Ralstonia solanacearum (bacterial wilt of potato).

Ralstonia solanacearum is included in the A2 (high risk) list of quarantine organisms by the European and Mediterranean Plant Protection Organization (EPPO). EPPO Code for R. solanacearum is RALSSO, while the phytosanitary categorization of the species in EPPO A2 list is no.58, EU: I/A2 (EPPO, 2018). Bacterial wilt disease was first reported in southern USA in the late nineteenth century on tomato plants (Smith, 1896). Infected plant materials (e.g. potato tubers) transmit the pathogen over long distances; hence, quarantine inspections and plant sanitary practices are the cornerstone of disease management (EPPO, 2018). R. solanacearum strains in the race 3 group are a select agent under the US Agricultural Bioterrorism Protection Act of 2002 (USDA, 2005). Peculiarly, the organism, if not yet already present in North America in pelargonium (Strider et al., 1981), was introduced with cuttings of this host by American companies producing these cuttings for their markets in countries like Kenya and Guatemala (Norman et al., 1999, 2009; Kim et al., 2002; Williamson et al., 2002; O'Hern, 2004). A similar situation led to introductions of the pathogen from Kenya into some northern European nurseries. Once the source (contaminated surface water) was recognized and proper control measures (use of deep soil water, disinfection of cutting producing premises and replacement of mother stock), the problem was solved and the disease in greenhouses eradicated (Janse et al., 2004). Similarly race 1 has been introduced into greenhouses with ornamental plants (rhizomes, cuttings or fully grown plants) such as Epipremnum, Anthurium, Curcuma spp. and Begonia eliator from tropical areas (Norman and Yuen, 1998, 1999; Janse et al., 2006; Janse, 2012). Introduction can and did occur from Costa Rica and the Caribbean, Indonesia, Thailand and South Africa. However, this idea of placing pathogens on bioterrorist list for unclear and perhaps industry-driven reasons and its effects, is strongly opposed in a recent publication from leading phytobacteriologists. This is because R. solanacearum is an endemic pathogen, causing endemic disease in most parts of its geographic occurrence, moreover normal quarantine regulations are already in place where the disease is not present or only sporadically and are thought to be more efficient and less damaging to trade and research than placing this pathogen on select agent lists and treating it as such (Young et al., 2008). Peculiarly, it has been used in the control of a real invasive species, the weed kahili ginger (Hedychium gardenarium) in tropical forests in Hawaii. This is not without risks because strains occurring on this weed host were thought to be non-virulent, but later appeared to be virulent on many edible and ornamental ginger species as well (Anderson and Gardner, 1999; Paret et al., 2008). Another threat for these countries could be strains belonging to race 1, biovar 1 (phylotype I) that have already been reported from field-grown potatoes in Portugal (Cruz et al., 2008).

Weeds as Pathogen Hosts and Disease Risk for Crops in the Wake of a Reduced Use of Herbicides: Evidence from Yam (Dioscorea alata) Fields and Colletotrichum Pathogens in the Tropics

The transition toward sustainable agriculture requires rethinking cropping systems in the light of less intensive and chemically reliant practices. Weed management is one of the target practices to evolve cropping systems with decreased impact on the environment. While softened management will lead to increased weeds/crops coexistence, it is of importance to assess the relative benefits and drawbacks of new practices. Among the potential drawbacks of weeds/crops coexistence, disease risk may increase if weeds are hosting pathogens. In this study, we assessed the potential of weeds for hosting pathogenic generalist fungi known to translate into disease in crops. We first describe prevalence in fields after harvest and relate prevalence to species characteristics and communities. Then, we directly test the idea that weeds serve as inoculums sources during cropping with a natural experiment. This study highlights variation in host skill among feral weeds for Colletotrichum species, including potential congeneric sub-specialization on different weeds within communities. Last, prevalence within fields was more correlated to focal crop inoculation rates compared to local weed load, but there was a significant correlation effect with prevalence on weeds in the vicinity of fields, suggesting that weeds are mediating disease levels at the local scale, too. Results pointed to the importance of weed host skill in disease risk yet open the door to the potential control of pathogens via targeted weed management.

