Soil Line Research Articles

Normalized Temperature Drought Index (NTDI) for Soil Moisture Monitoring Using MODIS and Landsat-8 Data

As the fundamental regulator of energy exchange in the vegetation–soil–atmosphere circulation system, soil moisture is a key parameter for drought monitoring and is indispensable to the land surface hydrological processes. In order to overcome the constraints of the Perpendicular Drought Index, PDI (performs poorly over the fields with dense vegetation and hard to construct the soil line), and the Temperature Vegetation Drought Index, TVDI (requires similar atmospheric forcing and large enough dimension of mapping area), in soil moisture monitoring, a new drought index (Normalized Temperature Drought Index, NTDI) is proposed to explore the spatiotemporal changes of soil moisture by substituting red and near-infrared reflectances with vegetation index and normalized land surface temperature on the basis of the PDI framework. Victoria, Australia, was selected as the study area as it experiences many severe droughts and has been affected for more than ten years. Time series of satellite-based data were applied to evaluate the effectiveness and applicability of the NTDI at the regional scale. Results indicated that the expression of the soil line representing the water condition of the bare soil is easier to obtain in the new trapezoid framework and has good fits with the coefficients of determination (R2) of more than 0.8. Compared with PDI, TVDI and Modified PDI (MPDI) at the cropping sites, NTDI exhibits a relatively better performance in soil moisture monitoring for most days where the R2 achieved can reach to more than 0.7 on DOY 242, 254 and 272. Meanwhile, spatial–temporal mappings of the four drought indices from satellite data were conducted, and the NTDI presented the slightly seasonal variation and effectively described the real spatial characteristics of regional drought. Overall, the NTDI seems to a viable approach and can provide insight into spatial and temporal soil moisture monitoring at different scales.

Nondestructive Evaluation of Cotton Genotypes for Resistance to Reniform Nematode

Identification of new sources of reniform nematode (Rotylenchulus reniformis Linford and Oliveira) resistance in cotton is critical to expanding host plant resistance to manage this important pathogen. Phenotyping plants in early breeding generations without destructive sampling would be useful for introgression of nematode resistance from exotic germplasm resources; therefore, a rapid, nondestructive method was developed to assess host plant resistance to the reniform nematode based on the number of females infecting the roots. In one set of experiments, the root system was cut off at 0, 1, 2.5, or 5 cm below the soil line and used to assess the number of females infecting this portion of the root system. Resistance could be accurately determined while leaving up to 5 cm of roots with the shoot. In a second set of experiments, the rate of plant recovery and reproductive development was evaluated using a combination of root retention (0, 1, 2.5 cm, or all root) and shoot retention (leaves at top two nodes, leaves at bottom two nodes, no leaves, all leaves) treatments. Plants more rapidly recovered using a treatment combination in which the top leaves and 2.5 cm roots were kept. This combination performed similarly to plants with neither shoots nor roots modified and was harvested 20 days sooner compared to some other treatment combinations.

First Report of Blight Caused by Rhizoctonia solani AG4-HGI on Pinellia ternata in Guizhou, China.

Pinellia ternata (Thumb.) has been used for over 1000 years as a traditional Chinese herbal medicine (Ying et al. 2007) and is widely cultivated in Guizhou Province, China. It is cultivated over an area of 2000 hectares, and is of great value to underdeveloped regions. In April 2020, blight was observed in a field of P. ternatain Bijie County, Guizhou Province, China (27°30'N, 105°28'E). Around 20 hectares of P. ternata were surveyed and the disease incidence ranged from 10 to 12%. The disease symptoms included light brown lesions formed on the stems near the soil line. The color of the lesions became darker, and the stems became constricted around the lesions and broke, associated with the leaf blight. To identify the causal agent of this blight, 22 diseased plants (about 30 d-old） were collected, the margins of the infected parts were cut into small pieces (5 mm) and surface disinfested with 1% NaOCl for 10 min, 75% ethanol for 30 s, and rinsed three times in sterile distilled water. The pieces were blotted dry with sterile filter paper and placed on potato dextrose agar (PDA, Hopebio, China), incubated at 28℃ in darkness until fungal hyphae growth was visible. Sixteen cultures with different morphologies were recovered from the samples. When representative isolates of each culture type were inoculated onto plants, one produced similar blight symptoms. The representative isolate was called CD-1. The colony color was first white but turned light brown after grown on PDA for 6-7 d, and produced dark brown sclerotia. The hyphae were branched at right angles, with a slight constriction at the base of the branches and a septum near the junction where the branch separates from the main hyphae. Hyphal cells were stained with 0.5% Safranin O and 3% KOH and were observed to be multinucleate. These morphological features indicated that CD-1 likely is R. solani (Sneh et al. 1991). When paired with tester strains AG1 and AG4(provided by Dr. Genhua Yang, Yunnan Agricultural University). CD-1 showed anastomosis with isolate of AG4 (Fenille et al. 2002). Genomic DNA was extracted from the isolate (Thangaraj et al. 2018) using a fungal genomic DNA extraction kit (Tiangen, China). The internal transcribed spacer (ITS) regions were amplified using the primers ITS1/ITS4 (White et al. 1990). A 535 bp fragment was amplified that showed 99% coverage and 99.4% identity with an isolate of R. solani AG4-HGI (GenBank: HG934417). The gene sequence was deposited in GenBank as accession #OL518945. Pathogenicity tests were performed using 30 d-old plants planted in sterilized soil in pots. Cut mycelial discs (diameter 6 mm) from 3-day-old PDA cultures and placed beside stems of 21 healthy plants. Nine plants treated with agar plugs were control samples. Inoculated plants were maintained at 24 ± 5℃ in a green house and watered every two days with sterilized water. Typical blight symptoms developed on the inoculated plants at d 3-5 post inoculation, whereas the control plants remained healthy. The experiments were repeated three times, and the isolates was re-isolated from the inoculated plants and identified as R. solaniAG4 by morphological features and molecular method. R. solani has been reported to cause blight of many plants such as coffee (Ren et al. 2018) and sesame (Cochran et al. 2018). To the best of our knowledge, this is the first report of R. solani AG4-HGI causing disease on P. ternate, both in China and worldwide. This finding suggests that this pathogen may cause a threat to cultivation and production of P. terenata.

