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Potato is affected by several diseases. Although, resistance can be obtained by introgression of major resistance genes from wild species, this has rarely been durable. Hence, other sources of resistance are highly needed. New research with a focus on loss of function mutations has led to the identification of disease susceptibility (S) genes in plants. The research in this thesis was aimed at the identification and characterization of potato S genes involved in the interaction with Phytophthora infestans and Botrytis cinerea. We selected 11 Arabidopsis thaliana S genes and silenced their potato orthologs by RNAi in the potato cultivar Desiree. The silencing of six genes resulted in resistance to P. infestans. Moreover, silencing of StDND1 reduced the infection of B. cinerea. Microscopic analysis showed that spore attachment and/or germination of P. infestans and B. cinerea was hampered on the leaf surface of StDND1-silenced potato plants. On StDMR1- and StDMR6-silenced potato plants, hyphal growth of P. infestans was arrested by the hypersensitive response-like cell death. Our results demonstrate that impairment of plant S genes may open a new way for breeding potatoes with resistance to pathogens like P. infestans and B. cinerea.

Powdery mildew (PM) is a major fungal disease that threatens thousands of plant species. PM is caused by Podosphaera leucotricha in apple and Erysiphe necator in grapevine. Powdery mildew is controlled by frequent applications of fungicides, having negative effects on the environment, and leading to additional costs for growers. To reduce the amount of chemicals required to control this pathogen, the development of resistant apple and grapevine varieties should become a priority. PM pathogenesis is associated with up-regulation of specific MLO genes during early stages of infection, causing down-regulation of plant defense pathways. These up-regulated genes are responsible for PM susceptibility (S-genes) and their knock-out causes durable and broad-spectrum resistance. All MLO S-genes of dicots belong to the phylogenetic clade V. In grapevine, four genes belong to clade V. VvMLO7, 11 and 13 are up-regulated during PM infection, while VvMLO6 is not. Chapter 2 reports the genome-wide characterization and sequence analysis of the MLO gene family in apple, peach and woodland strawberry, and the isolation of apricot MLO homologs. Twenty-one homologues were found in apple, 19 in peach and 17 in woodland strawberry. Evolutionary relationships between MLO homologs were studied and syntenic blocks constructed. Candidate genes for causing PM susceptibility were inferred by phylogenetic relationships with functionally characterized MLO genes and, in apple, by monitoring their expression following inoculation with the PM causal pathogen P. leucotricha. In apple, clade V genes MdMLO11 and 19 were up-regulated, whereas the two other members of clade V, MdMLO5 and 7, were not up-regulated. The clade VII gene MdMLO18 was also up-regulated upon P. leucotricha infection. Chapter 3 reports the knock-down, through RNA interference, of MdMLO11 and 19, as well as complementation of the mutant phenotype by expression of the MdMLO18 gene in the Arabidopsis thaliana triple mlo mutant Atmlo2/6/12. The knock-down of MdMLO19 resulted in a reduction of PM disease severity up to 75%, whereas the knock-down of MdMLO11, alone or combined with MdMLO19, did not cause any reduction or additional reduction of susceptibility compared to MdMLO19 alone. Complementation by MdMLO18 did not restore susceptibility. Cell wall appositions (papillae), a response to PM infection, were found in both susceptible plants and PM resistant plants where MdMLO19 was knocked-down, but were larger in resistant lines. The expression analysis of 17 genes related to plant defense, and quantification of phenolic metabolites in resistant lines revealed line-specific changes compared to the control. Chapter 4 evaluates the presence of non-functional alleles of the MdMLO19 S-gene in apple germplasm. The screening of the re-sequencing data of 63 apple genotypes led to the identification of 627 SNP in five MLO genes (MdMLO5, MdMLO7, MdMLO11, MdMLO18 and MdMLO19). Insertion T-1201 in MdMLO19 caused the formation of an early stop codon, resulting in a truncated protein lacking 185 amino-acids and the calmodulin-binding domain. The presence of the insertion was evaluated in a collection of 159 apple genotypes: it was homozygous in 53 genotypes, 45 of which were resistant or very resistant to PM, four partially susceptible and four not assessed. These results strongly suggest that this insertion is causative for the observed PM resistance. The absence of a clear fitness cost associated to the loss-of-function of MdMLO19, might have contributed to