly referred as mealycup sage, is a perennial herbaceous plant belonging to the Salvia genus of the Lamiaceae family. It originates from the Mediterranean region, North America, and Europe and is globally cultivated due to its appealing and captivating flowers. Moreover, mealycup sage is utilized as traditional Chinese medicinal plant for treatment of cardiovascular diseases (Li et al. 2018). In October 2023, powdery mildew-like symptoms were observed on Salvia farinacea plants cultivated in a garden located in Xinxiang City, Henan Province, China (113.93, 35.29). The leaves were covered with white and thin masses of mycelia, conidiophores and conidia of the fungus. About 100 plants were checked and 90 % were infected. There were a large number of white colonies with irregular or continuous round lesions on the adaxial and abaxial surfaces of the leaves, covering approximately 80% of the leaf area. The slightly or straight curved conidiophores (n = 30) were 46 to 145× 8 to 11 µm in size and consisted of foot cells, shorter cells and conidia. The ellipsoidal to oval conidia (n = 30), containing fibrosin bodies, were 24 to 35 × 12 to 19 µm in size and had a length/width ratio of 1.8 to 2.1. No chasmothecia were observed on leaves. These morphological features were consistent with those of Podosphaera xanthii (Braun and Cook 2012). Following the previously described method (White et al. 1990; Bradshaw et al. 2022; Zhu et al. 2022a), the sequences of ITS and Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) regions were amplified with specific primers ITS1/ITS4 (ITS1 5'-TCCGTAGGTGAACCTGCGG-3' ; ITS4 5'-TCCTCCGCTTATTGATATGC-3') and PMGAPDH1/PMGAPDH3R (PMGAPDH1 5'-GGAATGGCTATGCGTGTACC-3'; PMGAPDH3R 5'-CCCCATTCGTTGTCGTACCATG-3'), and the resulting sequences were uploaded in GenBank (Accession No. OR761885 and PP236082, respectively). BLASTn analysis showed that the sequence shared 560/565 (99%) and 272/272 (100%) homology with P. xanthii (MW301281) on Impatiens balsamina (Zhu et al. 2022b) and with P. xanthii (ON075658) on Cucumis melo (Bradshaw et al. 2022), respectively. The phylogenetic analysis clearly illustrated that the collected isolate of P. xanthii clustered in the same clade. The pathogenicity was tested according to the method previously described (Zhu et al. 2021). The fungus was inoculated onto the leaf surfaces of three healthy plants by blowing conidia from infected leaves with pressurized air. Non-inoculated plants were treated as control. Both the control and inoculated plants were separately placed in growth chambers under 60% humidity; light/dark, 16 h/8 h; and a temperature of 18°C. After a period of 12-15 days, the leaves of the inoculated plants exhibited signs of powdery mildew, whereas the control group remained unaffected. Therefore, the fungal pathogen was identified and confirmed as P. xanthii (isolate PXSF202310). Previously, P. xanthii was reported on Impatiens balsamina and S. farinacea from China and Korea (Zhu et al. 2021; Choi et al. 2022). As far as we know, this is the first documentation of P. xanthii on S. farinacea in central China. The presence of P. xanthii can lead to a deterioration in plant health and stunted growth, thereby negatively impacting both the decorative and medicinal value of S. farinacea. The recognition of P. xanthii on S. farinacea enhances our comprehension of this pathogen hosts and provides fundamental information for forthcoming disease control studies.

First Report of Agroathelia rolfsii Causing Southern Blight on Lippia (Phyla canescens) in Guangzhou, China.