Temporal stability and spatial dependency of pixel-specific red-NIR soil lines from multi-temporal satellite remote sensing imagery

ABSTRACT The temporal stability and spatial dependence of pixel-specific red-NIR soil line coefficients were studied using analyses of geostatistics and covariance. We used time series of MODIS 8-day composite reflectance data from 2000 to 2019 of bare soil surfaces near Kabul, Afghanistan. The goal was to explore the feasibility of reducing the soil background signal in red-NIR vegetation indices (VIs). The results were that 1) Red-NIR pixel-specific soil line coefficients can be obtained using multi-temporal remote sensing imagery; 2) pixel-specific Red-NIR soil line coefficients derived from multi-temporal remote sensing imagery exhibit clear spatial dependency related to the soil line coefficients of soils sampled in the field, and 3) Red-NIR pixel-specific soil line coefficients were stable for years, but showed instability for periods longer than 16 years. These results imply that A) some changes in soil surface characteristics can be deduced using long-term observations, B) pixel-specific Red-NIR soil lines at any location can be estimated by kriging interpolation. Consequently, soil line coefficients derived from multi-temporal satellite observations can be used to derive information about vegetation, such as the germination stages pixel-by-pixel or above-ground biomass in sparsely vegetated lands. The derived values should be valid over wide areas, minimizing soil background effects.

First Report of Phytophthora inundata Associated with Decline and Death of Walnut (Juglans regia L.) in Italy.

Persian walnut (Juglans regia L.) is an important nut crop in Italy. In recent years, incidence of walnut decline and death has increased in many Italian commercial orchards. In early summer 2020, we observed a serious decline in approximately 5% of trees in a waterlogged area of a Veneto-region walnut orchard (J. regia, cv Lara). Symptoms included extensive foliar wilt and canopy decline associated with collar and root rot. Symptomatic tissues excised from larger roots of affected trees were surface disinfested for 1 min in a 1% NaOCl solution, rinsed for 5 min in sterile distilled water, and placed onto P5ARPH selective medium (Jeffers and Martin 1986). A Phytophthora-like organism was consistently isolated. Pure cultures were obtained by single-hyphal transfers onto potato dextrose agar (PDA). Isolates were identified as Phytophthora inundata based on morphological characteristics (Brasier et al. 2003), sequences of internal transcribed spacer (ITS) amplicons from universal primers ITS6 (Cooke et al. 2000) and ITS4 (White et al. 1990) and sequences of cytochrome c oxidase, subunit II (Cox II) from Fm75 and Fm78 primer pair (Martin and Tooley 2003). On carrot agar (CA), colonies had a characteristic stellate to broad lobed patterns. On this medium, optimal growth was at 28-30 °C (7,3 mm/day) and the upper temperature limit for mycelial growth was 37°C. Mycelial disks of isolate CREADC-Om306, grown on CA, were floated in Petri plates with soil extract solution and incubated under continuous fluorescent light at room temperature (25+/-2 °C). Within 48 to 72 h, sporangia were produced that were persistent, non-papillate, ovoid or ovoid-obpyriform, measuring 55.0 to 80.7 (length) x 41.3 to 65.2 (width) µm (averages 64.3+/-10.2 x 47.9+/-9.7 µm). Oospores and chlamydospores were absent. BLAST analysis of the amplicons from CREADC-Om306 revealed ITS sequences (854-bp; GenBank accession no. OK342200) and Cox II sequences (568-bp; GenBank accession no. OK349677) that shared 100% identity with published P. inundata sequences available in GenBank (acc. n. AF266791 for ITS; MT458994 for Cox II). Pathogenicity tests were conducted in the greenhouse on six 2-year-old walnut (J. regia, cv Lara) plants. Four of the plants were inoculated with CREADC-Om306 on two opposite sides of each plant's stem at 1-2 cm above soil line. A cork borer was used to remove a 5-mm disk of bark that was replaced by a 5-mm diameter mycelial plug from 10-day-old cultures of the pathogen on PDA. Two control plants were treated in the same way except the bark wounds were inoculated with sterile PDA plugs. Plants were kept in greenhouse at 24 ± 2°C. After 3 months, lesions had developed from all points of inoculation with. P. inundata (mean lesion length 55,25+/-6,22 mm) and the pathogen was reisolated from the lesion margins of all inoculated plants. The control plants remained symptomless and did not yield the pathogen. P. inundata is widely distributed across the world as a plant pathogen on several native as well as horticultural crops, especially in riparian or other areas subject to flooding or waterlogging. This report is the first to document P. inundata as a pathogen on Persian walnut and adds it to the diverse list of known susceptible perennial native, ornamental, and agricultural hosts of this organism. In addition to P. inundata, which belongs to the major Phytophthora ITS Clade 6, other members of the clade including P. megasperma (Belisario et al. 2012) and P. gonapodyides (Belisario et al. 2016) have been described as walnut pathogens.

First report of Diplodia corticola causing dieback of white oak (Quercus alba) in Tennessee.

Diplodia corticola is a fungal pathogen causing oak dieback in Quercus (oak) spp. in parts of North America, northern Africa, and Europe (Ferreira et al., 2021; Smahi et al., 2017; Tsopelas et al., 2018). In August 2021, a single mature white oak (Q. alba) exhibiting wilt symptoms, vascular discoloration, and interveinal chlorosis was observed in Cove Lake State Park in Campbell County, Tennessee, U.S.A. Small sections of phloem tissues were cut from the margins of discolored vasculature of a single wilt symptomatic branch with a sterile scalpel and surface sterilized following Parra et al. (2020). Surface sterilized wood chips were plated onto potato dextrose agar amended with antibiotics (PDA++) following Gazis et al. (2018). Three days after plating, we recovered a single fungal isolate from wood chips that when grown in ½ PDA resembled D. corticola, having irregular margins and white aerial mycelia that progressively turned greyish-black 15 days after sub-culturing (Alves et al., 2004). Total genomic DNA was extracted from the isolate following Gazis et al. (2018). The internal transcribed spacer (ITS) was then amplified using the ITS1 and ITS4 primers and the subsequent PCR product was sequenced. Resulting reads were assembled into a consensus sequence and identity was assigned using BLAST on the NCBI nucleotide database. The assembled sequence (accession OM716006) had a 100% identity match with D. corticola type culture CBS 112549 (accession NR_111152). To complete Koch's postulates and identify potential host range, 5 red oaks (Q. rubra; 2-3 yrs old; caliper 14.7 ± 2 mm) and 5 white oaks (Q. alba; 2-3 yrs old; caliper 22.8 ± 2.3 mm) were inoculated with D. corticola (isolate DC_2.5). Trees were inoculated 15 cm above the soil line in a greenhouse with a 3 mm diameter plug of a 10-day old culture of D. corticola grown on PDA following Sitz et al. (2017). As a negative control, 5 red and 5 white oaks were inoculated with a 3 mm diameter plug of PDA. For each species, trees were sampled when seepage was observed from D. corticola inoculated sites (15 days post-inoculation for red and white oaks). At time of sampling, bark adjacent to inoculation sites on each tree was removed and cankers were photographed. Using a sterile scalpel, four wood chips were cut from canker margins and placed onto PDA++. For all trees, canker areas were measured using ImageJ software (Rasband, 2012). Recovered isolate identities were confirmed by extracting total genomic DNA as described above (Gazis et al. 2018) and PCR amplification of the ITS, large ribosomal subunit (LSU), and elongation factor 1-α (ef1-α) following (Ferreira et al., 2021). Diplodia corticola was reisolated from wood chips of D. corticola inoculated red (5/5 trees) and white (5/5 trees) oaks and ITS (accession OM716954), LSU (accession OM716955), and ef1-α (accession OM752198) sequences matched D. corticola type culture 112549 ITS (100% identity), LSU (99.76%-100% identity; accession KF766323), and ef1-α (98%-98.9% identity; accession XM_020275852). All D. corticola inoculated trees exhibited seepage from inoculation sites with streaking present in vasculature. Cankers were significantly larger in D. corticola inoculated red (2.34 ± 1.36 cm; P=0.042) and white (2.96 ± 0.52 cm; P=0.00029) oaks compared to agar inoculated trees. To the best of our knowledge, this is the first report of D. corticola causing decline of oaks in Tennessee.

