Plant Disease

Beyond the significant impact of Cassava witches broom disease (CWBD), caused by the fungus Rhizoctonia (syn. Ceratobasidium) theobromae on cassava cultivation in French Guiana and Brazil, this disease also poses a potential threat to cacao trees in the region, since the fungus is responsible for Vascular Streak Dieback (VSD) of cacao in South East Asia. Cross-pathogenicity trials were conducted in several cassava fields in French Guiana by planting young cacao plants adjacent to diseased cassava plants. Vascular necrosis was observed in some cacao plants, and the presence of R. theobromae in the cacao tissues was confirmed through PCR diagnostics using primers specific to the fungus. Sequence analysis indicated 100% similarity between samples from both hosts and 97.53 to 99.74% identity with R. theobromae isolates previously reported from cassava in the Americas and Southeast Asia. Additionally, symptomatic cacao in a mixed cacao-cassava farm yielded R. theobromae-positive PCR results, suggesting a natural infection. Ongoing work includes artificial inoculations and controlled cross-pathogenicity trials under screenhouse conditions to attempt reproduction of the symptoms. While current data do not yet establish definitive causality, the findings indicate potential host jump and warrant rapid communication to researchers, policy makers, and farmers to safeguard cacao production and Theobroma biodiversity in the Amazon region.

Whitebark pine (Pinus albicaulis), a wide-ranging high-elevation conifer in western North America, is listed as threatened in the U.S. and as endangered in Canada. A major threat to whitebark pine is the non-native, invasive white pine blister rust disease, caused by the fungal pathogen Cronartium ribicola. In many pathosystems (including white pine blister rust), seedling inoculation trials are used to identify parent trees with genetic resistance. However, many of these trials use only one spore density for inoculation, and little information exists on the effectiveness of quantitative disease resistance (QDR) under varying spore densities and the corresponding implications for field performance. In this study, we examine the levels of infection and survival present within six whitebark pine seedling families previously rated for QDR (three susceptible and three resistant families) under six widely varying inoculum densities. The susceptible families showed very high infection and mortality at all inoculum densities, while performance of the resistant families varied with spore density treatment. The information gathered from the study will be useful in updating the projections of the future of whitebark pine populations under field conditions in areas of different rust hazard. The results also serve as a caution to those working in other pathosystems where seedling inoculation trials based on one spore density level are used to rate the resistance level of parent trees and their associated progeny.

Soft rot Pectobacteriaceae (SRP) are among the most economically important plant pathogenic bacteria and are especially known to be problematic in potato production. The epidemiology of disease transmission has been investigated for almost a century, and several aspects have been highlighted as plausible infection routes. However, it is generally accepted that the major source of disease is the latently infected mother tuber, but several parameters are still influencing disease prevalence including contaminated equipment, soil water status as well as temperature. Management of the disease is limited to hygiene practices, dry storage and seed certification systems but several studies have also proven biocontrol agents such as bacteriophages (phages) as promising tools. Despite the severity of SRP on potato production, little is known about the genetic diversity of SRPs in Denmark, and since only few isolates are available, the possibility to design a broadly effective phage cocktail is limited. Here we describe a three-year field study utilizing an agri-citizen science approach where Danish farmers provided symptomatic potato plants or tubers, together with metadata such as date, location, potato variety and origin. By using whole genome sequencing (Illumina and Nanopore) together with metadata we were able to investigate and monitor the epidemiological disease spread across the country using 103 complete genomes, sampled across all three years. In this study we provide epidemiological evidence of disease origins and a suite of phages that could be used as a biocontrol tool for early disease intervention. Our results revealed several clonal clades across diverse locations (SNPs < 20) which strongly indicate common origin. A total of 17 Pectobacterium phages were tested and did target > 80% of clonal clades. Based on the clonality across the soft rot isolates we propose the possibility to set in early on using phages targeting strains relevant for soft rot development, with the possibility of a surveillance program together with customizing the phage preference.

