Mark Mazzola

Soil immune responses

Soil microbiomes may be harnessed for plant health Soil microorganisms are central to the provision of food, feed, fiber, and medicine. Engineering of soil microbiomes may promote plant growth and plant health, thus contributing to food security and agricultural sustainability (1, 2). However, little is known about most soil microorganisms and their impact on plant health. Disease-suppressive soils offer microbiome-mediated protection of crop plants against infections by soil-borne pathogens. Understanding of the microbial consortia and mechanisms involved in disease suppression may help to better manage plants while reducing fertilizer and pesticide inputs.

Characterization of fungi (Fusarium and Rhizoctonia) and oomycetes (Phytophthora and Pythium) associated with apple orchards in South Africa

Several species of fungi and oomycetes including Fusarium, Rhizoctonia, Phytophthora and Pythium have been reported as root pathogens of apple where they contribute to a phenomenon known as apple replant disease. In South Africa, little is known about specific species in these genera and their pathogenicity toward apple. Therefore, these aspects were investigated along with the development and optimization of qPCR tests for detection and quantification of the most virulent oomycete species. In eight investigated orchards, the oomycete Phythophthora cactorum was widely distributed, while nine Pythium species were differentially distributed among the orchards. Pythium irregulare was the most widely distributed and the most virulent species along with P. sylvaticum, P. vexans and Ph. cactorum. Seven binucleate Rhizoctonia anastomosis groups (AGs) were also differentially distributed among the orchards, with the majority appearing to be non-pathogenic while certain AG-I and AG-F isolates exhibited low virulence on apple. In the genus Fusarium, F. oxysporum was widely distributed, but isolates were non-pathogenic. Fusarium solani and F. avenaceum were less frequently encountered, with only some isolates having low virulence. qPCR data obtained from seedling roots inoculated with the most virulent Pythium species (P. irregulare, P. sylvaticum and P. vexans) and the genus Phytophthora were not always reproducible between trials, or isolates of the same species. In general, seedling growth inhibition was associated with the presence of a low amount of pathogen DNA (±40xa0fgxa0μl−1 to 2xa0pgxa0μl−1) in roots. Pythium irregulare, although having the lowest DNA concentrations in roots, was the only species for which a significant negative correlation was found between seedling weight and pathogen DNA concentration.

Deciphering microbial landscapes of fish eggs to mitigate emerging diseases.

Animals and plants are increasingly suffering from diseases caused by fungi and oomycetes. These emerging pathogens are now recognized as a global threat to biodiversity and food security. Among oomycetes, Saprolegnia species cause significant declines in fish and amphibian populations. Fish eggs have an immature adaptive immune system and depend on nonspecific innate defences to ward off pathogens. Here, meta-taxonomic analyses revealed that Atlantic salmon eggs are home to diverse fungal, oomycete and bacterial communities. Although virulent Saprolegnia isolates were found in all salmon egg samples, a low incidence of Saprolegniosis was strongly correlated with a high richness and abundance of specific commensal Actinobacteria, with the genus Frondihabitans (Microbacteriaceae) effectively inhibiting attachment of Saprolegniato salmon eggs. These results highlight that fundamental insights into microbial landscapes of fish eggs may provide new sustainable means to mitigate emerging diseases.

Characterisation and detection of Pythium and Phytophthora species associated with grapevines in South Africa

Replant and decline diseases of grapevines not only cause quantitative and qualitative yield losses, but also result in extra costs when vineyards have to be replanted. This study investigated the role of Pythium and Phytophthora in the decline syndrome in South Africa by determining (1) the species associated with nursery and established vines, and (2) pathogenicity of Ph. sp. niederhauserii and P. vexans relative to known grapevine pathogens. Quantitative real-time PCR (qPCR) assays were also developed for detection of the most prevalent oomycete groups. In total, 26 Pythium and three Phytophthora species were identified from grapevine nurseries and established vineyards. The most common infections in sampled nursery vines were caused by P. vexans (16.7%), followed by P. ultimum var. ultimum (15.0%) and P. irregulare (11.7%). In established vineyards, P. irregulare (18.0%) and P. vexans (6.2%) were also among the three most prevalent species, along with P. heterothallicum (7.3%). Three Phytophthora species were also identified from the sampled established vines, of which Ph. cinnamomi (5.1%) was predominant, followed by Ph. sp. niederhauserii (1.1%). In established vineyards a higher incidence and more diverse species composition was observed in spring and winter, than in summer. Pathogenicity studies showed that some Ph. sp. niederhauserii and P. vexans isolates were as aggressive as the known grapevine pathogens Ph. cinnamomi and P. irregulare. Sensitive qPCR assays were developed for the detection of P. ultimum var. ultimum, P. irregulare, P. vexans and the genus Phytophthora. These assays will be invaluable in limiting pathogen dispersal through screening of nursery material. This is especially important since pathogenic species were also isolated from healthy looking vines in nurseries.