Phytophthora Foot Rot of Deltoid Synurus Caused by Phytophthora cryptogea

Foot rot symptoms were repeatedly observed on plants of deltoid synurus (Synurus deltoides) growing in a field of the Wild Vegetable Research Institute located in Pyeongchang, Korea during disease surveys in July 2020 and June 2021. The symptoms appeared as wilting of the plant leaves, and the plant stems and petioles at or above the soil line turned dark and rotted. The incidence of diseased plants in the field was 5‒10%. Five isolates of Phytophthora sp. were obtained from lesions of the diseased plants and investigated for their morphological and molecular characteristics. All the isolates were identified as Phytophthora cryptogea based on the morphological and molecular characteristics. Three isolates of P. cryptogea were tested for pathogenicity on deltoid synurus plants using artificial inoculation. All the tested isolates caused foot rot symptoms on the inoculated plants. The symptoms were similar to those observed in plants from the field investigated. To our knowledge, this is the first report of P. cryptogea causing Phytophthora foot rot in deltoid synurus.

Universal quadratic soil spectral reflectance line and its deviation patterns' relationships with chemical and textural properties: A global data base analysis

Quantitative evidence for the fundamental spectral shape similarity existing between different soils is presented utilizing Pearson's correlation and SAM (Spectral Angle Mapper) analysis in-between signatures representing thousands of soil samples collected from different environments around the world. A universal quadratic soil line (UQSL) was found to highly fit these representative signatures as well as spectra from the ISRIC (International Soil Reference and Information Center) global database including soil samples collected from 58 countries. K-means clustering of ISRIC spectral reflectance deviations from the UQSL facilitated identification of 5 characteristic deviation patterns. Significant correlation was found for a set of 1000 soil samples between changes in multiple chemical properties and corresponding reflectance changes between these clusters. Future availability of large global soil data bases may allow improved determination of the UQSL and generalization of its deviation patterns relationships with PBC (Physical, Biological and Chemical) properties for wide areas monitoring of soil conditions.

First report of sclerotium rot in sword bean caused by Athelia rolfsii

In 2018, sclerotium rot was observed in sword beans (Canavalia gladiata DC.) grown in a field at Gyeongsangnam-do Agricultural Research and Extension Services. The disease mainly affected the stems and pods of sword beans that touched the ground. The stems that touched the ground were water-soaked at the start of the infection and rotted, slowly withered, and eventually died. White mycelia spread across the lesions, and numerous sclerotia formed on the petiole near the soil line. The sclerotia were globoid, 1–3 mm in diameter, and white to brown in color. Hyphae on PDA were 4–8 μm wide, and clamp connections were observed. Pathogenicity of isolated fungus on sword beans was confirmed according to Koch’s postulates. For molecular identification, we sequenced the entire ITS rDNA sequence of the causative fungus and analyzed its phylogenetic relationships. The fungi isolated from sword beans were in a clade r-1 containing the reference strains of A. rolfsii and S. delphinii. The 540-bp PCR amplicons corresponding to A. rolfsii-specific amplification were produced in all three isolates. Mycological characteristics, ITS rDNA sequencing, the species-specific PCR analysis, and pathogenicity in the host plant identified this fungus as Athelia rolfsii (Curzi) C.C. Tu & Kimbr. This is the first report of sclerotium rot on sword bean caused by A. rolfsii.

First Report of Paraphoma chrysanthemicola Causing Crown and Leaf Rot of Atractylodes lancea in China.

Atractylodes lancea is an important traditional Chinese medicinal plant whose rhizome is used for treating complaints such as rheumatic diseases, digestive disorders, night blindness and influenza. Jiangsu Province is the optimal cultivation location for high-quality A. lancea rhizome. Since June 2019, symptoms of crown rot and leaf rot were observed in about 10-20% of the A. lancea in a plantation (31° 36' 1" N, 119° 6' 40" W) in Lishui, Jiangsu, China. Lesions occurred on the stem near the soil line and on the leaves (Fig. 1A). Disease incidence reached approximately 80-90% by September, 2021 (Fig. 1B) and resulted in severe loss of rhizome and seed yields. For pathogen isolation, ten samples of symptomatic stem segments and ten diseased leaves were collected, surface-sterilized using 5% NaClO solution, rinsed with sterile water, cut into 0.5-2 cm segments, and plated to potato dextrose agar (PDA), and then incubated at 30°C in darkness. Pure cultures of four isolates showing morphological characteristics of Paraphoma spp. were obtained, identified as a single P. chrysanthemicola strain, and named LSL3f2. Newly formed colonies initially consisted of white mycelia; the five-day-old colonies developed a layer of whitish grey mycelia with a grey underside. 20-day-old colonies had white mycelium along the margin and with a faint yellow inner circular part with irregular radial furrows, and the reverse side looking caramel and russet (Fig. 1C). Pycnidia were subglobose (diameter: 5 to 15 μm; Fig. 1D). Unicellular, bicellular or strings of globose or subglobose chlamydospores developed from hyphal cells (Fig. 1E and 1F). The internal transcribed spacer (ITS) region and large subulin-28S of LSL3f2 were cloned using primers ITS1/ITS4 and LR0R/LR7 (Aveskamp et al. 2010, Li et al. 2013), and deposited in GenBank (OK559658 and OK598973, respectively). BLASTn search and phylogenetic analysis showed the highest identity between LSL3f2 and P. chrysanthemicola sequences (Fig. 1G) and confirmed LSL3f2 as P. chrysanthemicola. Koch's postulates were completed using one-month-old vegetatively propagated A. lancea plantlets growing on autoclaved vermiculite/peat mixture at 26°C with a light/dark cycle of 12/12 hours. Each plantlet was inoculated with 5 ml of conidial suspension in water (1 × 108 cfu/ml) by applying to soil close to the plantlet, with sterile water used as a mock control (n = 10). By 20 days post-inoculation, inoculated plantlets showed a range of disease symptoms consistent to those observed in infested fields (Fig. 1H). Pathogenicity was additionally confirmed using detached leaves inoculated with a colonized agar plug of LSL3f2 or an uninoculated control comparison (diameter = 5 mm) and incubated at 26℃ in the dark. Five to seven days post-inoculation, detached leaves showed leaf rot symptoms including lesions, yellowing and withering consistent with those in infested fields, while control leaves remained healthy (n = 10, Fig. 1I). The pathogen was reisolated from the diseased plantlets and detached leaves, in both cases demonstrating the micromorphological characteristics of LSL3f2. P. chrysanthemicola has been reported to cause leaf and crown rot on other plants such as Tanacetum cinerariifolium (Moslemi et al. 2018), and leaf spot on A. japonicain (Ge et al. 2016). However, this is the first report of P. chrysanthemicola causing crown and leaf rot on A. lancea in China.

