Potential Inoculum Source Research Articles

Konya İli Böğürtlen, Ahududu ve Kuşburnu Yetiştiriciliğinde Potansiyel Bir Tehdit: Ateş Yanıklığı Hastalığı

In the present study, totally 49 samples, which showed the symptoms of leaf and shoot blight and cankers with brown discoloration of necrotic tissues on mature branches, were collected from 22 districts and areas of Konya Province between 2017 and 2019. Presence rate of E. amylovora in collected samples, showing symptoms of the disease, from the province was determined to be 40% for blackberry and raspberry and 33% rosehip for rosehip in three years. Bacteria consistently isolated from the diseased tissues were identified on the basis of biochemical, physiological, and molecular tests, comparing with a reference strain of E. amylovora, isolated from blackberry (Kbb 371). Twenty seven representative bacterial strains were gram-negative, rod-shaped, mucoid, fermentative, positive for levan formation and acetoin production, no growth at 36°C, positive for gelatin hydrolysis, and negative for esculin hydrolysis, indole, urease, catalase, oxidase, arginine dehydrolase, reduction of nitrate, acid production from lactose, and inositol. All strains induced a hypersensitive response in tobacco (Nicotiana tobacum cv. White Burley) 24 h after inoculation with a 108 CFU ml-1 bacterial suspension in sterile distilled water. The strains were identified as E. amylovora using the species-specific primers set A/B (1), which amplified a 1-kb DNA fragment in PCR, and the matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) method. In order to fulfill the Koch postulates, pathogenicity test was confirmed by injecting bacterial suspensions of 108 CFU ml-1 in sterile distilled water into the shoot tips of 3-year-old blackberry R. fruticosus cv. Chester, raspberry R. idaeus cv. Heritage and rosehip R. canina. All tests were repeated three times. The bacterium was re-isolated from inoculated plants and identified as E. amylovora. Phytosanitary measures are needed to prevent any further spread of the bacterium as potential inoculum sources to new blackberry, raspberry and rosehip growing areas.

First Report of Potato Viruses Infecting Lamium purpureum in Ukraine

Weeds as reservoirs for destructive plant pathogens have a significant impact on the viral epidemiology, ecology and, as a result, on local economy, and are therefore being investigated in many parts of the world. Thus, the aim of this study was to investigate virus occurrence in red dead-nettle plants (Lamium purpureum L.) widespread in urban and field conditions throughout the in the Kyiv region of Ukraine. Methods. Field crop observations, visual diagnosis, biological testing of the virus, immunoassay (ELISA), polymerase chain reaction with reverse transcription (RT-PCR), sanger sequencing of partial genome sequences of PVX, PVY, PVS, PVM. Results. The results obtained in the study indicate that Lamium plants could be alternative weed hosts of number important viral diseases including potatoes and other vegetables. Serological and molecular test results evidence plants were infected by Potato virus X, Potato virus Y, Potato virus M, Potato virus S and therefore Lamium L. species can serve as a potential source of inoculum for wide range of vegetables and ornamentals. This study is the first report of Lamium plants being naturally infected with Potato virus M and Potato virus S in central Europe. Conclusions. These plants are alternative host of mixed infection with viruses belonging to different families: Alphaflexiviridae, Betaflexiviridae and Potyviridae.

First report of Colletotrichum truncatum causing anthracnose in cassava in Brazil.