First Report of Leaf Blight on Parthenium hysterophorus Caused by Nigrospora sphaerica in Malaysia

Weeds may act as inoculum reservoirs for fungal pathogens that could affect other economically important crops (Karimi et al. 2019). In February 2019, leaves of the ubiquitous invasive weed, Parthenium hysterophorus L. (parthenium weed) exhibiting symptom of blight were observed at Ladang Infoternak Sg. Siput (U), a state–owned livestock center in Perak, Malaysia. Symptoms appeared as irregularly shaped, brown–to–black necrotic lesions across the entire leaf visible from both surfaces, and frequently on the older leaves. The disease incidence was approximately 30% of 1,000 plants. Twenty symptomatic parthenium weed leaves were collected from several infested livestock feeding plots for pathogen isolation. The infected tissues were sectioned and surface–sterilized with 70% ethyl alcohol for 1 min, rinsed three times with sterile distilled water, transferred onto potato dextrose agar, and incubated at 25°C under continuous dark for 7 days. Microscopic observation revealed fungal colonies with similar characteristics. Mycelium was initially white and gradually changed to pale orange on the back of the plate but later turned black as sporulation began. Conidia were spherical or sub–spherical, single–celled, smooth–walled, 12 to 21 μm diameter (mean = 15.56 ± 0.42 μm, n= 30) and were borne on a hyaline vesicle. Based on morphological features, the fungus was preliminarily identified as Nigrospora sphaerica (Sacc) E. W. Mason (Wang et al. 2017). To confirm identity, molecular identification was conducted using isolate 1SS which was selected as a representative isolate from the 20 isolates obtained. Genomic DNA was extracted from mycelia using a SDS–based extraction method (Xia et al. 2019). Amplification of the rDNA internal transcribed spacer (ITS) region was conducted with universal primer ITS1/ITS4 (White et al. 1990; Úrbez–Torres et al. 2008). The amplicon served as a template for Sanger sequencing conducted at a commercial service provider (Apical Scientific, Malaysia). The generated sequence trace data was analyzed with BioEdit v7.2. From BLASTn analysis, the ITS sequence (GenBank accession number. MN339998) had at least 99% nucleotide identity to that of N. sphaerica (GenBank accession number. MK108917). Pathogenicity was confirmed by spraying the leaf surfaces of 12 healthy parthenium weed plants (2–months–old) with a conidial suspension (106 conidia per ml) collected from a 7 day–old culture. Another 12 plants served as a control treatment and received only sterile distilled water. Inoculation was done 2 h before sunset and the inoculated plants were covered with plastic bags for 24 h to promote conidial germination. All plants were maintained in a glasshouse (24 to 35°C) for the development of the disease. After 7 days, typical leaf blight symptoms developed on the inoculated plants consistent with the symptoms observed in the field. The pathogen was re–isolated from the diseased leaves and morphological identification revealed the same characteristics as the original isolate with 100% re–isolation frequency, thus, fulfilling Koch’s postulates. All leaves of the control plants remained symptomless and the experiment was repeated twice. In Malaysia, the incidence of N. sphaerica as a plant pathogen has been recorded on several important crops such as watermelon and dragon fruit (Kee et al. 2019; Ismail and Abd Razak 2021). To our knowledge, this is the first report of leaf blight on P. hysterophorus caused by N. sphaerica from this country. This report justifies the significant potential of P. hysterophorus as an alternative weed host for the distribution of N. sphaerica. Acknowledgement This research was funded by Universiti Putra Malaysia (UPM/GP–IPB/2017/9523402). References Ismail, S. I., and Abd Razak, N. F. 2021. Plant Dis. 105:488. Karimi, K., et al. 2019. Front Microbiol. 10:19. Kee, Y. J., et al. 2019. Crop Prot. 122:165. Úrbez–Torres, J. R., et al. 2008. Plant Dis. 92:519. Wang, M., et al. 2017. Persoonia 39:118. White, T. J. et al. 1990. Page 315 in: PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, CA. Xia, Y., et al. 2019. Biosci Rep. 39:BSR20182271.

Ralstonia solanacearum (bacterial wilt of potato).