First Report of Dieback Caused by Phytophthora ramorum on Golden Chinquapin, Chrysolepis chrysophylla, in California.

Golden or giant chinquapin, Chrysolepis chrysophylla (Fagaceae), is a slow growing, evergreen shrub or tree, native to the west coast of the US. In April 2015, several declining chinquapin trees were identified on Bolinas Ridge near Mt. Tamalpais in Marin Co. CA, a Phytophthora ramorum infested region. Branch dieback was observed but no bole or branch cankers were observed. Discolored xylem tissue (5-mm2) was cultured on corn meal agar selective medium, CMA-PARP. An organism morphologically resembling P. ramorum was isolated from one water sprout piece. The ITS sequences obtained from pure cultures, symptomatic branches, and water sprouts were identical, and one was deposited into GenBank. The ITS sequence and cox2 sequence obtained from a culture were a 100% match to the ex-type of P. ramorum. Koch's postulates were tested on 3 small plants (0.5m tall, 3.8-liter pots). Plants were inoculated with 6-mm plugs taken from the margin of a 7-day-old P. ramorum colony, growing on V8 juice agar. Plugs were placed on wounded stems, approximately 2 cm above the soil line, and wrapped in Parafilm. An equal number of plants were treated with uncolonized V8 agar plugs as controls. The potting mix of 2 additional plants of the same size was infested with 200 ml of a zoospore suspension (105 spores ml-1) which was drenched around the base of the stem. Each base was scored with a razor blade before applying the suspension. Two plants were scored and drenched with sterile water as controls. After 7 days, plug-inoculated plants began to wilt and leaves began to turn grayish green. After 15 days, plants began to collapse and isolations were made from stem cankers and roots onto CMA-PARP. After 12 days, leaves of plants that were soil-drenched with zoospores were a dull green color and after 38 days, dry and brittle. Lesions were seen on the roots and stems when isolations were made at 35 days. P. ramorum was consistently isolated from stems of wound-inoculated plants and from roots and stems of soil-inoculated plants. No P. ramorum was isolated, nor symptoms were observed on any control plants. P. ramorum causes sudden oak death on many members of the Fagaceae as well as ramorum blight on more than 130 hosts (Cobb et al., 2020). To our knowledge, this is the first official detection of P. ramorum causing disease on C. chrysophylla in wildlands. Unlike other Fagaceae hosts of P. ramorum, external bole cankers were not seen on infected chinquapin trees. Branch isolations onto PARP media was difficult and detection by PCR from symptomatic vascular tissue was more efficacious. Losses of chinquapin in US forests due to P. ramorum would reduce forest diversity and could cause the loss of habitat for many animal species.

First Report of Seedling Stem Rot on Jinxianlian (Anoectochilus roxburghii) Caused by Fusarium oxysporum in China.

Anoectochilus roxburghii is an important Chinese herbal medicine plant belonging to Orchidaceae and known as Jinxianlian. This orchid is cultivated and mostly adopted to treat diabetes and hepatitis. About 2 billion artificially cultivated seedlings of Jinxianlian are required each year and approximately $600 million in fresh A. roxburghii seedlings is produced in China. From 2011, sporadic occurrence of stem rot on Jinxianlian have been observed in greenhouses in Jinhua City (N29°05', E119°38'), Zhejiang Province. In 2018, nearly 30% of seedlings of Jinxianlian grown in greenhouse conditions were affected by stem rot in Jinhua City. Symptoms initially occurred in the stem at the soil line causing dark discoloration lesions, rotted tissues, wilting, and eventually leading to the death of the plants. A total of 23 diseased seedlings collected from seven different greenhouses were surface sterilized with 1.5% sodium hypochlorite for 3 min, then rinsed in water. Pieces of tissues disinfected from each sample were plated on 2% potato dextrose agar (PDA), and incubated at 25°C in the dark for 5 days (Kirk et al. 2008). A total of 19 isolates were recovered. They developed colonies with purple mycelia and beige or orange colors after 7 days of incubation under 25°C on PDA and carnation leaf agar (CLA) media (Kirk et al. 2008; Zhang et al. 2016). Colonies on PDA had an average radial growth rate of 3.1 to 4.0 mm /d at 25°C. Colony surface was pale vinaceous, floccose with abundant aerial mycelium. On CLA, aerial mycelium was sparse with abundant bright orange sporodochia forming on the carnation leaves. Microconidia were hyaline and oval-ellipsoid to cylindrical (3.7 to 9.3 × 1.3 to 2.9 μm) (n=19). Macroconidia were 3 to 5 septate and fusoid-subulate with a pedicellate base (27.4 to 35.6 × 3.2 to 4.2 μm) (n=19). These morphological features were consistent with Fusarium oxysporum (Sun et al. 2008; Lombard et al., 2019). To confirm the identification based on these morphological features, the internal transcribed spacer region (ITS) and translation elongation factor1 (TEF) were amplified from the DNA of 3 out of 19 isolates chosen at random respectively using the set primer ITS1/ITS4 and EF1/ EF2 (Sun, S., et al. 2018; Lombard et al., 2019). BLAST analysis revealed that the ITS sequences (OK147619, OK147620, OK147621) had 99% identity to that of F. oxysporum isolate JJF2 (GenBank MN626452) and TEF sequence (OK155999, OK156000, OK156001) had 100% identity to that of F. oxysporum isolate gss100 (GenBank MH341210). A multilocus phylogenetic analysis by Bayesian inference (BI) and maximum likelihood (ML) trees based on ITS and TEF indicated that the pathogen grouped consistently with F. oxysporum. Three out of 19 isolates chosen at random were selected to evaluate pathogenicity. Uninfected healthy A. roxburghii seedlings about 40 day-old planted in sterilized substrates were sprayed with distilled water containing 2 x 106 conidia per ml suspensions as inoculums, and plants sprayed with distilled water alone served as controls. Plants were then incubated at 25°C and 85% relative humidity. Ten plants were inoculated for each isolate. After 10 days, all plants inoculated developed stem rot symptoms, while control plants remained healthy. Cultures of Fusarium spp. were re-isolated only from inoculated plants with the frequency of 100% and re-identified by morphological characteristics as F. oxysporum, fulfilling Koch's postulates. To the best of our knowledge, this is the first report of F. oxysporum causing stem rot on A. roxburghii seedlings. As F. oxysporum is a devastating pathogenic fungus with a broad host range, measures should be taken in advance to manage stem rot of A. roxburghii.

