Minna Haapalainen

Chocolate spot (CS) disease, caused by Botrytis species, severely damages faba beans. This study analyzed interactions between three faba bean genotypes with varying levels of resistance and 21 Botrytis isolates from six species that were genotyped using 10 NEP markers. The isolates were cultured, sporulated under near-UV light, and inoculated onto 6–8-week-old, fully expanded, detached leaflets. All isolates induced CS symptoms, revealing significant interactions among lines and isolates. Botrytis fabae 19B053-4 was the most virulent, followed by B. fabiopsis 19B175 and B. fabae 17B4. Conversely, B. cinerea 17B10-1 caused the slowest lesion development. Notably, more resistant hosts consistently remained less susceptible across all isolates, indicating no crossover interaction. AUDPC values confirmed B. fabae as the primary, highly aggressive specialized pathogen, whereas related species pose opportunistic threats. Tracking cumulative disease progress identified quantitative host resistance through slowed infection rates, offering a framework for crop protection.

Fungal pathogens often cause severe post-harvest losses in stored carrot yields. In 2020 and 2021, carrot samples were collected in Finland from 26 fields with different soil types. The proportion of symptomatic carrots was assessed at two time points during the cold storage period. To analyse the occurrence of different fungal pathogens in the symptomatic carrots, DNA samples were prepared and tested for the presence of fungal pathogens by species-specific PCR. The most frequently detected pathogens were Cylindrocarpon spp. with 61% occurrence, followed by Fusarium avenaceum (44%), Mycocentrospora acerina (42%), Botrytis cinerea (12%), and Alternaria spp. (0.8%). B. cinerea and F. avenaceum were more common in carrots grown on organic soil, while M. acerina was more common in carrots grown on mineral soils. Mixed fungal infections were overrepresented in the symptomatic samples, in particular, Cylindrocarpon spp. often occurred together with F. avenaceum and M. acerina in the same samples. Amplicon sequencing of the fungal internal transcribed spacer region from the DNA of symptomatic tissues revealed that in addition to the pathogens— B. cinerea , M. acerina, Cylindrocarpon spp. and Sclerotinia sclerotiorum —the hyphomycete fungi Tetracladium spp. and the yeast Leucosporidium intermedium were abundant, and that Tetracladium spp. were more abundant in the carrots infected with M. acerina . A pathogenicity test confirmed that F. avenaceum is pathogenic to both the new season and old season carrots.

Fungal pathogens often cause severe post-harvest losses in stored carrot yields. In 2020 and 2021, carrot samples were collected in Finland from 26 fields with different soil types. The proportion of symptomatic carrots was assessed at two time points during the cold storage period. To analyse the occurrence of different fungal pathogens in the symptomatic carrots, DNA samples were prepared and tested for the presence of fungal pathogens by species-specific PCR. The most frequently detected pathogens were Cylindrocarpon spp. with 61% occurrence, followed by Fusarium avenaceum (44%), Mycocentrospora acerina (42%), Botrytis cinerea (12%), and Alternaria spp. (0.8%). B. cinerea and F. avenaceum were more common in carrots grown on organic soil type, while M. acerina was more common in carrots grown on mineral soils. Mixed fungal infections were overrepresented in the symptomatic samples, in particular, Cylindrocarpon spp. often occurred together with F. avenaceum and M. acerina in the same samples. Amplicon sequencing of the fungal internal transcribed spacer region from the DNA of symptomatic tissues revealed that in addition to the pathogens - B. cinerea , M. acerina , Cylindrocarpon spp. and Sclerotinia sclerotiorum - the hyphomycete fungi Tetracladium spp. and the yeast Leucosporidium intermedium were abundant, and that Tetracladium spp. were more abundant in the carrots infected with M. acerina . A pathogenicity test confirmed that F. avenaceum is pathogenic to both the new season and old season carrots.

Postharvest diseases can cause significant losses in carrot ( Daucus carota ) yield during storage. In this study, the effects of soil quality and microbial diversity in the field bulk soil and rhizosphere on the development of postharvest diseases were investigated. Microbial genera in bulk soil and rhizosphere samples were identified by Illumina Miseq amplicon sequencing of bacterial 16S and fungal ITS regions. Disease symptoms in the stored carrots were monitored after six months of cold storage, and the main pathogens were identified. The field sites with mineral soils, which had a higher pH, had a higher bacterial and fungal diversity than organic soils. The organically managed fields had a higher bacterial diversity than the conventionally managed fields. Community structure of both bacteria and fungi was largely driven by field site including its main chemical characteristics, with rhizosphere and bulk soil samples from the same site displaying comparable assemblages. Higher soil microbial diversity was associated with higher postharvest storability in the stored carrots. The weather during the growing season also had an effect on the observed fungal biodiversity, which was higher at sites that had lower temperatures and less rain prior to harvest.