Cassava (Manihot esculenta Crantz) presents significant economic importance in Brazil and other developing countries due to its use in human and animal feeding. In 2019, cassava plants sampled in Pará state (Brazil) presented necrotic and irregular leaf spots, characteristic symptoms of cassava anthracnose. About 90% of the plants were symptomatic, and disease severity was higher during months with high temperature and humidity. Fragments of symptomatic tissues were removed from the lesion transition area, surface disinfested (45 s in 70% ethanol, 1 min in 1% NaOCl, and rinsed twice in sterile water), and plated on potato dextrose agar. Cultures were incubated at 25 °C under continuous light for 7 days. Among the obtained isolates, seven presented grey felt-like mycelium with white sectors, reverse greyish, and hyaline, aseptate, smooth-walled, falcate conidia with average size 20.7-30.7 (26.1 ± 2.1) × 2.4-4.8 (3.5 ± 0.5) μm. Phenotypical features were similar to C. truncatum (Damm et al. 2019). The representative isolate UFT/Coll87 was chosen for further assays. The identity of the isolate was determined by maximum likelihood analysis using sequences of actin (ACT, GenBank accession number MT321653), β-tubulin (TUB2, MT856673) and glyceraldehyde-3-phosphate dehydrogenase (GAPDH, MT800857) partial regions. Colletotrichum isolate from cassava nested with C. truncatum isolates in a clade with 100% support, being confidently assigned to this species. Koch's postulates were fulfilled to confirm the pathogenicity of UFT/Coll87. Inoculation was carried out in three cassava plants by spraying a conidial suspension (106 conida mL-1) on unwounded leaves (three leaves per plant). Plants sprayed with sterile water represented negative control. Inoculated plants were kept in a humid chamber for 48 h, 25 °C, and a 12-h photoperiod. The experiment was repeated 2 times. Typical cassava anthracnose symptoms were observed 10 days after inoculation. No symptoms were observed in negative control. The pathogen was reisolated from symptomatic leaves and was phenotypically identical to the original isolate UFT/Coll87, fulfilling Koch's postulates. Colletotrichum fructicola, C. karstii, C. plurivorum, and C. siamense were reported causing cassava anthracnose in China (Liu et al. 2019). In Brazil, C. chrysophilum, C. fructicola, C. siamense and C. theobromicola were reported in association with cassava (Bragança et al. 2016; Oliveira et al. 2018; Machado et al. 2020). To our knowledge, this is the first report of C. truncatum causing cassava anthracnose worldwide. Our finding is important for disease management due to the high host range of C. truncatum. The pathogen can reduce the cassava yield, and the crop may serve as a potential inoculum source since it is commonly cultivated near to other crops that are also infected by C. truncatum.

Potential inoculum sources of Fusarium species involved in asparagus decline syndrome and evaluation of soil disinfestation methods by qPCR protocols.

Asparagus decline syndrome (ADS), one of the most important diseases affecting asparagus crops, causes important yield losses worldwide. Fusarium proliferatum, F. oxysporum and F. redolens are among the main species associated with ADS. To explore their potential inoculum sources and the effectiveness of soil disinfestation practices for ADS management, molecular methods based on a quantitative real-time polymerase chain reaction (qPCR) were developed. qPCR-based molecular tools demonstrated advantages in the sensitive and specific detection and quantification of fungal pathogens in comparison with less-accurate and time-consuming traditional culture methods. F. proliferatum, F. oxysporum and F. redolens could be specifically detected and accurately quantified in asparagus plants, soil and irrigation water collected from asparagus fields with ADS symptoms by means of the designed TaqMan qPCR protocols. Furthermore, these molecular tools were successfully applied for evaluation of the efficacy of diverse soil disinfestation treatments. Chemical fumigation with dazomet and biosolarization with pellets of Brassica carinata contributed to a significant reduction in the inoculum densities of the three Fusarium species in treated soils, which was correlated with production increases. The capability to accurately detect and quantify the main Fusarium species involved in ADS in plants, soil and water samples by means of qPCR will allow identification of high-risk fields that can be avoided or managed to reduce yield losses. Quantification of pathogen densities in the soil may also provide essential insights into the effectiveness of soil disinfestation methods for ADS management. © 2021 Society of Chemical Industry.

