Chitin in Strawberry Cultivation: Foliar Growth and Defense Response Promotion, but Reduced Fruit Yield and Disease Resistance by Nutrient Imbalances.

Strawberry cultivation is associated with high mineral fertilizer doses and extensive use of chemical plant protection products. Based on previous research, we expected that chitin application to peat substrate would increase the nutrient availability and activate the plant systemic defense response, resulting in higher strawberry yields and fewer disease symptoms. We set up two experiments in which the temporal variability and differences in initial nutrient concentrations of the growing media were taken into account. Chitin treatment resulted in the attraction of plant growth-promoting fungi toward the plant root, such as species from genera Mortierella and Umbelopsis. In addition, by the end of the experiments 87 mg of mineral nitrogen (N) per liter of substrate was mineralized, which can be related to the observed increase in plant shoot biomass. This, however, led to nutrient imbalances in plant shoots and fruit; N concentration in the leaves increased over 30%, exceeding the optimal range, while phosphorous (P) and potassium (K) deficiencies occurred, with concentrations lower than 50% of the optimal range. This may explain the decreased fruit yield and disease resistance of the fruit toward Botrytis cinerea. In contrast, chitin caused a clear defense priming effect in the strawberry leaves, with a strong induction of the jasmonic acid response, resulting in fewer foliar disease symptoms. Chitin causes positive effects on shoot growth and foliar disease resistance, but caution needs to be taken for nutrient imbalances leading to negative influences on root growth, fruit production, and disease susceptibility toward B. cinerea.[Formula: see text] Copyright © 2021 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Peat substrate amended with chitin modulates the N-cycle, siderophore and chitinase responses in the lettuce rhizobiome.

Chitin is a valuable peat substrate amendment by increasing lettuce growth and reducing the survival of the zoonotic pathogen Salmonella enterica on lettuce leaves. The production of chitin-catabolic enzymes (chitinases) play a crucial role and are mediated through the microbial community. A higher abundance of plant-growth promoting microorganisms and genera involved in N and chitin metabolism are present in a chitin-enriched substrate. In this study, we hypothesize that chitin addition to peat substrate stimulates the microbial chitinase production. The degradation of chitin leads to nutrient release and the production of small chitin oligomers that are related to plant growth promotion and activation of the plant's defense response. First a shotgun metagenomics approach was used to decipher the potential rhizosphere microbial functions then the nutritional content of the peat substrate was measured. Our results show that chitin addition increases chitin-catabolic enzymes, bacterial ammonium oxidizing and siderophore genes. Lettuce growth promotion can be explained by a cascade degradation of chitin to N-acetylglucosamine and eventually ammonium. The occurrence of increased ammonium oxidizing bacteria, Nitrosospira, and amoA genes results in an elevated concentration of plant-available nitrate. In addition, the increase in chitinase and siderophore genes may have stimulated the plant's systemic resistance.

The endophyte Verticillium Vt305 protects cauliflower against Verticillium wilt.

AIMS: To investigate the interaction between cauliflower and the isolate VerticilliumVt305, obtained from a field suppressive to Verticillium wilt of cauliflower, and to evaluate the ability of VerticilliumVt305 to control Verticillium wilt of cauliflower caused by V. longisporum. METHODS AND RESULTS: Single and combined inoculations of VerticilliumVt305 and V. longisporum were performed on cauliflower seedlings. Symptom development was evaluated, and fungal colonization was measured in the roots, hypocotyl and stem with real-time PCR. No symptoms were observed after single inoculation of VerticilliumVt305, although it colonized the plant tissues. Pre-inoculation of VerticilliumVt305 reduced symptom development and colonization of plant tissues by V. longisporum. CONCLUSIONS: VerticilliumVt305 is an endophyte on cauliflower plants and showed effective biological control of V. longisporum in controlled conditions. SIGNIFICANCE AND IMPACT OF THE STUDY: This work can contribute to the development of a sustainable control measure of V. longisporum in Brassicaceae hosts, which is currently not available. Additionally, this study provides evidence for the different roles of Verticillium species present in the agro-ecosystem.

Detection of Multiple Verticillium Species in Soil Using Density Flotation and Real-Time Polymerase Chain Reaction.