Lippia (Phyla canescens) is a fast-growing, mat-forming, and prostrate perennial plant well adapted to infertile, high-saline, and drought environments (Leigh, et al. 2004). It arrived in China from Japan as a flowering ground cover in 2001 (Cai, et al. 2004). In June 2022, southern blight appeared in our nursery of the Floriculture Research Institute of Guangdong Academy of Agricultural Sciences. High temperature and damp environment are major factors for this disease. The symptoms of top-layer plants were not easily detected, but they were slightly yellowed. A yellowish-brown water-soak lesion appeared on the stems and lowest leaves exposed to soil. White mycelium appeared in the middle stage. Finally, the surface plants showed water-soak decay, and a mass of beige to black-brown rapeseed-shaped sclerotia appeared on the residue and surrounding soil; these plants died. Sclerotia and mycelia were collected from disease tissue, and after surface sterilization, sclerotia was cultured on potato dextrose agar (PDA) at 28±2°C in an incubator without light. Eight fungal isolates with similar colony morphologies were consistently isolated by purifying from different sampling areas. The isolates exhibited obvious septa and a clamp connection structure within the white mycelium. The average growth rate was 26.86±0.06 mm/day. Numerous white granular sclerotia were produced on the mycelium 6 days later. The sclerotia with a diameter of 1.24±0.07mm (n=189) gradually changed from diage to yellow to brown. A typical strain B1 was selected for further identification, targeting its 18S rRNA and LSU rRNA sequences (Yang, et al. 2011; Xue, et al. 2019). Its 18S rRNA sequence (GenBank Accession No. OR517233, 1626 bp) is 99.63% and 99.57% identical to Athelia rolfsii (AY665774, 1179bp; KC670714, 1775bp; JF819726, 1781bp). Its LSU rRNA sequence (OR539570, 757 bp) is 99.87% identical to Agroathelia rolfsii (OR526537, 904 bp). For Athelia rolfsii, a synonym of Agroathelia rolfsii, by combining the morphological characteristics and molecular identification, the isolate pathogen B1 was confirmed to be Agroathelia rolfsii (the teleomorph of Sclerotium rolfsii). To fullfill Koch's postulates, we inoculated the mycelial plugs to healthy lippia stems and leaves which has grown for one year, with PDA plugs free of mycelium as the control. All the plants were kept in a greenhouse at 28±2°C with a 14-h photoperiod and 80% relative humidity. Each treatment was repeated thrice and vaccinated with 6 points. At 7 d following inoculation, all plants inoculated with B1 showed typical symptoms, but the control group was asymptomatic, and sclerotia appeared 17d after inoculation. Using the same protocol mentioned above, pathogenic fungal was reisolated only from treated groups, but not from the control group. Chose three of the pathogens for 18S rRNA and LSU rRNA sequencing, the results showed 100% identity to B1, the same as its microstructure. There are few reports about the disease on P. canescens. Sosa (2007) investigated the pathogens on P. canescens in Argentina, 16 fungi were found but no A. rolfsii. Sclerotium rolfsii were identified on P. nodiflora or P. lanceolata (Michaux) Greene in America (Farr, et al. 1989). To our knowledge, this is the first report in China. Because this pathogen has wide-ranging hosts and causes serious damage, the results from this study will offer guidance for the prevention and treatment of this 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.

First Report of White Mold Caused by Sclerotinia sclerotiorum on Echeveria gigantea in Mexico.

Echeveria gigantea, native of Mexico (Reyes et al. 2011), holds economic importance as it is marketed as a potted plant and cut flower due to its drought-tolerant capabilities and aesthetic appeal. In September 2023, a field sampling was conducted at the Research Center in Horticulture and Native Plants (18°55'56.6" N, 98°24'01.5" W) of UPAEP University. Echeveria gigantea cv. Quilpalli plants with white mold symptoms were found in an area of 0.5 ha, with an incidence of 40% and severity of 50% on severely affected stems. The symptoms included chlorosis of older foliage, necrosis at the base of the stem, and soft rot with abundant white to gray mycelium and abundant production of irregular sclerotia resulting in wilted plants. The fungus was isolated from 30 symptomatic plants. Sclerotia were collected, sterilized in 3% NaOCl, rinsed with sterile distilled water (SDW), and plated on Potato Dextrose Agar (PDA) with sterile forceps. Subsequently, a dissecting needle was used to place fragments of mycelium directly on PDA. Plates were incubated at 23 °C in darkness. A total of 30 isolates were obtained using the hyphal-tip method, one from each diseased plant (15 isolates from sclerotia and 15 from mycelium). After 6 days, colonies had fast-growing, dense, cottony-white aerial mycelium forming irregular sclerotia of 3.67 ± 1.13 mm (n=100). Each Petri dish produced 32.47 ± 7.5 sclerotia (n=30), after 12 days. The sclerotia were initially white and gradually turned black. The isolates were tentatively identified as Sclerotinia sclerotiorum based on morphological characteristics (Saharan and Mehta 2008). Two isolates were selected for molecular identification. Genomic DNA was extracted using the CTAB protocol. The ITS region and the glyceraldehyde 3-phosphate dehydrogenase (G3PDH) gene were sequenced for two randomly selected isolates (White et al. 1990; Staats et al. 2005). The ITS and G3PDH sequences of the SsEg9 isolate were deposited in GenBank (ITS-OR816006; G3PDH-OR879212). BLAST analysis of the partial ITS (510 bp) and G3PDH (915 bp) sequences showed 100% and 99.78% similarity to S. sclerotiorum isolates (GenBank: MT101751 and MW082601). Pathogenicity was confirmed by inoculating 30 120-day-old E. gigantea cv. Quilpalli plants grown in pots with sterile soil. Ten sclerotia were deposited at the base of the stem, 10 mm below the soil surface. As control treatment, SDW was applied to 10 plants. The plants were placed in a greenhouse at 23 °C and 90% relative humidity. After 16 days, all inoculated plants displayed symptoms similar to those observed in the field. Control plants did not display any symptoms. The fungus was reisolated from the inoculated stems, fulfilling Koch's postulates. The pathogenicity tests were repeated three times. Recently S. sclerotiorum has been reported causing white mold on cabbage in the state of Puebla, Mexico (Terrones-Salgado et al. 2023). To the best of our knowledge, this is the first report of S. sclerotiorum causing white mold on E. gigantea in Mexico. Information about diseases affecting this plant is very limited, so this research is crucial for designing integrated management strategies and preventing spread to other production areas.