Fusarium proliferatum is one of the main pathogens causing Fusarium basal rot of onion, which is a major problem in onion cultivation worldwide. In this work, three Fusarium proliferatum isolates from onion (Allium cepa), two from imported sets and one from a mature bulb grown in Finland, were tested for pathogenicity and mycotoxin production. Symptom development in the inoculated onion bulbs and production of fumonisins, beauvericin, and moniliformin were measured at 1–5 weeks post-inoculation (wpi). Symptoms were observed in all the bulbs inoculated with F. proliferatum, starting as water-soaked lesions and developing into dead brown scales. When colonizing the bulbs, all three F. proliferatum isolates were confirmed by qPCR to express the genes FUM1, required for fumonisin production, and SIX2, encoding a putative virulence factor. Fumonisin B₁ (FB₁) was detected already at 2 wpi in a symptomatic onion inoculated with Fpr047, and at 3 wpi, fumonisin production was confirmed for all three isolates. At 4 wpi, all the isolates had produced quantifiable amounts of FB₁, ranging from 3.9 to 177 µg/kg, and fumonisin B₂, ranging from 3.4 to 89 µg/kg. At 5 wpi, FB₁ was even detected in a symptomless sample. Beauvericin and moniliformin were quantified at 5 wpi in the symptomatic onion tissues with Fpr047 and Fpr049, but not with FUS16163, which was unable to produce moniliformin and produced less beauvericin than the other F. proliferatum isolates also in the rice culture. The results showed that onions infected with F. proliferatum can contain high amounts of fumonisins.

Carrot psyllid Trioza apicalis is a serious pest of cultivated carrot and also a vector of the plant pathogen ‘Candidatus Liberibacter solanacearum’ (Lso). To find out whether T. apicalis harbours other species of bacteria that might affect the Lso infection rate, the bacterial communities and metagenome in T. apicalis were studied. Lso haplotype C was detected in a third of the psyllids sampled, at different relative amounts. Surprisingly, T. apicalis was found to harbour only one secondary endosymbiont, a previously unknown species of gamma proteobacterium endosymbiont (Gpe), beside the primary endosymbiont ‘Candidatus Carsonella ruddii’ (CCr). The relative abundancies of these two endosymbionts were approximately equal. The genomes of CCr, Gpe and Lso were assembled from a T. apicalis metagenome sample. Based on the 16S rRNA gene, the closest relative of Gpe of T. apicalis could be a secondary endosymbiont of Trioza magnoliae. The 253.171 kb Gpe genome contains all the tRNA and rRNA genes and most of the protein-coding genes required for DNA replication, transcription and translation, but it lacks most of the genes for amino acid biosynthesis. Gpe has no genes encoding cell wall peptidoglycan synthesis, suggesting it has no cell wall, and could thus live as an intracellular endosymbiont. Like the CCr of other psyllids, CCr of T. apicalis retains a broad amino acid biosynthetic capacity, whilst lacking many genes required for DNA replication and repair and for transcription and translation. These findings suggest that these two endosymbionts of T. apicalis are complementary in their biosynthetic capabilities.

Post‐harvest diseases cause significant economic losses in the carrot production chain. In this study, storage losses and fungal pathogens causing them were analysed in the carrot yield from 52 different field plots in four areas in Finland in 3 years (2016–2018). Over 30,000 carrots were sampled and analysed at three time points during cold storage at 0–1°C. In March, after 5–6 months' storage, the average loss due to diseases was 20%–21% every year. Decay of the root tip was the most common disease symptom, followed by pits on the side and black rot in the crown, detected in 69.2%, 15.0% and 9.0% of the symptomatic samples, respectively. Both intensive carrot cultivation practice and early timing of harvest increased storage losses. Pathogens in 3057 symptomatic carrot tissue samples were isolated by culturing, and fungal species were identified. The most common fungal species detected were Mycocentrospora acerina, Botrytis cinerea and Fusarium spp., especially F. avenaceum. However, the frequency of different pathogens varied between the different years and time points during storage. Species‐specific PCR tests revealed that M. acerina and F. avenaceum were present in many early time‐point samples where they could not yet be detected by the culturing method. In Finland, this study on carrot post‐harvest diseases is the first large‐scale survey in which the fungal pathogens were isolated and identified by laboratory tests. In comparison with the previous studies, Fusarium spp. were detected more frequently in this study, while grey mould and Sclerotinia rot were less frequent.