Wet sieving of soil samples, followed by plating on semi-selective medium and microscopic analysis, is the most commonly used technique to quantify microsclerotia-forming Verticillium species in soil. However, the method is restricted to small samples, does not allow easy differentiation between species, and takes several weeks to complete. This study describes an alternative method to test 100-g soil samples for three Verticillium species (V. tricorpus, V. dahliae, and V. longisporum) using density flotation-based extraction of microsclerotia followed by new real-time polymerase chain reaction (PCR) assays. Primers for these real-time PCR assays were designed to the ribosomal DNA internal transcribed spacer for V. tricorpus and the ß-tubulin gene for V. dahliae + V. longisporum and V. longisporum. Tests with artificially and naturally infested soils showed that the new method is reproducible and sensitive (0.1 to 0.5 microsclerotia/g soil), allows differentiation among the three species, and can be completed in one day. The results of the new method and the wet-sieving method were highly correlated for V. tricorpus (R2 = 0.78), but not for V. dahliae/V. longisporum, probably due to the loss of germinability of V. dahliae/V. longisporum microsclerotia during prolonged dry storage of the soil.

First Report of Pilidium concavum Causing Tan-Brown Rot on Strawberry Fruit in Belgium.

In April 2010, pink-orange spore masses that later turned brown were observed on 7 to 50% of the transplant lots during a routine screening of Belgian strawberry (Fragaria × ananassa, cv. Elsanta) for the latent presence of Colletotrichum acutatum using the petiole freeze method (4). These spore masses contained hyaline, canoe-shaped to allantoid conidia (mean size 7.5 × 1.8 µm), which is not consistent with C. acutatum spore morphology. Subsequently, a spore mass was transferred onto potato dextrose agar (PDA) and a gray-to-brown colony with whitish, aerial mycelium was produced, which is also not consistent with C. acutatum isolates. To identify the fungus, the ITS1-5.8S-ITS2 rDNA region was amplified by PCR and sequenced. The 485-bp region was 100% identical to that of Pilidium concavum specimen voucher BPI 1107275 (GenBank Accession No. AY487094). P. concavum (Desm.) Höhn. (synanamorph Hainesia lythri; teleomorph Discohainesia oenotherae) is a pathogen of strawberry causing tan-brown rot of fruit and is a common secondary invader of roots and dead strawberry plant parts (3). A recent strain of P. concavum from strawberry, isolate UPL 50, obtained from Brazil (L. Zambolim, Univ. Fed. de Viçosa, personal communication) showed similar colony, microscopic (mean spore size of 6.8 × 1.8 µm), and molecular (ITS sequence 98% identical to that of P. concavum specimen voucher BPI 1107275) features as the Belgian isolate. Pathogenicity tests were conducted on mature strawberry fruits by submerging 15 fruits per isolate for 3 min in a conidial suspension (2 × 106 conidia ml-1 of water) obtained from a 2-week-old colony on PDA. Controls were submerged in sterile distilled water. The inoculated fruits were incubated in a moist chamber at 25°C. Sunken, yellowish brown lesions with pink and later orange-brown spore masses were observed starting 3 days after inoculation on 88 and 94% of the fruit for the Brazilian and Belgian isolate, respectively. The control fruits remained healthy. The fungal isolates were reisolated from symptomatic fruits and their identity was confirmed based on morphological features. During a strawberry field survey in July 2010 in Sint-Truiden (Belgium), lesions typical of those described above were observed on eight strawberry fruits (cv. Elsanta). The fungus was isolated from the symptomatic tissue of two fruits and characterized as described above. Since P. concavum was latently present on strawberry transplants and caused disease on the fruits in the field, we conclude that P. concavum is a potential threat for Belgian strawberry production. Moreover, no strawberry cultivars with resistance to the pathogen have been reported. The disease has previously been reported on strawberry in South America and Poland (1,2), but to our knowledge, this is the first report of P. concavum on strawberry in Belgium. Although the spore and colony morphology of P. concavum is different from C. acutatum, the spore masses of P. concavum can easily be confused with the spore masses of C. acutatum when using the freeze method. This suggests the need for microscopic analysis of these spore masses during routine analyses. References: (1) L. Cedeno et al. Interciencia 26:113, 2001. (2) U. P. Lopes et al. New Dis. Rep. 21:7, 2010. (3) J. L. Maas. Compendium of Strawberry Diseases. The American Phytopathological Society St. Paul, MN, 1998. (4) J. C. Mertely and D. E. Legard. Plant Dis. 88:407, 2004.