First Report of Stem and Foliage Blight of Chrysanthemum morifolium cv. Fubaiju Caused by Stagonosporopsis chrysanthemi in China.

Chrysanthemum (Chrysanthemum morifolium cv. Fubaiju) is used as medicinal herb (Chen et al. 2020). In October 2021, a leaf spot disease was observed on leaves of C. morifolium in Huanggang, Hubei province. Disease incidence was approximately 40%. Leaf lesions manifested as necrotic spots, coalesced, and expanded to form brown-black spots, leading to wilting of the leaves. On stems, the lesions manifested as dark brown necrotic spots. To identify the pathogen, 29 pieces (5 × 5 mm) from lesion margins were surface sterilized in 1% NaOCl and rinsed three times with sterile water. The pieces were transferred onto potato dextrose agar (PDA) for incubation at 25℃ for 3 d in the dark. Fifteen fungal colonies were successfully isolated. The colony morphology with flat wavy edge, sparse aerial mycelia, and surface olivaceous black were observed at 7 days post incubation. Subglobular pycnidia were brown with a short beak, and pycnidia diameters were thick (212 to 265 × 189 to 363 µm, n = 20). Ovoid conidia were aseptate and hyaline, conidia diameters were thick (4.0 to 9.8 × 1.8 to 4.7 µm, n = 100). The morphological characters of these isolates were consistent with those of Stagonosporopsis chrysanthemi (Zhao et al. 2021). Pure culture of representative HGNU2021-18 isolated from the diseased leaves subjected to molecular identification. Sequences of the rDNA internal transcribed spacer (ITS) region, 28S large subunit ribosomal RNA (LSU), ß-tubulin (TUB2), actin (ACT), and partial RNA polymerase II largest subunit (RPB2) genes were amplified from genomic DNA of isolate HGNU2021-18 using the following primer pairs: ITS1/ITS4 (White et al. 1990), LR0R/LR5 (Rehner et al. 1994), Btub2Fd/Btub4Rd (Woudenberg et al. 2009), ACT512F/ACT783R (Carbone et al.1999), and RPB2-5F2 (Sung et al. 2007)/fRPB2-7cR (Liu et al. 1999), respectively. The PCR products were purified and then sequenced by Sangon Biotech (China). Nucleotide sequences of ITS (544 bp, OM346748), LSU (905 bp, OM758418), TUB2 (563 bp, OM945724), ACT (294 bp, OM793715), and RPB2 (957 bp, OM793716) amplified from the isolate HGNU2021-18 were subjected to BLASTn analysis. The results showed that ITS, LSU, TUB2, ACT, and RPB2 shared 100.00%, 99.45%, 99.20%, 100.00%, and 100.00% sequence identity to the five published sequences (MW810272.1, MH869953.1, MW815129.1, JN251973.1, and MT018012.1, respectively) of the S. chrysanthemi isolate CBS 500.63. Phylogenetic analysis of the multilocus sequences of ITS, LSU, RPB2, ACT, and TUB2 belonging to different Stagonosporopsis species was performed in MEGA 7.0 (Chen et al. 2015). Isolate HGNU2021-18 was placed in a clade with S. chrysanthemi with 99% bootstrap support. Thus, the results of morphological and molecular analyses indicated that the disease symptoms on chrysanthemum plants were caused by S. chrysanthemi. Under conditions of 25°C and 85% relative humidity, pathogenicity test was performed on 2-month-old healthy plants using isolate HGNU2021-18. The leaves were inoculated with 5 mm diameter mycelial plugs or with sterile agar plugs (control). Six plants were used in each treatment. Disease symptoms were observed on treated plants at 2 weeks post inoculation which were those previously observed in the field, while the control plants remained symptomless. The pathogen was re-isolated from the diseased plants, and S. chrysanthemi was confirmed as the causal pathogen. This is the first report of S. chrysanthemi causing stem and foliage blight of chrysanthemum in China.