Molecular analyses of Pythium irregulare isolates from grapevines in South Africa suggest a single variable species

The Pythium irregulare species complex is the most common and widespread Pythium spp. associated with grapevines in South Africa. This species complex has been subdivided into several morphological and phylogenetic species that are all highly similar at the sequence level [internal transcribed spacer (ITS) and cytochrome c oxidase (cox) regions]. The complex includes Pythium regulare and Pythium cylindrosporum, which are morphologically distinct, and P. irregulare sensu stricto (s.s.) and Pythium cryptoirregulare, which are morphologically similar. The aim of the current study was to determine whether 50 South African grapevine P. irregulare isolates represented more than one phylogenetically distinct species. The isolates were characterised using nuclear (ITS and β-tubulin) and mitochondrial (cox1 and cox2) gene region phylogenies and two isozyme loci [glucose-6-phosphate isomerase (Gpi) and malate dehydrogenase (Mdh-1)]. Some of the gene sequence data were difficult to interpret phylogenetically, since some isolates contained two or more polymorphic ITS copies within the same isolate (intra-isolate variation) that clustered into different ITS sub-clades, i.e. the P. irregulare s.s. and P. cryptoirregulare sub-clades. The molecular data furthermore only revealed the presence of one phylogenetic species, P. irregulare. Morphological analyses of a subset of the isolates confirmed that the isolates were P. irregulare, and further showed that the P. cylindrosporum ex-type strain formed typical P. irregulare oogonia, and not the previously reported distinct elongated oogonia. Some of the molecular analyses suggested the occurrence of outcrossing events and possibly the formation of aneuploids or polyploids since (i) the nuclear and mitochondrial gene data sets were incongruent, (ii) polymorphic ITS copies were present within the same isolate, (iii) heterozygosities were observed in the β-tubulin gene and Gpi and Mdh-1 loci in some isolates and (iv) more than two β-tubulin alleles were detected in some isolates. Altogether, the data suggest that P. irregulare, P. cryptoirregulare, P. cylindrosporum, and possibly P. regulare should be synonimised under the name P. irregulare.

Bioremediation of Soils by Plant–Microbe Systems

Abstract Sustainable ecosystems can be designed to eliminate environmental toxins and reduce pathogen loads through the direct and indirect consequences of plant and microbial activities. We present an approach to the bioremediation of disturbed environments, focusing on petroleum hydrocarbon (PHC) contaminants. Treatment consists of incorporating a plant-based amendment to enhance ecosystem productivity and physiochemical degradation followed by the establishment of plants to serve as oxidizers and foundations for microbial communities. Promising amendments for widespread use are entire plants of the water fern Azolla and seed meal of Brassica napus (rapeseed). An inexpensive byproduct from the manufacture of biodiesel and lubricants, rapeseed meal is high in nitrogen (6% wt/wt), stimulates >100-fold increases in populations of resident Streptomyces species, and suppresses fungal infection of roots subsequently cultivated in the amended soil. Synergistic enzymatic and chemical activities of plant and microbial metabolism in root zones transform and degrade soil contaminants. Emphasis is given to mechanisms that enable PHC functionalization via reactive molecular species.

Anaerobic soil disinfestation is an alternative to soil fumigation for control of some soilborne pathogens in strawberry production