Phenotypic and genetic diversity of rice seed-associated bacteria and their role in pathogenicity and biological control.

AIMS: To study the phenotypic and genetic diversity of culturable bacteria associated with rice seed and to asses the antagonistic and pathogenic potential of the isolated bacteria. METHODS AND RESULTS: Seed of rice cultivar PSBRc14 was collected from farmers' fields of irrigated lowland in southern Luzon, Philippines. Isolations of distinct colonies yielded 498 isolates. Classification of the isolates according to similarities in cellular characteristics, whole-cell fatty acid composition, and colony appearance differentiated 101 morphotype groups. Predominant bacteria were Coryneform spp., Pantoea spp. and Pseudomonas spp. Other bacteria regularly present were Actinomycetes spp., Bacillus pumilus, B. subtilis, Burkholderia glumae, Enterobacter cloacae, Paenibacillus polymyxa, Staphylococcus spp. and Xanthomonas spp. The genetic diversity among isolates was assessed by BOX-PCR fingerprinting of genomic DNA and represented 284 fingerprint types (FPTs). Most FPTs (78%) were not shared among samples, while eight FPTs occurred frequently in the samples. Seven of these FPTs also occurred frequently in a previous collection made from rainfed lowlands of Iloilo island, Philippines. Sixteen per cent of the isolates inhibited in vitro the mycelial growth of the rice pathogens Rhizoctonia solani and Pyricularia grisea, whereas 4% were pathogens identified as Burkholderia glumae, Burkholderia gladioli and Acidovorax avenae ssp. avenae. CONCLUSIONS: This study reveals a broad morphological and genetic diversity of bacteria present on seed of a single rice cultivar. SIGNIFICANCE AND IMPACT OF THE STUDY: This line of work contributes to a better understanding of the microbial diversity present on rice seed and stresses its importance as a carrier of antagonists and pathogens.

Biosurfactants are involved in the biological control of Verticillium microsclerotia by Pseudomonas spp.

AIMS: To examine the effect of previously described bacterial antagonists on the viability of Verticillium microsclerotia in vitro and to elucidate the possible modes of action of bacterial strains in the suppression of Verticillium microsclerotia viability. METHODS AND RESULTS: A microplate assay was developed to test the suppressive effect of well-defined Pseudomonas spp. on the viability of Verticillium microsclerotia in vitro. Experiments using phenazine- and biosurfactant-deficient mutants indicated that biosurfactants and phenazine-1-carboxylic acid play a role in the suppression of microsclerotia viability by Pseudomonas spp. In addition, microsclerotia colonization tests revealed that Pseudomonas spp. are able to colonize the surface of the microsclerotia, but not the inner matrix. Growth response curves showed that the population levels of Pseudomonas spp. increased when they were in the vicinity of Verticillium microsclerotia, indicating that Pseudomonas spp. may utilize nutrients from the microsclerotia for their growth. CONCLUSIONS: Pseudomonas spp. seem to be good candidates for Verticilllium microsclerotia biocontrol. Biosurfactant production is one of the main mechanisms involved in their mode of action. SIGNIFICANCE AND IMPACT OF THE STUDY: This line of work may contribute to a better understanding of biological control agents and their working mechanisms.

Verticillium wilt of cauliflower in Belgium.

Since several years commercial cauliflower in Belgium is severely affected by a vascular wilt disease. Plants wilt and a vascular discoloration of the cauliflower stems can be observed. The pathogen causes significant yield and quality losses. The first objective of this study was to isolate and characterise the causal agent of this disease. Verticillium species could be isolated out of the vascular tissue of symptomatic cauliflower plants. Morphological and physiological characteristics indicated that the isolates from cauliflower could be identified as the new and brassica-related species V. longisporum. Secondly, the susceptibility of several cauliflower cultivars to V. longisporum was evaluated by way of field experiments in naturally infested soil. At harvest, vascular discoloration was evaluated by a visual score. These experiments pointed to considerable differences between cultivars in their susceptibility to V. longisporum. The final goal of this study was to find a way to control the wilt disease on cauliflower. A field experiment showed that vascular discoloration of cauliflower was significantly reduced when soil was incorporated with broccoli residues, two Brassica green manures (B. juncea or B. nigra) or two commercial biological products based on Talaromyces flavus or Trichoderma sp..