First Report of Laurel Wilt Disease Caused by Raffaelea lauricola on Sassafras in Virginia.

Laurel wilt is a lethal vascular disease affecting native Lauraceae in North America. The causal fungus, Raffaelea lauricola T.C. Harr., Fraedrich & Aghayeva and its symbiont, redbay ambrosia beetle, Xyleborus glabratus Eichhoff are native to Asia (Fraedrich et al. 2008, Harrington et al. 2008). Since their introduction near Savannah, Georgia in 2002 (Fraedrich et al. 2008), laurel wilt has spread rapidly, resulting in extensive mortality of native redbay (Persea borbonia [L.] Spreng.) [Hughes et al. 2017] and is a threat to other native Lauraceae, such as sassafras (Sassafras albidum [Nutt.] Nees) (Bates et al. 2013) and northern spicebush (Lindera benzoin [L.] Blume) [Olatinwo et al. 2021]. In June 2021 a sassafras sapling showing wilt and dieback was observed along a roadside in Scott County, Virginia, which borders a laurel wilt-positive Tennessee county (Loyd et al. 2020). The trunk (approximately 5 cm diameter) was submitted to the Virginia Tech Plant Clinic. Although beetle holes were observed, X. glabratus was not found. Discolored sapwood chips were excised and plated on malt extract agar amended with cycloheximide (200 ppm) and streptomycin (100 ppm) [CSMA]. A fungus was consistently recovered and the morphology of conidiophores and conidia, and presence of blastoconidia and mucoid growth, aligned with the description of R. lauricola (Harrington et al. 2008). Two R. lauricola-specific primer sets (Dreaden et al. 2014) were used to amplify DNA extracted from a representative isolate (0248-2021) and confirm R. lauricola. For further confirmation, the LSU region of the rDNA was sequenced (Lloyd et al. 2020). The sequence of the isolate (GenBank accession no. OL583842) showed 100% identity (573/573 bp) to R. lauricola ex-type strain sequence, CBS 121567 (accession no. MH877762) (Harrington et al. 2008, Vu et al. 2018). The isolate was also confirmed by the National Identification Services by sequencing. To confirm pathogenicity, 15 sassafras seedlings (height = 60-100 cm, diameter = 8-10 mm) were inoculated with a conidial suspension harvested from 10-day CSMA cultures of 0248-2021, as follows: two 0.4 mm diameter holes were drilled 10 cm above the soil line at a 45° angle on opposite sides of the stem, leaving at least 3 cm between holes. Ten µl of the conidial suspension (5 x 107/ml) was transferred into each hole and sealed with parafilm. Two sassafras seedlings were inoculated with sterile water. Seedlings were maintained with 12 h photoperiod at 27° ± 2° C. Off-color foliage and loss of turgor were observed 10 days post-inoculation on conidia-inoculated seedlings; at two weeks, these were completely wilted and had sapwood discoloration. Water-inoculated plants showed no symptoms. Sapwood from 15 cm above the inoculation point was excised from 0248-2021-inoculated plants (n=2) and water-inoculated plants (n=1) and plated on CSMA. R. lauricola was recovered from symptomatic plants, but not from water-inoculated plants. The identity of the recovered fungus was confirmed with two species-specific primers sets (Dreaden et al. 2014). It is likely that laurel wilt is more prevalent in the area of the roadside find. Both sassafras and northern spicebush are widespread in Virginia and their range extends into the northeastern US and lower Canada. Laurel wilt poses a serious threat to these species and their ecosystems. For example, spicebush and sassafras are primary hosts of the native spicebush swallowtail butterfly (Papilio troilus L.) [Nitao et al. 1991].

Recognition of the Bare Soil Using Deep Machine Learning Methods to Create Maps of Arable Soil Degradation Based on the Analysis of Multi-Temporal Remote Sensing Data

The detection of degraded soil distribution areas is an urgent task. It is difficult and very time consuming to solve this problem using ground methods. The modeling of degradation processes based on digital elevation models makes it possible to construct maps of potential degradation, which may differ from the actual spatial distribution of degradation. The use of remote sensing data (RSD) for soil degradation detection is very widespread. Most often, vegetation indices (indicative botany) have been used for this purpose. In this paper, we propose a method for constructing soil maps based on a multi-temporal analysis of the bare soil surface (BSS). It is an alternative method to the use of vegetation indices. The detection of the bare soil surface was carried out using the spectral neighborhood of the soil line (SNSL) technology. For the automatic recognition of BSS on each RSD image, computer vision based on deep machine learning (neural networks) was used. A dataset of 244 BSS distribution masks on 244 Landsat 4, 5, 7, and 8 scenes over 37 years was developed. Half of the dataset was used as a training sample (Landsat path/row 173/028). The other half was used as a test sample (Landsat path/row 174/027). Binary masks were sufficient for recognition. For each RSD pixel, value “1” was set when determining the BSS. In the absence of BSS, value “0” was set. The accuracy of the machine prediction of the presence of BSS was 75%. The detection of degradation was based on the average long-term spectral characteristics of the RED and NIR bands. The coefficient Cmean, which is the distance of the point with the average long-term values of RED and NIR from the origin of the spectral plane RED/NIR, was calculated as an integral characteristic of the mean long-term values. Higher long-term average values of spectral brightness served as indicators of the spread of soil degradation. To test the method of constructing soil degradation maps based on deep machine learning, an acceptance sample of 133 Landsat scenes of path/row 173/026 was used. On the territory of the acceptance sample, ground verifications of the maps of the coefficient Cmean were carried out. Ground verification showed that the values of this coefficient make it possible to estimate the content of organic matter in the plow horizon (R2 = 0.841) and the thickness of the humus horizon (R2 = 0.8599). In total, 80 soil pits were analyzed on an area of 649 ha on eight agricultural fields. Type I error (false positive) of degradation detection was 17.5%, and type II error (false negative) was 2.5%. During the determination of the presence of degradation by ground methods, 90% of the ground data coincided with the detection of degradation from RSD. Thus, the quality of machine learning for BSS recognition is sufficient for the construction of soil degradation maps. The SNSL technology allows us to create maps of soil degradation based on the long-term average spectral characteristics of the BSS.