Colletotrichum gloeosporioides sensu lato associated with anthracnose in Bauhinia forficata

Bauhinia forficata is an important plant commonly used in traditional medicine, and as an ornamental plant in parks and gardens. Its use as an ornamental plant is compromised by the symptoms of anthracnose. Although anthracnose caused by Colletotrichum species is a widespread plant disease and observed across several Brazilian regions, the species associated with this disease are unknown for most host species. The present study aimed to identify the Colletotrichum species associated with anthracnose of B. forficata in Brazil, as well as the prevalence and aggressiveness of each. Multilocus phylogenetic analyses including the loci APN2, APN2-MAT/IGS, CAL, GAP2/IGS, GAPDH, GS and TUB2 reveals that C. chrysophilum, C. siamense and C. tropicale are associated with B. forficata anthracnose across three different Brazilian geographic regions. Colletotrichum siamense was prevalent in all sampled regions and was also the most aggressive species on B. forficata leaves. The Colletotrichum species here reported have been found on several commercial crops. Since B. forficata commonly occurs as a native plant in areas near commercial fields, it may represent a potential source of inoculum to these crops.

Succession of the fungal endophytic microbiome of wheat is dependent on tissue-specific interactions between host genotype and environment

Fungi living inside plants affect many aspects of plant health, but little is known about how plant genotype influences the fungal endophytic microbiome. However, a deeper understanding of interactions between plant genotype and biotic and abiotic environment in shaping the plant microbiome is of significance for modern agriculture, with implications for disease management, breeding and the development of biocontrol agents.For this purpose, we analysed the fungal wheat microbiome from seed to plant to seeds and studied how different potential sources of inoculum contributed to shaping of the microbiome. We conducted a large-scale pot experiment with related wheat cultivars over one growth-season in two environments (indoors and outdoors) to disentangle the effects of host genotype, abiotic environment (temperature, humidity, precipitation) and fungi present in the seed stock, air and soil on the succession of the endophytic fungal communities in roots, flag leaves and seeds at harvest. The communities were studied with ITS1 metabarcoding and environmental climate factors were monitored during the experimental period.Host genotype, tissue type and abiotic factors influenced fungal communities significantly. The effect of host genotype was mostly limited to leaves and roots, and was location-independent. While there was a clear effect of plant genotype, the relatedness between cultivars was not reflected in the microbiome. For the phyllosphere microbiome, location-dependent weather conditions factors largely explained differences in abundance, diversity, and presence of genera containing pathogens, whereas the root communities were less affected by abiotic factors. Our findings suggest that airborne fungi are the primary inoculum source for fungal communities in aerial plant parts whereas vertical transmission is likely to be insignificant.In summary, our study demonstrates that host genotype, environment and presence of fungi in the environment shape the endophytic fungal community in wheat over a growing season.

Ecology‐based analysis of a recent association between Spartium junceum and 16SrV phytoplasma

AbstractAn outbreak of Spartium witches’ broom (SpaWB) in Sicily prompted us to identify and characterize associated phytoplasmas. Over 80 samples of Spanish broom (Spartium junceum) and around 270 individuals of the potential vector Livilla spectabilis were collected and analysed. Single and mixed infections of 16SrV and ‘Candidatus Phytoplasma spartii’ were detected in Spanish broom samples and for the first time in L. spectabilis. The 16SrV isolates were further characterized by multilocus sequence typing (MLST) to determine their phylogenetic relationship with flavescence dorée phytoplasma (FDp) and to evaluate the risk of host‐jumping to grapevine. Phylogenetic analysis of most of the analysed genes using the MLST approach grouped S. junceum 16SrV‐C isolates with FDp isolates infecting grapevine and Scaphoideus titanus. Notably, phylogenetic analysis of the vmpA gene clustered the S. junceum isolates with FDp genotypes transmitted by S. titanus. This study extends the knowledge of SpaWB epidemiology, focusing on the possible risk of a 16SrV host jump from Spanish broom to grapevine. Spanish broom was identified as a reservoir and potential inoculum source of phytoplasmas that cause severe disease in cultivated crops. Furthermore, the L. spectabilis psyllid may be involved in the epidemiology of this 16SrV‐C phytoplasma, although in the absence of in vivo transmission trials. The study further confirms the strong ability of phytoplasmas to adapt to new hosts and vectors, thus leading to potential phytosanitary emergencies.