Ralstonia solanacearum is included in the A2 (high risk) list of quarantine organisms by the European and Mediterranean Plant Protection Organization (EPPO). EPPO Code for R. solanacearum is RALSSO, while the phytosanitary categorization of the species in EPPO A2 list is no.58, EU: I/A2 (EPPO, 2018). Bacterial wilt disease was first reported in southern USA in the late nineteenth century on tomato plants (Smith, 1896). Infected plant materials (e.g. potato tubers) transmit the pathogen over long distances; hence, quarantine inspections and plant sanitary practices are the cornerstone of disease management (EPPO, 2018). R. solanacearum strains in the race 3 group are a select agent under the US Agricultural Bioterrorism Protection Act of 2002 (USDA, 2005). Peculiarly, the organism, if not yet already present in North America in pelargonium (Strider et al., 1981), was introduced with cuttings of this host by American companies producing these cuttings for their markets in countries like Kenya and Guatemala (Norman et al., 1999, 2009; Kim et al., 2002; Williamson et al., 2002; O'Hern, 2004). A similar situation led to introductions of the pathogen from Kenya into some northern European nurseries. Once the source (contaminated surface water) was recognized and proper control measures (use of deep soil water, disinfection of cutting producing premises and replacement of mother stock), the problem was solved and the disease in greenhouses eradicated (Janse et al., 2004). Similarly race 1 has been introduced into greenhouses with ornamental plants (rhizomes, cuttings or fully grown plants) such as Epipremnum, Anthurium, Curcuma spp. and Begonia eliator from tropical areas (Norman and Yuen, 1998, 1999; Janse et al., 2006; Janse, 2012). Introduction can and did occur from Costa Rica and the Caribbean, Indonesia, Thailand and South Africa. However, this idea of placing pathogens on bioterrorist list for unclear and perhaps industry-driven reasons and its effects, is strongly opposed in a recent publication from leading phytobacteriologists. This is because R. solanacearum is an endemic pathogen, causing endemic disease in most parts of its geographic occurrence, moreover normal quarantine regulations are already in place where the disease is not present or only sporadically and are thought to be more efficient and less damaging to trade and research than placing this pathogen on select agent lists and treating it as such (Young et al., 2008). Peculiarly, it has been used in the control of a real invasive species, the weed kahili ginger (Hedychium gardenarium) in tropical forests in Hawaii. This is not without risks because strains occurring on this weed host were thought to be non-virulent, but later appeared to be virulent on many edible and ornamental ginger species as well (Anderson and Gardner, 1999; Paret et al., 2008). Another threat for these countries could be strains belonging to race 1, biovar 1 (phylotype I) that have already been reported from field-grown potatoes in Portugal (Cruz et al., 2008).

First Report of the Rust Disease Caused by Puccinia crepidis-japonicae on New Host Sonchus arvensis from Pakistan.

Sonchus arvensis (Asteraceae) is a traditional medicinal herb. The aerial parts are a rich source of vitamins, essential amino acids, and minerals, which may help in treatment of fever, inflammation, detoxication, and blood circulation (Li et al. 2018). In October 2018, typical rust symptoms were observed on S. arvensis leaves and stems in Buner district, Malakand division, Khyber PakhtunKhwa Province, Pakistan (34.39°N; 72.61°E). Almost 40% of leaves and stems of five S. arvensis plants displayed severe rust. The specimens were examined microscopically and compared with type specimen description in published literature (Dietel 1908; Hiratsuka et al. 1992). The fungus was identified as Puccinia crepidis-japonicae based on the characteristic of spore morphology and phylogenetic study based on the internal transcribed spacer (ITS) and large subunit (LSU) sequence data. Uredinia were amphigenous, rounded, or somewhat elliptical, naked, small patches, equally spread, brown. Urediniospores (n=30) were globose to ellipsoid, brownish yellow and measured 22.4–24.7 × 20.2–22.1 µm. Urediniospore walls were brownish orange and finely echinulate and 1.7 to 2.1 µm thick with 2 to 3 germ pores. Telia were amphigenous, rounded or elliptic, scattered, dark brown to blackish. Teliospores (n=30) were ellipsoid, subglobose or long ellipsoid, rounded at both ends, not thickened at apex, warted, reddish brown, and measured 31.3–39 × 24.6–26.8 µm. Teliospore walls were reddish-black and about 1.5–2.5 µm thick, and the pedicles were short, hyaline, fragile, become tapered toward apex, and measured 14.4–18.7 × 4.7–9 µm. DNA was extracted from urediniospores, and the combined region of ITS and LSU (28S) were amplified using Rust2Inv (forward primer) and LR6 (Reverse primer) according to the protocol outlined by Aime (2006). A BLASTn search (http://www.ncbi.nlm.nih.gov) showed that the combined ITS and LSU region shared 99% identity (792/804 bp) to the P. crepidis-japonicae accessions (KY798395 from Hawaii, USA) with 100% query cover. The resulting sequence was deposited in GenBank (Accession No. MN093335). Both morphological and molecular characteristics indicatethat this species was P. crepidis-japonicae. To test pathogenicity and fulfill the Koch's postulates, a urediniospore suspension (1 × 104 spores/ml) was sprayed on three 6-week-old plants of S. arvensis, and one as negative control, incubated at 22–24°C. Uredinia were observed on the leaves after 10 days of inoculation, whereas the control plants remained symptomless. Microscopic examination confirmed that the symptoms on plants obtained from the field and greenhouse inoculations were morphologically identical. This fungus has been observed previously on Crepis japonica in China, Hong Kong, Japan, Korea, and Taiwan, on Prenanthes spp. in China and on Youngia tenuifolia and Y. fusca in China (Farr and Rossman 2021). Pereira et al. (2002) suggested that P. crepidis-japonicae may play a significant role as a biocontrol agent against its weed host. To the best of our knowledge, there are no other reports of this fungus on any other hosts in Pakistan. The specimen has been vouchered in LAH Herbarium, Department of Botany, University of the Punjab, Lahore, Pakistan (LAH36343). This is the first report of P. crepidis-japonicae on S. arvensis as a new host from Pakistan.