Developing an Injury Severity to Yield Loss Relationship for Soybean Gall Midge (Diptera: Cecidomyiidae).

Soybean gall midge, Resseliella maxima Gagné (Diptera: Cecidomyiidae), is a newly identified pest confirmed on soybean, Glycine max (L.) Merr. (Fabales: Fabaceae). To date, soybean gall midge has been found in Nebraska, Iowa, South Dakota, Minnesota, and Missouri, and has caused severe economic loss to commercial fields since 2018. Much is still unknown about this pest, so research efforts have been focused on biology and management. Larvae feed on the inside of the stem just above the soil line and are difficult to access and time-consuming to sample. In order to accelerate nondestructive sampling efforts, we developed an injury rating system to quantify the severity of plant injury from soybean gall midge larvae. Research plots from 2019 and 2020 in Iowa and Nebraska were evaluated for injury throughout the growing season and yield was measured. Our objective was to describe the relationship between injury severity and yield loss caused from soybean gall midge. A nonlinear regression model was developed to validate our injury rating system and to express the relationship between season long injury severity and yield loss. Results from our analysis indicate the injury rating system we developed correlates well with yield loss caused by larvae and may be an important tool for understanding the economic impact of this emergent pest of soybeans.

First Report of Fusarium falciforme (FSSC 3 + 4) Causing Rot of Industrial Hemp (Cannabis sativa) in California.

Industrial hemp (Cannabis sativa) is a newly legal crop in California that is grown for cannabidiol oil, fiber and seed. In August 2019, whole plant decline and root rot were observed affecting <5% of plants in two industrial fields in Fresno County, CA. Symptoms included chlorotic, collapsed foliage, stem vascular discoloration, and root rot with abundant mycelial growth. Stem and root segments (1-2 cm) from three to five diseased plants were agitated in 0.1% tween-20 and soaked in 70% ethanol for 30 s and 1% NaOCl for 2 min. After incubating for 5 to 7 days on 1:10 potato dextrose agar (PDA) amended with tetracycline, Fusarium selective medium (FSM), and PARP (pimaricin + ampicillin + rifampicin + pentachloronitrobenzene [PCNB] agar) medium, white to pale cream aerial mycelium emerged from tissue of all plants on PDA and FSM but not PARP. Isolates cultured on 0.1% potassium chloride agar formed heads of microconidia on long monophialides consistent with the Fusarium solani species complex (FSSC) (Leslie and Summerell 2008). To obtain pure cultures of two isolates (CS529 and CS530), a single-hyphal tip was excised and grown on PDA. DNA was extracted from actively growing mycelium (PrepMan Ultra kit). The translation elongation factor gene (EF-1α) was amplified via PCR using EF1/EF2 primers (O'Donnell et al. 1998). Sequences of the two isolates were identical and deposited under accession number MW892973 in GenBank. The 599 bp sequence was 99.33% identical to FSSC 3 + 4 (Fusarium falciforme) accessions FD_01443_EF-1a based on FUSARIUM-ID BLAST analysis. To evaluate pathogenicity, stems of hemp plants (cv. 'Berry Blossom'; n=8 plants per isolate) were wounded by penetrating the epidermis in an area about 0.5-cm square by 1-mm deep and 8-inches above the soil line. A 0.5 cm-diameter plug of 7-day old F. falciforme-colonized PDA was placed against the wound. Inoculation sites were loosely wrapped with parafilm for 2 days. A negative control consisted of a sterile PDA plug (n=3). Treatments were arranged in a completely randomized design in a greenhouse. The experiment was conducted once, due to regulatory restrictions at campus facilities. At 61 days post-inoculation, external stem lesions were significantly larger in diameter (P < 0.05; Tukey's HSD) in plants inoculated with CS529 (8 ± 1 mm) compared to the control (2 ± 0 mm), and larger but not significant for CS530 (6 ± 1 mm). Internal stem lesions (i.e., rot in stele) were observed in plants inoculated with CS529 (9 ± 3 mm); stem rot was very minor in plants treated with CS530 (1 ± 1 mm) and nonexistent for control plants. No other disease symptoms were observed. F. falciforme was isolated from stems of CS529- and C530-inoculated plants. Sequences of re-isolates matched 100% with accession MW892973. These results suggest that F. falciforme causes rot in hemp in California. These studies specifically confirm stem rot abilities; field observations of root rot indicate root rotting abilities, but further tests are needed for confirmation. This is the first report of F. falciforme causing disease in industrial hemp. FSSC was described as causing foot rot in hemp in Italy (Sorrentino et al. 2019), but these isolates belonged to phylogenetic species 5 (F. solani) not F. falciforme. In addition, F. falciforme was reported as causing root rot in hydroponically grown cannabis (Punja and Rodriguez 2018). These studies provide the foundation for development of management tools for hemp disease.

First Report of Pectobacterium versatile Causing Potato Soft Rot in Oregon and Washington.