First Report of Powdery Mildew Caused by Podosphaera xanthii on Youngia japonica subsp. elstonii in Hainan Province, China.

Youngia japonica (L.) DC. is a polymorphic annual herb of the Asteraceae family. Although this plant originated in Asia, it is now world-widely distributed. In China, Y. japonica is used for edible or folk medicine to treat viral infections and various kinds of inflammation (Yu et al. 2021). As a traditional Chinese medicinal herb, Y. japonica used for the treatment of inflammatory diseases, such as angina, leucorrhea, mastitis, conjunctivitis, and rheumatoid arthritis (Chen et al. 2006). During the spring of 2023, powdery mildew symptoms were observed on 60% of Y. japonica subsp. elstonii plants in a greenhouse on the Hainan Medical University campus (19° 58' 53″ N; 110° 19' 47″ E) in Haikou, Hainan Province, China. Powdery mildew colonies covered the leaf surfaces and stems of affected plants, causing discoloration and defoliation. Mycelia were superficial and hyphal appressoria were nipple-shaped. Conidiophores (n =30) were unbranched, cylindrical, 99 to 166 × 11 to 16 µm, and produced three to five immature conidia in chains with a crenate outline. Foot cells (n =30) were cylindrical, straight or sometimes curved at the base, and 35 to 61 µm long. Conidia (n =100) were ellipsoid-ovoid to doliiform, 21 to 40 ×13 to 21 µm (length/width ratio = 1.4 to 2.3), with well-developed fibrosin bodies, and produced germ tubes from the lateral position. Based on these morphological characteristics, the pathogen was provisionally identified as Podosphaera xanthii (Braun and Cook 2012). The teleomorph was not observed. A specimen was deposited in the Hainan Medical University Plant Pathology Herbarium as HMYJ-23. To confirm the genus identification and ascertain a putative species, genomic DNA was extracted from mycelium, conidiophores, and conidia using a fungal DNA kit (Omega Bio-Tek, USA). The rDNA internal transcribed spacer (ITS) region was amplified with primers ITS1/ITS4 (White et al. 1990) and sequenced directly. The resulting 575-bp sequence was deposited in GenBank (accession no. OR229712). A BLASTn search in GenBank of this sequence showed 99% similarity with the ITS sequences of P. xanthii isolates from China (MT260063, OP765400, MW422608, and MT739423), Thailand (LC270778, LC270779, and LC270780), and Argentina (AB525914). Additionally, the 613-bp 28S rDNA region was amplified using the primer pairs NL1 and NL4 (O'Donnell 1993; accession no. OR240257). This region shared 100% similarity with P. xanthii isolates (MK357436, LC371333, LC270780, OP765401, and AB936277) as well. To confirm pathogenicity, five healthy potted plants of Y. japonica subsp. elstonii were inoculated by gently pressing a powdery mildew-infected leaf onto the young leaves. Five non-inoculated plants served as controls. All plants were maintained in a greenhouse at 24 to 30°C, 70% relative humidity, with a 16-h photoperiod. After 7 days, inoculated leaves showed powdery mildew symptoms whereas no symptoms were observed on control plants. The fungal colonies observed on inoculated plants were morphologically identical to those found on the originally infected leaves collected from Hainan Province. Based on the morphological characteristics and molecular identification, the fungus was identified as P. xanthii. To our knowledge, this is the first record of P. xanthii infecting Y. japonica subsp. elstonii in Hainan province, China. We are concerned that the pathogen could become a threat to the widespread planting of Y. japonica subsp. elstonii in the future.

First Report of Powdery Mildew Caused by Podosphaera xanthii on Sphagneticola trilobata in Hainan Province, China.