A novel monopartite begomovirus and satellites associated with yellow mosaic disease of Sida spp. in India.

Begomoviruses (family Geminiviridae) cause severe diseases in many economically important crops and non-cultivated plants in the warmer regions of the world. Non-cultivated weeds have been reported to act as natural virus reservoirs. In January 2016, Sida plants with yellow mosaic symptoms were found at the edge of an agricultural field in Gujarat, India. Sequence analysis of the viral genomic components cloned from a diseased Sida plant indicated the presence of a distinct monopartite begomovirus (proposed as sida yellow mosaic Gujarat virus) along with a betasatellite (ludwigia leaf distortion betasatellite) and an alphasatellite (malvastrum yellow mosaic alphasatellite). Our results emphasize that this weed may harbor a begomovirus-alphasatellite-betasatellite complex. This host serves as a potential source of virus inoculum, which can be transmitted by whiteflies to other cultivated crops.

Enrichment of salt-tolerant CO2–fixing communities in microbial electrosynthesis systems using porous ceramic hollow tube wrapped with carbon cloth as cathode and for CO2 supply

Microbial inocula from marine origins are less explored for CO2 reduction in microbial electrosynthesis (MES) system, although effective CO2–fixing communities in marine environments are well-documented. We explored natural saline habitats, mainly salt marsh (SM) and mangrove (M) sediments, as potential inoculum sources for enriching salt-tolerant CO2 reducing community using two enrichment strategies: H2:CO2 (80:20) enrichment in serum vials and enrichment in cathode chamber of MES reactors operated at −1.0 V vs. Ag/AgCl. Porous ceramic hollow tube wrapped with carbon cloth was used as cathode and for direct CO2 delivery to CO2 reducing communities growing on the cathode surface. Methanogenesis was dominant in both the M- and SM-seeded MES and the methanogenic Archaea Methanococcus was the most dominant genus. Methane production was slightly higher in the SM-seeded MES (4.9 ± 1.7 mmol) compared to the M-seeded MES (3.8 ± 1.1 mmol). In contrast, acetate production was almost two times higher in the M-seeded MES (3.1 ± 0.9 mmol) than SM-seeded MES (1.5 ± 1.3 mmol). The high relative abundance of the genus Acetobacterium in the M-seeded serum vials correlates with the high acetate production obtained. The different enrichment strategies affected the community composition, though the communities in MES reactors and serum vials were performing similar functions (methanogenesis and acetogenesis). Despite similar operating conditions, the microbial community composition of M-seeded serum vials and MES reactors differed from the SM-seeded serum vials and MES reactors, supporting the importance of inoculum source in the evolution of CO2-reducing microbial communities.

Association of a begomovirus-satellite complex with yellow vein and leaf curl disease of hollyhock (Alcea rosea) in India.

Geminiviruses cause considerable yield loss in several crop plants worldwide. In 2016, several hollyhock plants displaying yellow mosaic and leaf curling symptoms were noticed in a nursery of Jawaharlal Nehru University, New Delhi, India. Analysis of the collected samples indicated an association of monopartite and bipartite begomoviruses with satellites. Three begomoviruses (including a member of a new begomovirus species), two alphasatellites, and a betasatellite were isolated from yellow-mosaic-disease-affected plants. Similarly, a begomovirus, two alphasatellites, and a betasatellite were found to be associated with leaf curl disease of hollyhock. These begomoviruses and satellites were found to be recombinants. By harboring diverse begomoviruses and satellite DNAs, hollyhock may serve as a potential source of virus inoculum.

Can Amplicephalus funzaensis Linnavuori 1968 (Hemiptera: Cicadellidae) transmit phytoplasmas to strawberry?