Potato (Solanum tuberosum cv. Norkotah) tubers with symptoms of soft rot were submitted to Oregon State University, Hermiston Agricultural Research and Extension Center Plant Clinic in 2019. One submission in May, originated from a field with poor emergence and seed piece decay (~20% affected) in Umatilla County, Oregon. The second submission, in September, originated from a field in Washington. From each submission, ~100 mg tissue at the margin of infection was washed with distilled water, excised, macerated in 500 L sterile distilled water for 5 minutes. The resulting solution was streaked on crystal violet pectate (CVP) medium and incubated at 28°C for 24 hours. One colony, representative of the many white colonies that formed depressions on CVP plates, was isolated from each submission. Bacterial isolates from Oregon and Washington were named JB56A and JB133A, respectively, and preserved in Luria-Bertani (LB) broth with 15% glycerol at -80°C for long-term storage. Genomic DNA was extracted from JB56A and JB133A cultures grown in LB broth overnight at 30°C using the Wizard SV Genomic DNA kit. The partial dnaX gene (537 bp) was amplified from genomic DNA of each isolate using dnaXf/dnaXr primers (Slawiak et al. 2009) and sequenced. These sequences were deposited to the NCBI GenBank Database, accession numbers MW930747 (JB56A) and MW930748 (JB133A). BLAST analyses (Altschul et al. 1990) using default parameters indicated that the dnaX sequences of JB56A and JB133A were 99.2% (533/537) and 98.7% (530/537) identical to that of P. versatile SCC1 (CP021894). A condensed maximum likelihood tree was built using the partial dnaX sequence of the two query strains, twelve Pectobacterium reference strains to include all known species of Pectobacterium, and four Dickeya species as an outgroup (Fig. S1). JB56A and JB133A formed a monophyletic clade with P. versatile SCC1. Potato (cv. Upstate Abundance) tuber and stem bioassays (Ma et al. 2018) were conducted twice to assess the pathogenicity of these isolates. Tubers were wounded with a sterile 2 mm wide wooden applicator stick and 5 μl culture grown in LB broth overnight (~109 CFU) was pipetted into the wound. Tubers were incubated at 29°C for 24 hours and cut through puncture sites to observe symptoms. Stems of four- or five-week-old plants were wounded with a sterile toothpick about 10 cm above the soil line and a smear of JB56A or JB133A grown on LB agar was inserted into the wound using a toothpick and incubated in a greenhouse for 72 hours. Positive controls (D. dianthicola ME23) and negative controls (no bacteria) were included in both assays. Tubers and stems exhibited disease symptoms after 24 and 72 hours, respectively, following inoculation with JB56A, JB133A, and D. dianthicola ME23. No symptoms were observed for negative controls. The identity of bacteria re-isolated from the margin of stem lesions was confirmed by partial dnaX sequence analyses. P. versatile was recently described as a distinct species based on whole genome sequence comparisons (Portier et al. 2019). In 2018, we isolated P. versatile from potato stems with blackleg disease in New York, and a recent study found that it was isolated in the US from an iris in 1946 (Ma et al. 2021; Portier et al. 2019). However, the geographic distribution and importance of this pathogen in the US remains largely unknown. To our knowledge, this is the first report of potato soft rot caused by P. versatile in Oregon and Washington, two important potato producing states.

First Report of Southern Blight, Caused by Athelia rolfsii (syn. Sclerotium rolfsii) on Hellebores in North America.

Lenten rose (Hellebores hybridus) is an herbaceous perennial grown in landscapes and valued for early spring flowers and high levels of deer resistance. An additional benefit as a landscape plant comes from the high level of disease resistance, with only three fungal pathogens reported in North America. In August of 2021, a Lenten rose plant within a mature landscape in Silver Spring, MD, USA, (lat 39.116629 long 77.043198) was found with a collapsed canopy and brown stems near the soil line. Small clusters of brown sclerotia-like objects were seen along the stem. Samples of the sclerotia and diseased tissue were dipped in 70 percent ethanol for 15 sec, transferred to 5 percent NaClO for 30 sec, immersed in sterile water for one minute, then plated onto Potato Dextrose Agar. Sclerotia-like objects germinated and white mycelia covered the plates within five days of germination. Hyphae emerged from diseased tissue within two days and also grew rapidly. Cultures from sclerotia-like objects and diseased tissue produced white sclerotia which melanized to brown spherical sclerotia ranging in size from two to four mm. Culture samples (1 cm square) were excised from the culture plates and transferred to the base of three two-year old potted hellebore plants. Control plants had blocks of PDA placed at the base of the plants. Plants were placed in plastic bags for two days to maintain humidity, then maintained at room temperature without plastic bags. Petioles turned brown and collapsed within seven days of inoculation. White, fan-like hyphae were present along with maturing sclerotia. Samples from surface sterilized tissue and sclerotia produced the same culture morphology as the originally isolated cultures. Non-inoculated plants remained healthy, and the pathogen was not isolated from non-inoculated plants. Individual DNA samples were prepared from original cultures and the re-isolated cultures. Molecular identification was performed by amplification of the internal rRNA transcribed spacer region (ITS1/4, White et al. 1990 ), the large subunit rRNA (LSU), and the elongation factor-1A (EF1a). Amplification products were cloned into TOPO-TA pcr4 vector and sequenced (Macrogen USA). Sequences were submitted to GenBank for IT1/4 (OK172559) and LSU (OK172560). Homology to ITS1/4 was found with Athelia rolfsii (MN622806), to LSU with Athelia rolfsii (MT225781) and for EF1a with Athelia rolfsii (MW322687). This is the first report of Athelia rolfsii on Hellebores in North America (Farr, D.F &amp; Rossman, A.Y. Fungal Databases, U.S. National Fungus Collections, ARS, USDA. Retrieved September 10, 2021). This report is unique in that few pathogens are known to infect Hellebores(Taylor et al. 2011) and southern blight is not commonly isolated in landscape plantings at Maryland latitudes. 1. White et al. PCR Protocols: A Guide to Methods and Amplifications. Academic Press, San Diego, 1990 2. Taylor, R.K., Romberg, M.K. &amp; Alexander, B.J.R. A bacterial disease of hellebore caused by Pseudomonas viridiflava in New Zealand. Australasian Plant Dis. Notes 6, 28–29, 2011.

First report of Pectobacterium versatile causing blackleg of potato in Serbia.