Sphagneticola trilobata (L.) Pruski is a perennial creeping herb of the Asteraceae family, which is native to South America. It was introduced into Southern China as a groundcover in the 1970s (Zhang et al. 2023). Now it is mainly used for folk medicine to treat various kinds of inflammatory, incuding joint pain, rheumatic diseases, arthritis, in addition to treating persistent wounds, ulcers, and edemas (Gonçalves et al. 2022). In February and November 2023, powdery mildew symptoms were observed on 60% of S. trilobata plants on the Hainan Medical University campus (19° 58' 53″ N; 110° 19' 47″ E) in Haikou, Hainan Province, China. Powdery mildew colonies covered the leaf surfaces and stems of affected plants, causing discoloration and defoliation. Mycelia were superficial and hyphal appressoria were nipple-shaped. Conidiophores (n =30) were unbranched, cylindrical, 74 to 161 × 10 to 14 µm, and produced three to five immature conidia in chains with a crenate outline. Foot cells (n =30) were cylindrical, straight or sometimes curved at the base, and 27 to 56 µm long. Conidia (n =100) were ellipsoid-ovoid to doliiform, 17 to 30 ×14 to 28 µm (length/width ratio = 1.1 to 1.9), with well-developed fibrosin bodies, and produced germ tubes from the lateral position. Based on these morphological characteristics, the pathogen was provisionally identified as Podosphaera xanthii (Braun and Cook 2012). The teleomorph was not observed. A specimen was deposited in the Hainan Medical University Plant Pathology Herbarium as HMST-23. To confirm the genus identification and ascertain a putative species, genomic DNA was extracted from mycelium, conidiophores, and conidia using a fungal DNA kit (Omega Bio-Tek, USA). The rDNA internal transcribed spacer (ITS) region was amplified with primers ITS1/ITS4 (White et al. 1990) and sequenced directly. The resulting 577-bp sequence was deposited in GenBank (accession no. OR784549). A BLASTn search in GenBank of this sequence showed 100% similarity with the ITS sequences of P. xanthii isolates from China (MT260063, MN203658, OP765400, and MT739423), Thailand (LC270780), and Vietnam (KM260731, KM260730, and KR779870). Additionally, the 28S rDNA region was amplified using the primer pairs NL1 and NL4 (O´Donnell 1993; accession no. OR784550). This region shared 100% similarity with P. xanthii isolates (LC371334, LC270782, AB936277, and OP765401) as well. Powdery mildew from Hainan sample belonged to the P. xanthii group with strong bootstrap values support 99% in maximum likelihood phylogenetic tree based on ITS and 28S gene sequences. To confirm pathogenicity, five healthy potted plants of S. trilobata were inoculated by gently pressing a powdery mildew-infected leaf onto 15 young leaves. Five non-inoculated plants served as controls. All plants were maintained in a greenhouse at 24 to 30°C, 70% relative humidity, with a 16-h photoperiod. After 7 days, inoculated leaves showed powdery mildew symptoms whereas no symptoms were observed on control plants. The fungal colonies observed on inoculated plants were morphologically identical to those found on the originally infected leaves collected from Hainan Province. Based on the morphological characteristics and molecular identification, the fungus was identified as P. xanthii. In different countries and regions, P. xanthii has been previously reported on S. trilobata in Taiwan (Yeh et al. 2021). To our knowledge, this is the first record of P. xanthii infecting S. trilobata in Hainan Province, China. S. trilobata is often planted as an ornamental plant on both sides of the road, and we are concerned that it may serve as a new host, spreading this pathogen to other economic crops.

Transversotrema hafniensis n. sp. infection in Poecilia reticulata by cercariae released from Melanoides tuberculata in Denmark.