Phytoplasmas are plant pathogenic bacteria of the class Mollicutes that lack cell walls, are restricted to the phloem of their plant hosts, are difficult to culture, and are transmitted by insect vectors. Phytoplasmas from 16SrI and 16SrVII groups have been associated with diseases in urban trees in the Bogota plateau and with potato and strawberry crops in Cundinamarca,Colombia. The objective of this work was to evaluate if the vector Amplicephalus funzaensis (Hemipera: Cicadellidae) could transmit phytoplasmas to Fragaria x ananassa under semi-controlled conditions. A transmission assay on F. x ananassa var. Monterrey was performed with A. funzaensis from a population naturally infected with phytoplasmas, whose host was Cenchrus clandestinus. Seven months after herbivory by these insects, the plants did not show symptoms associated to phytoplasmas, even though more than one third of the insects used carried phytoplasmas. In total, 120 A. funzaensis individuals were tested for the presence of phytoplasmas by molecular methods; of these, 46 (38%) were positive for phytoplasmas, showing the existence of insect populations with a high number of individuals that are a potential source of inoculum for the pathogen transmission. Additionally, for the molecular identification of A. funzaensis, a DNA barcode was generated from the cyitochrome c oxidase (COI) gene.

PATHOGENICITY OF Macrophomina SPECIES COLLECTED FROM WEEDS IN COWPEA

ABSTRACT Charcoal rot caused by Macrophomina phaseolina is a major cowpea disease causing substantial losses to growers. In the semi-arid region of Brazil, cowpea is one of the most widely used alternatives for crop rotation during the off-season of melon. This favors Macrophomina multiplication because both crops are hosts of this pathogen. The objective of this study was to verify the pathogenicity of Macrophomina phaseolina and M. pseudophaseolina on cowpea. The Macrophomina spp. isolates used were obtained from the roots of Trianthema portulacastrum and Boerhavia diffusa, weed species prevalent in melon production areas in North-east Brazilian. The experiment was carried out in a greenhouse. Cowpea plants cv. ‘Paulistinha’ were inoculated with 30 M. phaseolina isolates, 30 M. pseudophaseolina isolates and a reference isolate of M. phaseolina obtained from cowpea roots. All Macrophomina isolates were able to cause disease on cowpea and there were no statistical differences between both Macrophomina species regarding disease incidence and severity. Moreover, 65.2 and 100.0% of the M. phaseolina isolates, and 56.2 and 92.8% of the M. pseudophaseolina isolates, obtained from T. portulacastrum and B. diffusa, respectively, were as severe to cowpea as the M. phaseolina reference isolate from cowpea. These results emphasize the need to establish management practices aiming to control T. portucalastrum and B. diffusa from cowpea production areas, as they can act as potential sources of inoculum and survival for Macrophomina spp.

Ralstonia solanacearum causing potato bacterial wilt: host range and cultivars’ susceptibility in Rwanda

AbstractMembers of the Ralstonia solanacearum species complex (RSSC), causing potato bacterial wilt or brown rot, are highly contagious and there are no known cultivars with durable resistance to the pathogen. This study hypothesized (a) that crops intercropped or rotated with potato, plants in the same family, and plants grown in the neighbouring fields can host the pathogen and they can be potential sources of primary inoculum, and (b) that potato cultivars currently multiplied by the public tissue culture laboratory in Rwanda are less susceptible to the pathogen. Fourteen plant species and potato, and nine potato cultivars were tested for susceptibility to an RSSC phylotype II strain under greenhouse conditions. The bacteria induced symptoms on potato, tomato, tree tomato, sweet pepper, and eggplant only. Among the plant species with symptoms, potato, tomato, and tree tomato wilted completely. There was a significant difference in days to symptom expression and to complete wilting (p < .0001). While all tested potato cultivars were found to be susceptible, the number of days to first symptom expression, days to complete wilting, area under the disease progress curve (AUDPC), and the number and weight of harvested tubers varied considerably. Cultivars Cruza, Kinigi, and CIP‐58 were less susceptible whereas the cultivars Gikungu, Kirundo, and Victoria were highly susceptible. There is a strong need to search for other sources of resistance. The results indicate that some plant species that might serve as a reservoir of the bacterium should be avoided in the vicinity of potato crops.