Potato blackleg is frequently observed on the production fields in the Bačka region of Vojvodina province, which is one of the largest potato-growing areas in Serbia. This disease usually occurs during June and July. In July 2020, blackleg symptoms in the form of stem necrotic lesions, vascular discoloration, hollow stems, and wilting of whole plants were noted on potato cultivar VR808 on a field 28 ha in size located in Maglić village (GPS coordinates 45.349325 N, 19.542768 E). Disease incidence was estimated at 20-25%. Isolations were performed from 12 potato samples on Crystal Violet Pectate medium (CVP). Stem sections consisted of brown lesions and healthy tissue (c.10 cm) were surface sterilized with ethyl alcohol 70% (w/v) and rinsed with sterile distilled water. Small pieces of tissue were taken at the edges of stem lesions (between healthy and diseased tissue) were soaked in phosphate buffer saline for 20 min and plated using a standard procedure (Klement et al. 1990). Single colonies that formed pits after 48 hours at 26 °C were re-streaked onto Nutrient Agar (NA) where creamy white colonies with smooth surfaces were formed. A total of 30 isolates were selected and DNA isolated from the colonies was further analyzed by polymerase chain reaction (PCR) using the partial dnaX gene (DNA polymerase subunit III gamma/tau) with primer pair dnaXf/dnaXr for Pectobacterium and Dickeya species identification (Slawiak et al. 2009). A single characteristic band of 535 bp was amplified in all isolates (Slawiak et al. 2009). DNA sequence alignment showed two distinct groups of isolates (Fig.S1), which were genetically uniform within each group. Using BLASTn search, it was established that the dnaX sequence of the first group (consisting of 19 Serbian potato isolates) had 99.79% identity with NCBI-deposited Pectobacterium versatile strains 14A and 3-2 from potato from Belarus (Acc. No. CP034276 and CP024842, respectively) as well as SCC1 from Finland (Acc. No. CP021894). The remaining 11 dnaX sequences had 100% identity with Pectobacterium carotovorum subsp. carotovorum strain CFBP7081 originating from water in Spain (Acc. No. MK516961). The partial dnaX sequences of three Serbian P. versatile isolates (Pv1320, Pv1520, and Pv1620) and one P. carotovorum subsp. carotovorum (Pcc2520) were deposited in GenBank under Acc. No. MW839571, MW805306, MW839572, and MW805307, respectively. These results, indicating combined infection in the observed field, signify the first identification of P. versatile in Serbia. Multilocus sequence analysis (MLSA) performed with proA (proAF1/ proAR1) and mdh (mdh2/mdh4) genes (Ma et al. 2007; Moleleki et al. 2013) grouped three tested Serbian potato P. versatile isolates together with P. versatile strains from NCBI (Fig.S2). For both tested genes, BLASTn search revealed 100% homology with P. versatile strain SCC1 from Finland. Three Serbian P. versatile potato isolates were deposited under Acc. Nos. MZ682623-25 for proA and MZ682620-22 for mdh genes. According to the routine tests suggested for Pectobacteriaceae (Schaad et al. 2001), Serbian isolates possessed microbiological traits identical to P. versatile description (Portier et al. 2019). Pathogenicity was performed on potato cultivar VR808 with three selected P. versatile isolates (Pv1320, Pv1520, and Pv1620) in the following assays: (i) surface-sterilized tuber slices with holes in the center filled with 100 µL of bacterial suspensions (adjusted to 109 CFU mL-1) to test the isolates' ability to cause soft rot, and (ii) young, four-week old plants with developed 3rd true leaf (c. 30 cm tall) were inoculated by injecting stems with bacterial suspension adjusted to 107 - 108 CFU mL-1 at a height 5 cm above the soil line. Negative controls were treated with sterile distilled water. Inoculated plants were kept under controlled conditions (25 °C temperature and >70% relative humidity). Each assay was replicated twice. Soft rot appeared on tuber slices 24 h after inoculation. On inoculated stems, initial symptoms manifested as greasy elongated spots at inoculation sites two days after inoculation (DAI), and subsequently extended along the vascular tissue and became necrotic. Whole plant's decay was recorded in five DAI, while negative controls remained healthy. To complete Koch's postulates, bacteria were re-isolated from symptomatic potato plants and confirmed by PCR and sequencing of dnaX. This first report of P. versatile in potato indicates that blackleg currently present in Serbia is caused by a diverse bacterial population. This pathogen was first identified in genome comparison as 'Candidatus Pectobacterium maceratum' (Shirshikov et al. 2018) and was later renamed as Pectobacterium versatile sp. nov. (Portier et al. 2019). Thus far, bacterium Pectobacterium carotovorum subsp. brasiliensis has been recognized as dominant pathogen on most of the infected fields in Vojvodina province, and was recently noted on one plot subjected to a combined infection with Dickeya dianthicola (Marković et al. 2021). Findings achieved in this study are highly relevant, as they point to the diversity in potato blackleg pathogens, likely due to the increasingly widespread distribution of imported seed potatoes.

Phytophthora cactorum causing bleeding canker of Acer x freemanii in southern Idaho.

Since 2018, bleeding cankers have been observed on maple trees in multiple home gardens in southwest Idaho. The cankers ooze a dark sap and and are approximately 10 cm to 35 cm in diameter. Cankers typically occur on the main trunk but are also present on scaffold branches in severe infecrions. Symptoms of foliar chlorois, branch dieback, and premature autumn senescence were also associated with the disease. Phytophthora DNA was detected in symptomatic material from five trees using real-time PCR (Miles et al., 2017). In July 2019 recovery of a causal agent from a symptomatic Acer x freemanii tree was attempted. Excisions were made from the interface of healthy and diseased tissue around the cankers using a chisel. The tissue was then placed in sealed plastic ziplock bags at 4°C for 7 days. Hyphae were then removed with forceps and placed onto potato dextrose agar (PDA) amended with penicillin G (0.2 g/liter) and streptomycin sulfate (0.8 g/liter). Colonies resembling Phytophthora cactorum were consistently observed after 5 days at 21°C. Tentative P. cactorum identification was based on the presence of abundant papillate and caducous sporangia on a short pedicel; sporangia were approximately 30 μm long and 26 μm wide (Bush et al., 2006; Hudler, 2013). Individual hyphal tips were transferred to fresh PDA plates and sequencing of both the rDNA ITS region and Cytochrome c oxidase subunit I (COI) was completed for a representative isolate (D19-130). DNA extraction, PCR and sequencing were as previously described (Woodhall et al. 2013; Robideau et al., 2011). The resulting DNA sequences for rDNA ITS (MW315449) and COI (MW881040) were both 100% identical (723/723 bp and 728/728 bp) with sequences from cultures previously identified as P. cactorum (MH171627 and MH136858). To determine pathogenicity, 14 month-old maple (A. x freemanii) trees in individual containers with potting mix were wounded 15 mm above the soil line with a single 10 mm incision using a sterile razor blade and inoculated by placing a 10 mm2 fully colonized PDA plug of isolate D19-130 on the wound. The inoculum and wound were then covered with a damp cotton ball that was secured loosely with parafilm. Control plants consisted of uninoculated plants and wounded plants inoculated with a PDA agar plug. Each treatment was replicated five times and placed in a controlled environment chamber set at 24ºC and 90% relative humidity. All treatments were sprayed with water daily to ensure the cotton balls remained damp. After 8 weeks, black lesions, up to approximately 25 mm above the soil line, were observed on the stem base of all P. cactorum-inoculated plants. No black lesions were observed on non-inoculated plants or plants inoculated with a PDA agar plug. P. cactorum was isolated from lesions, as described above, except polystyrene foam boxes containing moist paper towels were used instead of bags. This report confirms P. cactorum as a causal agent of bleeding canker of maple in Idaho for the first time. It has been shown that several Phytophthora species can infect maple (Jung and Burgess, 2009; Huddler, 2013). P. cactorum has a wide host range but certain strains have been associated with lethal bleeding stem cankers in maple and other deciduous trees worldwide (Huddler, 2013). Knowledge of the causal agent of bleeding canker on maple will help determine appropriate disease management practices.