Cucurbit yellow vine disease (CYVD), caused by the bacterium Serratia ureilytica, is a phloem-associated disease of cucurbits. This study characterized the spatial and temporal distribution of S. ureilytica in Cucurbita pepo cultivar Delicata plants under greenhouse conditions using a GFP-tagged isolate (P01). Seedlings were sampled weekly for four weeks. Transverse sections from the stem, petiole, leaf, shoot apex, and root were imaged by laser scanning confocal and fluorescent dissecting microscopy. In parallel, bacterial abundance in each plant tissue was assessed by quantifying colony-forming units (CFU) via droplet plating over a 4-week time course. Across plant tissues and time points, S. ureilytica fluorescent signal was primarily concentrated in the inner and outer periphery of the bicollateral vascular bundles, with higher magnification images revealing mainly symplastic localization within phloem-associated cells. Consistent with the imaging results, bacterial quantification data showed a high abundance of CFUs in the main stem across weeks, with an irregular pattern of presence in the distal tissues at later time points. These results suggest that S. ureilytica is predominantly localized within phloem-associated cells and spreads both acropetally and basipetally during infection.

Australia is the largest producer of Pyrethrum (Tanacetum cinerariifolium) globally. Amongst the constraints on production are the fungal pathogens Didymella tanaceti and Stagonosporopsis tanaceti, which pose a significant threat to the industry, causing substantial yield losses. While the infection biology of S. tanaceti is well characterised, knowledge of D. tanaceti and its potential interaction with S. tanaceti on plants remains limited, hindering disease management. We developed fluorescently labelled strains of both pathogens via Agrobacterium tumefaciens-mediated transformation (ATMT). Binary vectors carrying the mNeonGreen or tdTomato fluorescent protein genes were introduced into D. tanaceti and S. tanaceti, respectively, and expression of the fluorescent proteins was confirmed by microscopy. Genome sequencing revealed single-copy T-DNA insertions in all transformants, with minor genomic rearrangements at insertion sites. Detached leaf assays demonstrated that transformed strains retained pathogenicity, producing disease symptoms indistinguishable from those of the wild type. These fluorescently labelled variants enabled detailed visualisation of D. tanaceti infection biology and its interactions with S. tanaceti, including co-infection dynamics. Co-infection assays using fluorescent strains further facilitated simultaneous visualisation and differentiation of both pathogens within host tissues. Importantly, these tools also allowed the first description of the early stages of infection by D. tanaceti in pyrethrum leaves. This study represents the first successful transformation of D. tanaceti and S. tanaceti, providing valuable resources to investigate their infection processes.

Ralstonia pseudosolanacearum (Rps) belongs to the Ralstonia solanacearum species complex (RSSC). It is a vascular pathogen that causes lethal bacterial wilt disease in many plants, including tomato and eggplant. In this study, we infiltrated tomato leaves with the phytopathogenic bacterium at 109 CFU/mL and observed the development of necrotic scars in the infiltrated area at 48 hours post-infiltration. Interestingly, this response was followed by petiole bending toward the ground of the compound leaf. This was followed by the gradual senescence of the infiltrated leaflet only. In addition, the terminal leaflet infiltrated with the pathogen exhibited epinasty. None of the above symptoms were observed in leaves infiltrated with the known virulent deficient hrpB::{Omega} mutant. Surprisingly, all of the above symptoms were observed in leaves infiltrated with another well-known virulence-deficient mutant phcA::{Omega}. It indicated that the necrotic lesion caused in tomato leaves was hrp-dependent. Infiltration in eggplant leaves caused necrotic scarring and leaf senescence, which were relatively delayed. Necrotic scarring without petiole bending or senescence in tomato leaves was also observed due to infiltration of Pseudomonas aeruginosa SPT08, a tomato endophyte having plant growth promotion activity. The patho-phenotypes such as petiole bending, epinasty, and senescence observed in the case of tomato in this study were not reported earlier. We believe these phenotypes produced in tomato after leaf infiltration may be useful to study the virulence of this pathogen.

Apple blotch, caused by Diplocarpon coronariae, is an increasingly important fungal disease that leads to premature leaf fall and significant yield losses in apple orchards. Breeding resistant cultivars offers a sustainable strategy to reduce disease impact, as all commercial apple cultivars are susceptible to this pathogen. This study aimed to investigate the disease resistance of Malus baccata Jackii-derived offspring to D. coronariae through artificial inoculation and to identify loci associated with resistance. Simple interval mapping was performed using phenotypic and genotypic data from 122 individuals of an F1 population (Idared x M. baccata Jackii), together with analyses of M. baccata Jackii-derived open-pollinated populations. Our results indicate that resistance to apple blotch is a complex, polygenic trait, with four important QTLs identified on linkage groups 1, 2, 12 and 13. Disease severity was strongly affected by inoculum, phenotyping method and environmental factors. These findings have direct implications for apple breeding programmes aimed at developing apple blotch-resistant cultivars.