Corroboration That Highly Resistant Impatiens Cultivars Are Not Immune to Downy Mildew Disease: A Report of Crop Losses from Two California Producers

Two new series of Impatiens walleriana (impatiens) cultivars, Beacon and Imara XDR, were released to commercial growers in the United States in 2019 to 2020. Field trials show these new cultivar series are highly resistant to impatiens downy mildew (IDM). However, neither of these two impatiens series are completely immune to the disease, and preventive fungicide programs are still recommended for use throughout production to maintain plant health. Here we report two destructive outbreaks of IDM from Imara XDR in two commercial production facilities in California, one in 2019 and one in 2020. The disease outbreaks were caused by a known rDNA genotype of Plasmopara destructor (synonym = P. obducens). Modified Koch’s postulates showed that the pathogen could infect and cause disease in both Beacon and Imara XDR plants. Mefenoxam applied by both growers may have been ineffective due to resistance in P. destructor populations, which has been demonstrated on several previous occasions. Given these findings, fungicide programs intended to supplement genetic resistance should not be overly reliant upon application of mefenoxam and should utilize effective materials from different mode of action groups, in rotation. Fungicides to supplement genetic resistance are particularly appropriate in frost-free areas or in any circumstances that provide a potential inoculum source.

Identification of inoculum sources of Fusicladium eriobotryae in loquat orchards in Spain

Fusicladium eriobotryae is the causal agent of loquat scab, the main disease damaging fruit, leaves and young twigs of this crop. A two-growing season study (2015–2016 and 2016–2017) was carried out in two loquat orchards (cv “Algerie”) to determine the inoculum sources of F. eriobotryae by direct observation of conidia, pathogen isolation on culture media and detection using a new real time PCR protocol developed in this study. One-year-old twigs, fruit peduncles and fruit mummies were randomly sampled three times per growing season on each orchard, and inflorescences only at flowering. Conidia of F. eriobotryae were not found and the isolation of the pathogen was neither possible from any sample in both seasons. Specific primers FUG2F and FUG2R, were designed to detect and quantify DNA of F. eriobotryae on plant material, with a limit of detection (LOD) established at 48.6 fg/μl. The DNA of the pathogen was not detected by real time PCR in fruit mummies nor inflorescences. It was detected in fruit peduncles and twigs in the season 2016–2017 with concentrations ranging from 50 to 2742 fg/μl, confirming that this two loquat organs might act as potential inoculum sources for F. eriobotryae. The detection of F. eriobotryae only in this season agrees with the predictions of an epidemiological model for this pathogen. Our results indicate that in years with a high disease pressure, fruit twigs and peduncles might act as a source of inoculum of new infections the following year.

Molecular Epidemiology of Xanthomonas perforans Outbreaks in Tomato Plants from Transplant to Field as Determined by Single-Nucleotide Polymorphism Analysis.