Alternatives to soil fumigation are needed for soil-borne disease control. Our goal was to test anaerobic soil disinfestation (ASD) as an alternative to soil fumigation for control of critical soil-borne pathogens in California strawberry production. Controlled environment experiments were conducted at 25°C and 15°C testing different materials as carbon sources for ASD using soil inoculated with Verticillium dahliae. Field trials were conducted in three locations comparing ASD with 20Mg ha−1 rice bran (RB) against fumigated and untreated controls, steam, mustard seed meal and fish emulsion. In ASD treated soils, temperature and anaerobicity were critical for control of V. dahliae, but multiple carbon inputs reduced inoculum by 80-100%. In field trials, ASD with RB provided control of a number of pathogens, and in three of four trials produced marketable fruit yields equivalent to fumigation. Little weed control benefit from ASD was found. ASD with RB also induced changes in the soil microbiome that persisted through the growing season. When equivalent yields were obtained, net returns above harvest and treatment costs with ASD RB were 92–96% of those with bed fumigation based on average prices over the previous 5 years. ASD can be a viable alternative for control of some soil-borne pathogens. Growers are adopting ASD in CA strawberry production, but research to determine optimal soil temperatures, anaerobicity thresholds and carbon sources for effective control of specific pathogens is needed. n nThis article is protected by copyright. All rights reserved.

Targeted Metabolic Profiling Indicates Apple Rootstock Genotype-Specific Differences in Primary and Secondary Metabolite Production and Validate Quantitative Contribution From Vegetative Growth

Previous reports regarding rhizodeposits from apple roots are limited, and complicated by microbes, which readily colonize root systems and contribute to modify rhizodeposit metabolite composition. This study delineates methods for collection of apple rhizodeposits under axenic conditions, indicates rootstock genotype-specific differences and validates the contributions of vegetative activity to rhizodeposit quantity. Primary and phenolic rhizodeposit metabolites collected from two apple rootstock genotypes, G935 and M26, were delineated 2 months after root initiation by utilizing gas chromatography/liquid chromatography—mass spectrometry (GC/LC-MS), respectively. Twenty-one identified phenolic compounds and 29 sugars, organic acids, and amino acids, as well as compounds tentatively identified as triterpenoids were present in the rhizodeposits. When adjusted for whole plant mass, hexose, erythrose, galactose, phloridzin, kaempferol-3-glucoside, as well as glycerol, and glyceric acid differed between the genotypes. Phloridzin, phloretin, epicatechin, 4-hydroxybenzoic acid, and chlorogenic acid were among the phenolic compounds found in higher relative concentration in rhizodeposits, as assessed by LC-MS. Among primary metabolites assessed by GC-MS, amino acids, organic acids, and sugar alcohols found in relatively higher concentration in the rhizodeposits included L-asparagine, L-cysteine, malic acid, succinic acid, and sorbitol. In addition, putative ursane triterprenoids, identified based on accurate mass comparison to previously reported triterpenoids from apple peel, were present in rhizodeposits in high abundance relative to phenolic compounds assessed via the same extraction/instrumental method. Validation of metabolite production to tree vegetative activity was conducted using a separate set of micropropagated trees (genotype MM106) which were treated with a toxic volatile compound (butyrolactone) to inhibit activity/kill leaves and vegetative growth. This treatment resulted in a reduction of total collected rhizodeposits relative to an untreated control, indicating active vegetative growth contributes to rhizodeposit metabolites. Culture-based assays indicated an absence of bacterial or fungal endophytes in roots of micropropagated G935 and M26 plants. However, the use of fungi-specific primers in qPCR indicated the presence of fungal DNA in 30% of the samples, thus the contribution of endophytes to rhizodeposits cannot be fully eliminated. This study provides fundamental information for continued research and application of rhizosphere ecology driven by apple rootstock genotype specific rhizodeposition.

Control of bull’s-eye rot of apple caused by Neofabraea perennans and Neofabraea kienholzii using pre- and postharvest fungicides

Bulls-eye rot is a major postharvest disease of apple caused by several fungi belonging to the Neofabraea and Phlyctema genera. Chemical control of these fungi is a crucial component of disease management for apples that are conventionally grown. The efficacy of several preharvest and postharvest applied fungicides were evaluated to identify effective chemistries that can control bulls-eye rot incited by Neofabraea perennans and N. kienholzii on apples. In general, the preharvest fungicide thiophanate-methyl was found to be effective at reducing disease caused by N. perennans and N. kienholzii. Two postharvest fungicides, thiabendazole and pyrimethanil, also provided disease control that was far superior to other chemical compounds evaluated in this study. The efficacy of thiabendazole and pyrimethanil was unaffected by application method (fungicide dip compared with thermofog). Despite providing satisfactory control of bulls-eye rot, integration of these three chemicals into disease management programs should proceed judiciously with consideration of their impact on the development of fungicide resistance and influence on diversity in populations of apple postharvest pathogens.