The interactions between the phloem-limited pathogen ‘Candidatus Liberibacter solanacearum’ haplotype C and carrot (Daucus carota subsp. sativus) were studied at 4, 5, and 9 weeks postinoculation (wpi), by combining dual RNA-Seq results with data on bacterial colonization and observations of the plant phenotype. In the infected plants, genes involved in jasmonate biosynthesis, salicylate signaling, pathogen-associated molecular pattern- and effector-triggered immunity, and production of pathogenesis-related proteins were up-regulated. At 4 wpi, terpenoid synthesis-related genes were up-regulated, presumably as a response to the psyllid feeding, whereas at 5 and 9 wpi, genes involved in both the terpenoid and flavonoid production were down-regulated and phenylpropanoid genes were up-regulated. Chloroplast-related gene expression was down-regulated, in concordance with the observed yellowing of the infected plant leaves. Both the RNA-Seq data and electron microscopy suggested callose accumulation in the infected phloem vessels, likely to impair the transport of photosynthates, while phloem regeneration was suggested by the formation of new sieve cells and the upregulation of cell wall–related gene expression. The ‘Ca. L. solanacearum’ genes involved in replication, transcription, and translation were expressed at high levels at 4 and 5 wpi, whereas, at 9 wpi, the Flp pilus genes were highly expressed, suggesting adherence and reduced mobility of the bacteria. The ‘Ca. L. solanacearum’ genes encoding ATP and C4-dicarboxylate uptake were differentially expressed between the early and late infection stages, suggesting a change in the dependence on different host-derived energy sources. HPE1 effector and salicylate hydroxylase were expressed, presumably to suppress host cell death and salicylic acid-dependent defenses during the infection. [ ] Copyright © 2021 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license .

Fusarium oxysporum forma specialis cepae strains are pathogens causing Fusarium basal rot of onion, which is a major problem in onion cultivation worldwide. In this work, the pathogenicity and toxin production of different F. oxysporum isolates, originating either from mature harvested onions or from onion sets, were studied. The F. oxysporum isolates belonging to the f. sp. cepae phylogenetic clade had the full set of seven secreted in xylem (SIX) genes and the gene C5, whereas all the other F. oxysporum isolates tested were negative for these virulence‐related genes. However, in the pathogenicity test on onion seedlings the separation between the f. sp. cepae isolates and the others was not clear‐cut. One of the f. sp. cepae isolates that had low in planta expression levels of SIX9 showed only mild virulence on the seedlings. A different F. oxysporum isolate, which did not have the known virulence‐related genes, was pathogenic on seedlings, although non‐pathogenic on mature bulbs. Thus, the two different pathogenicity assays frequently used, seedling survival test and onion bulb inoculation test, measure partially different features of the fungal isolates. Coinfection with pathogenic and non‐pathogenic isolates caused more severe disease in the mature onion bulbs than the pathogenic isolate alone. All the isolates tested for toxin production capacity on rice medium produced beauvericin and, in addition, the non‐pathogenic isolates produced moniliformin. Small amounts of beauvericin were detected in the infected onion bulbs. The amount of beauvericin did not show significant correlation with the fungal colonisation level or symptom severity.

Fusarium basal rot (FBR) of onion is a serious disease problem worldwide. The Fusarium species causing FBR can also produce mycotoxins that are potentially harmful to humans and animals. In this study, a multiple reaction monitoring technique with ultra-high-performance liquid chromatography–tandem mass spectrometry (MRM UHPLC-MS/MS) was developed and validated for onion matrix to study Fusarium mycotoxins in the harvested onions. This study was focused on fumonisins B₁, B₂, and B₃ (FB₁, FB₂, and FB₃), beauvericin (BEA), and moniliformin (MON), which are the main mycotoxins produced by Fusarium oxysporum and Fusarium proliferatum. In the in-house validated protocol, the onion samples were extracted with methanol:water (3:1) using magnetic stirring for 15 min. FBs and BEA were determined directly from the filtered extracts, whereas MON required sample concentration prior to analysis. No cleanup of extracts was needed prior to analysis. The target mycotoxins were separated on an Acquity UPLC system BEH C18 column with gradient elution. Mycotoxins were identified and quantified using ¹³C-FB₁ as internal standard. Minor matrix effect was compensated using multi-point matrix-matched calibration curves with uninfected onion sample. For the mycotoxins studied, a good linearity was obtained (R² ≥ 0.99) and the recoveries were in the range of 67–122%, with the highest standard deviation for MON, 22%. The limits of quantification were from 2.5 to 10 ng g⁻¹ in onion matrix. The method was successfully employed for the analysis of mycotoxins in harvested onions showing FBR symptoms and found to be infected with F. oxysporum and F. proliferatum.