Outbreaks of bacterial spot on tomato (BST) caused by Xanthomonas perforans are a major concern for sustainable crop production. BST is a common occurrence in tomato transplants grown for field production. We hypothesized that BST outbreaks in commercial fields originate from X. perforans strains inadvertently introduced from commercial transplant facilities. To test this hypothesis, we used a genome-wide single-nucleotide polymorphism (SNP) analysis to characterize X. perforans strains recovered from tomato transplant facilities and fields in commercial production areas. X. perforans strains were isolated from symptomatic transplants prior to roguing at two commercial transplant growers. Then, the same groups of transplants were tracked to commercial fields to recover X. perforans strains from diseased plants prior to harvest. Whole-genome sequencing was carried out on 84 strains isolated from transplant and field plants from Florida and South Carolina. SNPs were called using three reference strains that represented the genetic variation of the sampled strains. Field strains showing genetic similarity to transplant strains had a difference of 2 to 210 SNPs. Transplant and field strains clustered together by grower within each phylogenomic group, consistent with expectations. The range of genetic divergence among strains isolated from field plants was similar to the range obtained from strains on transplants. Using the range of genetic variation observed in transplants, we estimate that 60% to 100% of field strains were an extension of the transplant strain population. Our results stress the importance of BST management to reduce X. perforans movement from transplant to field and to minimize subsequent disease outbreaks.IMPORTANCE Current management of Xanthomonas perforans on tomato plants mainly relies on the frequent application of pesticides. However, the lack of effective pesticides and the development of strain tolerance to certain bactericides limit the ability to control outbreaks in production fields. Better knowledge of probable sources of X. perforans inoculum during tomato production is required to refine management strategies. Tomato plants are typically established in the field using transplants. This study aimed to determine if strains from field epidemics were coming from transplant facilities or resulted from local field outbreaks. The overall goal was to identify potential sources of inoculum and subsequently develop strategies to reduce carryover from transplant production to the field. Our results indicate that tomato producers should shift disease management efforts to transplant facilities to reduce disease in the field. Improved transplant health should reduce the likelihood of bacterial spot outbreaks and subsequently reduce pesticide usage in the field.

Alternaria spp. on apparently healthy apples as a potential inoculum source for moldy core development and the effect of resistant and susceptible apple cultivars

Apple (Malus domestica Borkh) is a major fruit crop in Chile. Among apple diseases, moldy core (MC) in the fruit has a high prevalence (16 to 46%) in susceptible cultivars. Apparently healthy flowers and fruit were sampled at six growth stages, from the pink bud to the mature fruit, from the susceptible ‘Oregon Spur’ and from the resistant ‘Granny Smith’ apples during two growing seasons. At least seven fungal genera were detected colonizing flowers and the fruit of both cultivars. Small-spored Alternaria spp. were the primary species identified throughout all the growing stages. Independently of the growth stages, sepals, stamens and carpels of flowers were similarly colonized by Alternaria spp. in the susceptible (‘Oregon Spur’) and resistant (‘Granny Smith’) apple cultivars. However, in fruits of 4–6 cm in diameter, big differences in the frequency of Alternaria spp. in fruit carpels were observed between susceptible and resistant apples, with 55% and 1%, respectively. At least five Alternaria spp. were identified at the species level using plasma membrane ATPase, namely, A. alternata, A. arborescens, A. limoniasperae, A. tenuissima in sect Alternaria, A. kordkuyana in sect. Pseudoalternaria and Alternaria sp. in sect. Infectoriae. All Alternaria isolates, from sepals, stamens and carpels, from apparently healthy flowers and fruits were pathogenic, producing MC symptoms in apples. These results suggest that apparently healthy flowers and fruits may act as an important inoculum source of Alternaria spp. for MC infections and also suggest that control strategies against MC using fungicide applications should be carried at flowering, from pink bud onwards.

Latent Infection of Valsa mali in the Seeds, Seedlings and Twigs of Crabapple and Apple Trees is a Potential Inoculum Source of Valsa Canker

A real-time quantitative PCR assay using a species-specific primer pair was developed to rapidly and accurately quantify Valsa mali, the causative pathogen of apple Valsa canker (AVC), in crabapple seeds, crabapple seedlings, apple twigs and apple seeds. Surveys were conducted in different regions, and crabapple or apple seeds were collected for V. mali detection by qPCR assay. Our results showed that 12.87% to 49.01% of crabapple seeds collected from different regions were positive for V. mali. The exopleura and endopleura were the two major areas of V. mali infection in crabapple seeds. The presence of V. mali infection in crabapple seeds was also confirmed by a high-throughput sequencing approach. With the growth of crabapple seedlings, the concentration of V. mali gDNA in crabapple seedlings gradually increased until eight or more leaf blades emerged. One-year-old twigs from an apple scion nursery were infected with V. mali, and only apple seeds from infected apple trees showing evident Valsa canker symptoms carried V. mali. In conclusion, this study reports that crabapple seeds and apple seeds carried V. mali as latent inoculum sources. V. mali infected not only apple tissues but also crabapple seedlings, which are the rootstocks of apple trees. This study indicated that the inoculum sources for AVC vary. Application of a novel qPCR assay can potentially improve the accuracy of early diagnosis, and is helpful to reveal the epidemic regularity of AVC.