BACKGROUND: Exotic and ornamental fish are highly popular companion animals resulting in a significant transcontinental trade of fish, invertebrates and aquatic plants. A major issue is the diseases associated with these organisms, as they have a major impact on health of the fish in both public and private household aquaria. A secondary issue is the trade with these products, which potentially may expand the distribution area and spread a range of diseases to new habitats. RESULTS: We here describe how Poecilia reticulata (guppy), produced in a private household aquarium, were invaded by cercariae of an exotic trematode released by imported Melanoides tuberculata snails. The fish presented with severe clinical signs (tremor, flashing, scraping of body against objects). A standard parasitological examination and morphometric identification showed scale pocket infections with a digenean trematode species within the genus Transversotrema. Molecular identification by PCR, sequencing and phylogenetic analyses of a 2646 bp sequence encoding ribosomal RNA (partial 18 S, ITS1, 5.8 S, ITS2, partial 28 S) was performed. The 1107 bp sequence of mitochondrial DNA (cox1) showed that the parasite differed from previously described Transversotrema species in M. tuberculata. Morphometrics of adult and larval specimens of this isolate also differed from previously described freshwater species within the genus. The new species was described and is named after Copenhagen, for its geographic origin. CONCLUSIONS: The genus Transversotrema comprises a range of species, adapted to a microhabitat in scalepockets of teleosts. A combination of morphological and molecular characterization techniques has been shown to provide a good differentiation between species. The fish were not purchased from a pet shop but produced in the home aquarium. This indicated that an infection pressure existed in the aquarium, where the source of infection was found to be exotic intermediate host snails M. tuberculata, which originally were imported and purchased from a pet shop. The potential spread of fish diseases associated with trade of fish and snails to new geographic regions, where climate conditions are favourable, is discussed.

Molecular-Phylogenetic Characterization of Xanthomonas hortorum pv. pelargonii Strains Causing Leaf Spot of Geraniums in Iran.

Bacterial blight and leaf spot of geraniums is a destructive disease of cultivated Pelargonium species around the world. During 2020-2021, surveys were conducted in seven geranium-growing provinces of Iran to monitor the status of bacterial blight and leaf spot disease. The disease was observed in six surveyed provinces varying in the extent of occurrence and severity. Twenty-two Gram-negative pale-yellow bacterial strains resembling members of Xanthomonas were isolated from symptomatic leaves and stems. Pathogenicity and host range assays showed that the bacterial strains were pathogenic on Pelargonium grandiflorum, P. graveolens, P. peltatum, and P. zonale. All strains were positive for PCR test using the primer pair XcpM1/XcpM2 which is specific for Xanthomonas hortorum pv. pelargonii. Phylogenetic analysis using the sequences of gyrB and lepA genes showed that the 22 strains clustered in a clade among the sequences of X. hortorum pv. pelargonii strains retrieved from the GenBank, while distinct from the other pathovars of X. hortorum. BOX-PCR-based fingerprinting using BOX-A1R primer revealed that the strains isolated in this study were grouped into two clusters while no distinct correlation was observed between the host/area of isolation and BOX-PCR fingerprinting. None of the strains obtained in this study nor reference strain of the pathogen did produce bacteriocin against each other. Results obtained in this study shed light on the geographic distribution, taxonomic status and host range of the bacterial blight and leaf spot pathogen of geraniums in Iran, paving the path of further research on disease management.

First Report of Powdery Mildew Caused by Podosphaera xanthii on Acalypha indica in China.

Acalypha indica L. is an annual erect herb of the Euphorbiaceae family. This plant is found widely in the tropics and parts of Africa and Asia (Chakraborty et al. 2023). In China, A. indica is a vegetable and also used as a folk medicine due to its antipyretic and hemostatic, antibacterial and anti-inflammatory properties. In February 2022 and 2023, powdery mildew symptoms were observed on 70% of A. indica plants on the Hainan Medical University campus (19° 58' 53″ N; 110° 19' 47″ E) in Haikou, Hainan Province, China. Powdery mildew colonies covered the leaf surfaces and stems of affected plants, causing discoloration and defoliation. Mycelia were superficial and hyphal appressoria were nipple-shaped. Conidiophores (n =30) were unbranched, cylindrical, 66 to 150 × 10 to 15 µm, and produced three to five immature conidia in chains with a crenate outline. Foot cells (n =30) were cylindrical, straight or sometimes curved at the base, and 31 to 59 µm long. Conidia (n =100) were ellipsoid-ovoid to doliiform, 20 to 33 ×12 to 20 µm (length/width ratio = 1.3 to 2.4), with well-developed fibrosin bodies, and produced germ tubes from the lateral position. Based on these morphological characteristics, the pathogen was provisionally identified as Podosphaera xanthii (Braun and Cook 2012). The teleomorph was not observed. A specimen was deposited in the Hainan Medical University Plant Pathology Herbarium as HMAI-23. To confirm the genus identification and ascertain a putative species, genomic DNA was extracted from mycelium, conidiophores, and conidia using a fungal DNA kit (Omega Bio-Tek, USA). The rDNA internal transcribed spacer (ITS) region was amplified with primers ITS1/ITS4 (White et al. 1990) and sequenced directly. The resulting 575-bp sequence was deposited in GenBank (accession no. OR775733). A BLASTn search in GenBank of this sequence showed 99% similarity with the ITS sequences of P. xanthii on plants of Fabaceae, Malvaceae and Cucurbitaceae family from China (MH143485, MT242593, MK439611 and MH143483), Thailand (LC270779 and LC270778), Korea (MG754404), Vietnam (KM260704), and Puerto Rico (OP882310). Additionally, the 28S rDNA region was amplified using the primer pairs NL1 and NL4 (O´Donnell 1993; accession no. OR784547). This region shared 99% similarity with P. xanthii isolates (LC371333, LC270780, AB936277, and OP765401) as well. To confirm pathogenicity, five healthy potted plants of A. indica were inoculated by gently pressing a powdery mildew-infected leaf onto 15 young leaves. Five non-inoculated plants served as controls. All plants were maintained in a greenhouse at 24 to 30°C, 70% relative humidity, with a 16-h photoperiod. After 7 days, inoculated leaves showed powdery mildew symptoms whereas no symptoms were observed on control plants. The fungal colonies observed on inoculated plants were morphologically identical to those found on the originally infected leaves collected from Hainan Province. Based on the morphological characteristics and molecular identification, the fungus was identified as P. xanthii. In different countries and regions, P. xanthii has been previously reported on A. indica from Sudan and India (Amano 1986). To our knowledge, this is the first record of P. xanthii infecting A. indica in China. We are concerned that the pathogen could become a threat to the widespread planting of A. indica in the future.