2.Hairy root disease (HRD), caused by rhizogenic Agrobacterium, is an economically important disease affecting hydroponic tomato (Solanum lycopersicum L.) production worldwide. HRD-affected plants show extensive root proliferation, resulting in decreased energy expenditure towards fruit production. Host plant susceptibility to rhizogenic Agrobacterium is typically evaluated through artificial wounding-based infection bioassays. However, under natural infection settings, rhizogenic Agrobacterium can induce disease symptoms without deliberate, artificial wounding. We developed a soil-based, non-wounding bioassay that closely mimics natural rhizosphere interactions and permits quantitative and qualitative assessment of HRD symptoms. The assay measured root dry weight, documented agravitropic root development typical of HRD and confirmed in planta T-DNA gene expression using reverse transcriptase quantitative polymerase chain reaction (RT-qPCR). We used this bioassay to evaluate disease symptoms towards rhizogenic Agrobacterium in tomato cv. Moneymaker and the rootstocks Optifort, Maxifort, and Arnold. Optifort and Maxifort exhibited significantly higher root biomass than Arnold and Moneymaker, indicating more pronounced symptom development. The bioassay also differentiated virulence levels amongst various rhizogenic Agrobacterium strains isolated from HRD-affected plants. Together, these results show that our soil-based bioassay provides a robust and ecologically relevant platform for screening tomato genotypes and comparing virulence levels of rhizogenic Agrobacterium strains supporting resistance breeding and disease management efforts.

Leaf diseases pose a serious threat to faba bean production. Leaf blotch of faba bean, caused by Alternaria spp., has become increasingly widespread and destructive in several countries. Leaf diseases pose a serious threat to faba bean production. The infection of plant by pathogens can be influenced by various factors associated with the host plant, environmental conditions and presence of other microorganisms. The phyllosphere and endosphere play a critical role in plant health and disease development. This study aimed to evaluate the factors shaping the structure and diversity of fungal communities associated with faba beans. Plant samples were collected in 2004 from two intensively managed faba bean production fields in the central region of Latvia. Fungal assemblages were characterized using an ITS region metabarcoding approach based on Illumina MiSeq sequencing. Among the assigned amplicon sequence variant (AVS), 65% belonged to the phylum Ascomycota, while approximately 4% were classified as Basidiomycota. Alternaria and Cladosporium were the dominant genera across samples. The alfa and beta diversities of fungal communities was higher during flowering of faba beans to compare with ripening. The higher abundance of Basidiomycota yeasts were observed during flowering, in contrast, Cladosporium genus was significantly more abundant during ripening. Alternaria DNA was found on leaves that showed no symptoms of the disease. The diversity and composition of fungal communities were significantly influenced by sampling time and presence of leaf blotch, caused by Alternaria spp.

Soybean (Glycine max [L.] Merr.) productivity is frequently compromised by soil-borne pathogens. Macrophomina phaseolina (Mp), the causal agent of charcoal rot, can produce important soybean yield losses especially when hot and dry weather prevails. Integrating biological control agents with chemical seed treatments represents a promising strategy for improving disease management. This study aimed to (i) assess the in vitro compatibility of Trichoderma koningiopsis with commercial fungicide seed treatments, and (ii) evaluate the field performance of T. koningiopsis, alone or combined with compatible fungicides, across three soybean growing seasons. Compatibility assays revealed fungicide-specific effects, with Acronis(R) classified as non-fungitoxic and Topseed Extra as moderately fungitoxic. Across field seasons, Mp inoculation reduced seedling emergence, while several seed treatments improved emergence compared to the inoculated control, however, treatment effects varied markedly among years. Disease severity did not differ significantly among treatments in any season, and yield responses were strongly modified by environmental conditions rather than treatment effects. Temperature-response assays showed that T. koningiopsis exhibited optimal growth between 28 to 30{degrees}C and complete inhibition above 40{degrees}C, indicating high thermal sensitivity. The results demonstrate that T. koningiopsis can be integrated with compatible fungicides and may enhance early stand establishment under favorable conditions, but its field performance is strongly limited by high temperatures. These findings highlight the importance of environmental conditions when biological seed treatments are used.