Natural Hosts and Genetic Diversity of the Emerging Tomato Leaf Curl New Delhi Virus in Spain.

Knowledge about the host range and genetic structure of emerging plant viruses provides insights into fundamental ecological and evolutionary processes, and from an applied perspective, facilitates the design and implementation of sustainable disease control measures. Tomato leaf curl New Delhi virus (ToLCNDV) is an emerging whitefly transmitted begomovirus that is rapidly spreading and inciting economically important diseases in cucurbit crops of the Mediterranean basin. Genetic characterization of the ToLCNDV Mediterranean populations has shown that they are monophyletic in cucurbit plants. However, the extent to which other alternative (cultivated and wild) hosts may affect ToLCNDV genetic population structure and virus prevalence remains unknown. In this study a total of 683 samples from 13 cultivated species, and 203 samples from 24 wild species from three major cucurbit-producing areas of Spain (Murcia, Alicante and Castilla-La Mancha) from five cropping seasons (2012–2016) were analyzed for ToLCNDV infection. Except for watermelon, ToLCNDV was detected in all cultivated-cucurbit species as well as in tomato. Among weeds, Ecballium elaterium, Datura stramonium, Sonchus oleraceus, and Solanum nigrum were identified as alternative ToLCNDV plant hosts, which could act as new potential sources of virus inoculum. Furthermore, we performed full-genome deep-sequencing of 80 ToLCNDV isolates from different hosts, location and cropping year. Our phylogenetic analysis supports a Mediterranean virus population that is genetically very homogeneous, with no clustering pattern, and clearly different from Asian virus populations. Additionally, D. stramonium displayed higher levels of within-host genetic diversity than cultivated plants, and this variability appeared to increase with time. These results suggest that the potential ToLCNDV adaptive evolution occurring in wild plant hosts could serve as a source of virus genetic variability, thereby affecting the genetic structure and spatial-temporal dynamics of the viral population.

Bark and Grape Microbiome of Vitis vinifera: Influence of Geographic Patterns and Agronomic Management on Bacterial Diversity

In recent years, the concept of “microbial terroir” has been introduced in the frame of the more renowned notion of “vitivinicultural terroir,’ since several studies demonstrated that wine characteristics are related to regional microbial community compositions. Most of the existing research focused on grape berries microbiota, since it can directly impact wine quality. In this work we studied, for the first time through next-generation sequencing, the epiphytic bacterial community of vine bark and its relationships with grape microbiota. The study was carried out in two Italian wine appellations (situated in different regions) to explore the impact of biogeography, and the influence of two agronomical practices (biodynamic and conventional) was evaluated as well. Overall, our results show that grapevine bark harbors a rich epiphytic microbiota and displays a higher microbial biodiversity than grape berry. Moreover, this study suggests that geographic and anthropogenic factors impact both bark and grape bacteriomes, but to a different extent. The evidence of a “microbial terroir” seems to be even more marked in bark than in berries, possibly due to its permanence over time and to its physical proximity with soil. The importance of vine trunk bark, as potential source of inoculum for grapes and as interesting bacterial diversity habitat, is evidenced. This opens new fields of investigation, not only for researchers that aim at describing this little-known habitat within the vineyard, but also for stakeholders from the wine industry that want to understand the roles of microorganisms on the entire winemaking process, from vineyard to cellar.