First report of leaf blight caused by Stemphylium lycopersici on Salvia splendens in China.

Salvia splendens is a popular ornamental plant in China with extensive potentials, including value in traditional Chinese medicine and in environmental restoration function (Li et al. 2008). In September 2019, leaf blight disease was observed on road side plants of S. splendens in Bayi park, Nanchang city, Jiangxi province, China. The typical symptoms appeared as irregular necrotic spots or leaf blight, accompanied by extensive scorch necrosis or ultimately defoliation. Small segments cut from diseased leaves were surface sterilized in a 2% sodium hypochlorite solution for 2 min and rinsed three times with sterile distilled water. Then, the samples were placed on potato dextrose agar (PDA) plates incubated at 25°C in darkness. Pure cultures were obtained by the hyphal tip method. Morphologically, all 11 colonies were identical to each other on PDA. Two strains, YZU 191468 and YZU 191481, were selected for further study and deposited in the Fungal Herbarium of Yangtze University (YZU), Jingzhou, Hubei, China. The 7-day-old colonies were circular, 53 to 56 mm in diameter, and consisted of white mycelium with a buff margin, and were cinnamon colored in the center of the reverse side. To examine conidial morphology, the mycelium was transferred onto potato carrot agar (PCA) and incubated at 23°C with a period of 8 h light/16 h dark for 7 days. Conidia were normally solitary or two in a chain, ellipsoid or long ellipsoid, beakless, 10 to 23×30 to 60 µm in size (n=50). Based on morphology, the isolates were consistent with Stemphylium lycopersici (Yamamoto 1960). To confirm the identification, genomic DNA was extracted from both isolates and used to amplify the internal transcribed spacer rDNA region (ITS), glyceraldehydes-3-phosphate dehydrogenase (GAPDH) and calmodulin (CAL) genes with primer pairs ITS5/ITS4, gpd1/gpd2, and CALDF1/CALDR2, respectively (Woudenberg et al. 2017). Sequences were deposited in GenBank with accession numbers OP564983 and OP564984 (ITS), OP892529 and OP892530 (GAPDH), OP584970 and OP584971 (CAL). A neighbor-joining tree was constructed with Mega 7.0 based on the combined dataset with 1,000 bootstrap replicates. The resulting phylogenetic tree showed that the strains from S. splendens clustered with S. lycopersici (CBS 122639 and CBS 124980) supported with 100% bootstrap values. The molecular analyses confirmed that the species causing leaf blight symptoms was S. lycopersici. To test pathogenicity, healthy leaves of S. splendens were surface sterilized and inoculated by mycelium blocks (6 mm in diameter) and spore suspension (1×106 spore/mL) of representative strains YZU 191468 and YZU 191481, respectively. Controls were inoculated with blocks of PDA and sterile water. Each strain was inoculated on three leaves of a plant. One clean plant was used as control. The test was replicated three times. After inoculation, the plants were covered with plastic bags and incubated in a greenhouse (25℃, 80 % relative humidity, 8 h light/16 h dark). After 5 days, the inoculated leaves exhibited dark brown spots with white mycelium, followed by withering of necrotic tissues. There were no symptoms observed on the controls. The fungal isolates inoculated leaves had the same morphological characteristics as the strains used for inoculation. S. lycopersici has been found on eggplant and Zinnia elegans in China (He et al. 2019; Yang et al. 2017). To the best of our knowledge, this is the first report of S. lycopersici causing leaf blight on S. splendens in China. This finding offers a new reference for the management and control of S. splendens leaf diseases in China.