Breeding disease-resistant varieties (DRV) is a central strategy for reducing reliance on phytosanitary products. However, the successful deployment and long-term durability of these cultivars rely on acquiring field data across diverse production conditions, a step that remains frequently neglected, especially in perennial crops. Since 2018, the OSCAR observatory, a network of vineyard plots planted in France with varieties resistant to downy and powdery mildew, the two major pathogens of grapevine, has aimed to close this gap for viticulture. The observatory comprises over 199 commercial plots, covering 127 hectares across diverse agroclimatic conditions, all managed by winegrowers under their own production practices. The observatory currently monitors 30 disease-resistant grapevine varieties, tracking both their agronomic performance and the dynamics of key pathogens. Since 2018, while phytosanitary treatments have been reduced by an average of 79% compared to conventional plots, the incidence of downy and powdery mildew, remain low, even in years highly conducive to these diseases. However, the long-term survey also highlights the decline in efficacy of some resistances to downy mildew and the emergence of black rot, a disease effectively controlled by conventional phytosanitary programs. Beyond acting as a rapid warning system for resistance breakdown, the observatory promotes sustainable disease management in viticulture. It provides valuable insights to winegrowers on effective DRV management. It also delivers actionable feedback to breeders to guide more durable DRV breeding strategies. Highlights- OSCAR observatory monitors 199 plots of grapevine disease resistant varieties (DRV) - Grapevine DRV cuts fungicide uses by 79% while maintaining good disease control - Some resistances efficacy declines against downy mildew, but not powdery mildew - Black rot, a disease usually controlled by fungicide, is rising in OSCAR plots OSCAR provides useful feedback to breeders and winegrowers on DRV management

The fungal pathogen Zymoseptoria tritici poses a major global threat to wheat production, causing severe yield losses and necessitating intensive and costly fungicide applications. The increasing demand for durable genetic resistance has intensified interest in quantitative resistance loci, particularly those exhibiting multi-stage resistance (MSR), which suppress pathogen development continuously throughout the wheat life cycle. Many previously effective resistance genes are now showing declining efficacy, underscoring the urgent need for novel and long-lasting sources of resistance. In this study, we report the identification and genetic mapping of two quantitative resistance loci that address this need. The first locus, designated Stb23, is a major QTL on chromosome 1DS, with LOD scores exceeding 9 and explaining 6-36% of phenotypic variation at the seedling stage and 2-16% at the adult-plant stage. The second locus, designated Stb24, is a major QTL on chromosome 3DL, with LOD scores of approximately 10 and accounting for 11-30% of seedling-stage variation and 9-23% of adult-plant variation. Furthermore, two tightly linked KASP markers-snp_1D1217527 for Stb23 and snp_3D1077880 for Stb24-were developed and validated across three popular Australian bread wheat cultivars, providing practical tools for deploying these loci in breeding programs targeting improved resistance to Z. tritici. Key messageTwo significant major-effect resistance loci on chromosomes 1DS (proposed as Stb23) and 3DL (proposed as Stb24) were identified and characterized. Two tightly linked KASP markers with these loci were also discovered and validated for molecular-assisted breeding programs.

Cercospora species associated with soybean cause Cercospora leaf spot and purple seed stain, which are major diseases affecting soybean production worldwide and can lead to significant yield and seed quality losses. However, unstable and poor sporulation under laboratory conditions remains a critical challenge, hindering the recovery of genetically homogeneous isolates and the establishment of standardized experimental protocols. These limitations further restrict our understanding of the biology, epidemiology, and pathogenicity of these pathogens. In this study, we developed specialized legume-based culture media derived from soybean and pea tissues to mimic host-associated environmental conditions. We compared the sporulation efficacy of these media with commonly used artificial media, including potato dextrose agar (PDA) and V8 juice agar. Our results demonstrated that legume-based media consistently supported higher levels of sporulation than PDA and V8 across multiple strains, although conidial yields varied depending on the strain and medium concentration. Transcriptional analysis of sporulation-related genes revealed that while abaA, wetA, and steA did not show significant differential expression among media, velB exhibited distinct medium-dependent expression patterns. Further evaluation using additional field isolates confirmed that legume-based media provide a more reliable method for inducing sporulation than PDA. Overall, legume-based media represent a practical and effective approach for promoting sporulation in soybean-associated Cercospora species under laboratory conditions.

Phytophthora cinnamomi, the causal agent of Phytophthora root rot (PRR), poses a persistent threat to the United States avocado industry, the top domestic producer and consumer. Avocado growers are facing clonal A2 P. cinnamomi populations challenging their current PRR control methods. In this study, we characterized 125 isolates collected from orchards in California, Florida, Hawaii, Texas, and Puerto Rico for radial growth per day, optimal growth temperature, in vitro fungicide sensitivity, and virulence on DAnjou pear fruit and UC2001 avocado seedlings. Across all isolates, optimal growth occurred most frequently at a range from 22 to 25{degrees}C; however, a subset of isolates from Hawaii, Florida, and California exhibited higher optimal growth temperatures (28{degrees}C and 30{degrees}C) suggesting thermal adaptation in warmer regions. Potassium phosphite EC50 values spanned from 4.61 to 763.13 {micro}g/ml, with significantly higher insensitivity in isolates from California and Florida, reflecting the continued overuse of this fungicide in these major production states. In contrast, baseline sensitivities to ethaboxam, mandipropamid, mefenoxam, fluopicolide, and oxathiapiprolin were uniformly high, with narrow, unimodal EC50 distributions across states. Finally, a wide range of virulence among isolates was detected using avocado seedlings and DAnjou pear fruits with isolates from California and Puerto Rico being the most virulent. Together, this data documents extensive phenotypic diversity within clonal A2 P. cinnamomi populations including heat-adapted and phosphite-insensitive lineages, establishes multi-state fungicide sensitivity baselines, and underscores the need for continued surveillance, integrated fungicide stewardship (especially phosphonates), and rootstock screening against phenotypically diverse populations to sustain avocado PRR management and ensure the United States avocado industry sustainability and profitability.

BACKGROUNDEndophytic fungi are naturally inhabiting plant organs without causing disease symptoms. They can also contribute to their hosts pest and disease resistance by displaying entomopathogenic and/or antifungal traits. In this study, we evaluated the ability of 11 strains of avocado fungal endophytes to antagonize three important avocado plant pathogens: Colletotrichum gloeosporioides, Fusarium solani, and Phytophthora cinnamomi, and two insect pests: Sitophilus zeamais and Xyleborus bispinatus. RESULTSThe results show that Trichoderma spp. strains were the most effective against the evaluated plant pathogens in terms of growth inhibition, in direct contact assays or through metabolite production. Other fungi, such as Purpureocillium sp. and Pochonia sp., only exhibited pathogen inhibition through diffusible metabolites but displayed strong insecticidal capacity against the evaluated pests, hence being identified as promising multi-target biocontrol agents in the integrative analysis. CONCLUSIONOur findings evidence the potential of avocado fungal endophytes and their metabolites as multi-target biocontrol agents of crop pests and pathogens.

Chia (Salvia hispanica L.) is an emerging crop in Bangladesh valued for its medicinal properties and economic significance. In March 2024, target spot-like symptoms were observed in an experimental chia field (24.75{degrees} N, 90.50{degrees} E) at Bangladesh Agricultural University in Mymensingh, Bangladesh with disease incidence ranging from 23% to 47% across approximately 0.25 ha. Initially appearing as brick-red spots, these symptoms developed into target-shaped concentric rings, affecting leaves, stems, and inflorescences. A total of 24 fungal isolates were recovered from infected tissue; two representative isolates (BGECh-3 and BGECh-4) were randomly selected for details characterization. Pathogen identity was established through morphological traits, multilocus phylogenetic analysis of internal transcribed spacer (ITS) and elongation factor 1-alpha (EF-1) genes sequence, and pathogenicity confirmation through Kochs postulates, collectively identifying the causal agent as Corynespora cassiicola. The isolates demonstrated a broad host range, successfully infecting brinjal, chili, bottle gourd, country bean, tomato, and soybean. In vitro fungicide sensitivity assays with seven commercial fungicides showed that both isolates were highly sensitive to Goldzim (50% carbendazim), which completely inhibited mycelial growth at 10 {micro}g mL-{superscript 1}. Conza (10% Hexaconazole) and Amister top (18.2% azoxystrobin + 11.4% difenoconazole) reduced growth by up to 85% and 67%, respectively at equal concentration. Other fungicides showed comparatively lower efficacy even at higher concentrations. This study represents the first report of target spot disease of chia caused by C. cassiicola in Bangladesh and provides insights for effective disease management strategies.