First Report of Anthracnose on Euonymus japonicus Thunb. Caused by a Provisionally Novel Species of Colletotrichum in the Magnum Complex in South Korea.

Euonymus japonicus Thunb., also known as the evergreen spindle tree, is an evergreen tree, which is widely planted as a hedge plant along streets in South Korea. In April 2022, severe anthracnose symptoms were observed on the leaves of this tree in Jangsu in the Jeonbuk Province of the country (35°43'49.44″N, 127°34'53.7″E). About 80% of the leaves of each affected tree within a 0.03-ha area showed incidence of the disease on approximately 30 trees were planted along the roadside (~30 m). These symptoms typically included circular or irregularly shaped whitish-gray lesions with a diameter of 2.0 to 3.0 cm. In cases where some leaves were severely affected, larger blotches formed. To isolate the pathogen, about ten leaves showing anthracnose symptoms on each tree were randomly selected and brought to the laboratory. Fungal isolations were made from acervuli filled with conidial masses on infected evergreen tissues, followed by plating onto 2% potato dextrose agar (PDA) as well as incubated at 25℃. On the PDA, colonies were circular, raised, green-grey or dark grey, and had a distinct white margin. The conidia were single-celled, transparent, cylindrical with rounded ends, had smooth walls, with a length ranging from 12 µm to 16.7 µm and a width raging from 4 µm to 6.5 µm (av. = 14.1 X 5.0 µm, n=40). Of those that were successfully recovered with approximately 90% frequency, two monoconidial isolates were deposited to the culture collection at Chungnam National University in South Korea (Accession number: CDH059-060). To ensure the identity of the fungus, genomic DNAs were extracted from the selected isolates, CDH059-060, and were sequenced. This was achieved based on partial sequences of the internal transcribed spacer (ITS), actin and beta-tubulin (TUB2) gene regions which were amplified using ITS1F / ITS4 (Gardes and Bruns 1993; White et al. 1990), ACT-512F / ACT-783R (Carbone and Kohn 1999), and T1 / Bt2b (O'Donnell and Cigelnik 1997; Glass and Donaldson 1995) primer pairs, respectively. The resulting sequences were deposited to GenBank (OR984424-425) for ITS, (OR996289-290) for actin, and (OR996291-292) for TUB2. For a phylogenetic analysis, sequences from different gene regions (ITS, actin and TUB2) retrieved from GenBank were aligned, concatenated, and analyzed as a single dataset based on a maximum likelihood analysis. The phylogenetic result revealed that the fungus isolated in this study was positioned in a clearly distinct lineage, provisionally representing an undetermined species of Colletotrichum, which is most closely related to Colletotrichum liaoningense (Y.Z. Diao, C. Zhang, L. Cai & X.L. Liu, CGMCC3.17616 (KP890104 for ITS, KP890097 for actin, and KP890111 for TUB, Diao et al. 2017). Sequence comparisons revealed that this pathogen differed from C. liaoningense at 20 of 494 characters (∼4.0%) in the ITS and 2 of 251 (∼1.0%) in the actin sequences. For pathogenicity tests, three seedlings of E. japonicus were used. The leaves for each tree were treated with 10 ml of a conidial suspension by spraying (1x106 conidia ml-1 of the isolate, CDH059), while the three seedlings were treated with distilled water as control. After sprayed, the treated areas were sealed with plastic bags for a day to maintain humidity. Anthracnose symptoms identical to those observed in the field appeared seven days after inoculations, while no symptoms were observed in the control. Re-isolations were successfully achieved from the treatments, fulfilling Koch's postulates. Anthracnose associated with the provisionally novel species of Colletotrichum sp. on E. japonicus has not been recorded elsewhere, and in this regard, this is the first report of anthracnose caused by Colletotrichum sp. on E. japonicus in Korea. To effectively control the disease, more attention should be paid to the host range of the pathogen and other regions where the disease caused by the pathogen might occur in the country.