Ectomycorrhizal (ECM) fungi are well-known for their crucial roles in forest health and productivity, yet their responses to various forest management practices are understudied, particularly in oak-dominated forests. The purpose of this study was to better understand the effects of silvicultural treatments on the diversity and community composition of ECM fungi in an oak-hornbeam forest in northern Hungary. We analyzed ITS2 rDNA metabarcoding data of soil-borne fungi to compare richness and community composition of ECM fungi among forest treatment types (clear-cutting, gap-cutting, preparation-cutting, tree retention in clear-cut areas, and control) and between sampling years (2020 and 2021). We found 268 ECM fungal genotypes, with the most diverse phylogenetic clades being /russula-lactarius (52), /tomentella-thelephora (47), /inocybe (40), /sebacina (27), and /cortinarius (20). We found significant compositional difference of ECM fungi among silvicultural treatments in both years, with some variations in richness. There were also small, but still significant compositional differences between the two years. Treatment effect was partly explained by altered environmental variables, such as relative humidity and soil temperature. These results highlight the importance of forest structure and the abiotic environment in driving community dynamics of plant-symbiotic fungi, with potential implications for forest health and productivity.

BackgroundPotato is an important crop worldwide, yet its production is severely threatened by Phytophthora infestans, the causal agent of late blight. Alternatives to the current control strategies are needed, as these rely heavily on environmentally harmful treatments. The recruitment of beneficial microbes by plants upon stress ("cry-for-help" mechanism) may represent an opportunity to find new biocontrol agents but this has not yet been reported for potato. The aim of this study was to analyse whether foliar late blight infection induces shifts in the phyllosphere, rhizosphere and soil bacterial communities associated with two potato cultivars of differing sensitivity to late blight. Moreover, we aimed at isolating members of the plant microbiota to test whether bacteria putatively recruited upon infection would be particularly active in protecting the plant against late blight. ResultsControlled foliar infection triggered substantial, cultivar-specific shifts in the rhizosphere communities across two successive generations. Despite the number of differentially abundant ASVs detected being ten times higher in the second generation than in the first one, the same taxonomic groups were concerned by the shifts: Burkholderiales, Flavobacteriales, and Bacillales. Furthermore, the communities linked to the susceptible cultivar consistently shifted more strongly than the communities linked to the resistant cultivar. The obtained ASV sequences were used to identify 163 corresponding isolates. The inhibition potential of these strains against P. infestans spores was assessed through biological assays, which revealed the biocontrol potential of strains otherwise not yet known to inhibit phytopathogenic organisms, such as Advenella, Nocardioides and Phyllobacterium strains. Although we found no correlation between the relative abundance shift of the ASVs upon infection and the activity of the corresponding strains, we observed that the overall activity of strains isolated from the resistant cultivar was higher than that of the strains isolated from the susceptible one. ConclusionTaken together, the higher activity of the strains isolated from the resistant cultivar, along with its comparatively modest microbiome shifts upon infection suggest that the investigated resistant cultivar might harbour specific microbiota enriched in strains with efficient protective abilities against their host plants pathogens, which possibly contribute to its higher resistance against P. infestans.

Longitudinal data on disease susceptibility in forest trees are rare but essential for understanding host-pathogen dynamics and genetic variation in susceptibility traits. We present a long-term multisite common garden dataset quantifying susceptibility of Scots pine (Pinus sylvestris) to Dothistroma needle blight. The dataset comprises annual disease assessments collected from the same trees across 11 years, spanning 168 families and 21 Scottish provenances. This design enables partitioning of genetic and environmental sources of variation, evaluation of temporal stability in host response, and estimation of variance components and narrow-sense heritability of susceptibility. The data support analyses of phenotypic plasticity, provenance-level responses, and interactions between disease susceptibility and other adaptive traits. This resource will facilitate predictive modelling of host susceptibility under current and future environmental conditions.

RNA interference (RNAi) shows great potential to protect crops against fungal diseases, yet reported protection efficiencies vary greatly, and our understanding of the factors responsible for this variance remains limited. In this meta-analysis, we evaluated 89 studies that compare the efficiency of host-induced gene silencing (HIGS) and spray-induced gene silencing (SIGS) in controlling fungal diseases, focusing on biotrophic, hemibiotrophic, and necrotrophic fungi, the use of formulations, and the dsRNA design as explanatory factors for differences between reported efficiency values. Our results indicate that SIGS is slightly more effective, particularly in biotrophs. Surprisingly, SIGS studies using formulations did not outperform those applying naked dsRNA. We also assessed parameters of RNA design. Differences in dsRNA length and the number of constructs, and number of targets showed no consistent significant effect on resistance in either HIGS or SIGS. Interestingly, however, HIGS studies reported significantly higher efficiency when targeting genes closer to the 3 end and SIGS when targeting genes closer to the 5 end. We discuss potential reasons for the reported patterns, such as variability in dsRNA uptake mechanisms, intercellular trafficking and Dicer processing, and conclude that more research is needed to understand the biological mechanisms determining RNAi efficiency for fungal control.