the high frequency of the mutation in breeding germplasm and cultivars. Among the genotypes containing the homozygous insertion, ‘McIntosh’ and ‘Fuji’ are commonly used in apple breeding. After barley and tomato, apple is the third species with a reported natural non-functional mlo allele in its germplasm, with the important difference that the allele is present in a relatively large number of apple genotypes, most of which not related to each other. Chapter 5 reports the knock-down through RNA interference of four grapevine MLO genes, all members of clade V. VvMLO7, 11 and 13 are up-regulated in early stages of infection, whereas VvMLO6 is not responsive to the pathogen. Knock-down of VvMLO6, 11 and 13, alone or combined, did not decrease PM severity, whereas the knock-down of VvMLO7, alone or in combination with VvMLO6 and VvMLO11, caused a reduction of severity of 77%. Cell wall appositions (papillae), a response to PM attack, were present in both resistant and susceptible lines, but were larger in resistant lines. Thirteen genes involved in defense were less up-regulated in resistant plants, highlighting the reduction of PM disease severity. In Chapter 6 we discuss the results presented in this thesis. The pivotal role of MLO genes in the interaction of PM pathogens with apple and grapevine is described and further experiments aimed at addressing open questions are proposed. The results described in this thesis open interesting avenues in MLO genes research, particularly the finding that a natural mlo mutation in apple appeared to be more common than expected. This mutation is directly applicable in marker assisted breeding for durable PM resistance in apple.

Powdery mildew (PM) is a worldwide-occurring plant disease caused by ascomycete fungi of the order Erysiphales. A conspicuous number of plant species are susceptible to this disease, the occurrence of which is increasing due to the influence of climate change. Symptoms are easy to recognize by the powdery whitish fungal structures growing on the surface of plant organs. Severe infections cause significant losses in crops, such as tomato, cucumber and wheat, as well as in ornamentals, like rose and petunia. Accordingly, breeding crops with a robust immunity to this disease is of great economic importance. A significant step in this direction was the discovery of mlo (mildew locus o) mutant alleles of the barley HvMlo gene, which are responsible for the non-race specific resistance to the barley PM pathogen, Blumeria graminis f.sp. hordei (Bgh). During the years, this recessively inherited resistance was observed to be durable, contrary to the short life-span of resistances conferred by dominant resistance (R-) genes used in barley breeding programs. Studies on the histological mechanisms of the mlo-based resistance showed that the PM pathogen was stopped during penetration of the cell wall by the formation of a papilla. This structure prevents the formation of the feeding structure of the pathogen, called a haustorium. After sequencing many plant genomes, we are discovering that MLO genes are not only typical of this cereal, but are ubiquitously present in higher plant species in multiple copies per species, forming a gene family. The impairment of some members of a number of ever increasing plant species lead to broad-spectrum resistance towards their adapted PM pathogens. For example, in tomato the ol-2 gene, naturally harbored by the cherry tomato Solanum lycopersicum var. cerasiforme, represents the loss-of-function allele of the SlMLO1 gene, conferring resistance to the PM pathogen Oidium neolycopersici (On). Consequently, the use of mlo mutants represents a suitable alternative to the classical use of R-genes in breeding programs. In Chapter 2 , we describe the in silico identification of the complete tomato SlMLO gene family using the available information in the SOL genomic network database. In total, 16 tomato SlMLO members were cloned from leaf, root, flower and fruit of the susceptible tomato cv. Moneymaker to confirm the sequences retrieved from the database and to verify their actual expression in these tissues. We observed the presence of various types of splicing variants, although their possible functional meaning has not been investigated. Motif analyses of each of the translated protein sequences and phylogenetic studies highlighted, on one hand, amino acid stretches that characterize the whole MLO family, and, on the other hand, stretches conserved in MLO homologs that are phylogenetically related. Following a gene expression study upon On inoculation, we identified members of the SlMLO family that are upregulated few hours after pathogen challenge. Except SlMLO1, none of the three newly identified homologs in clade V, thus phylogenetically close to SlMLO1, are induced. Interestingly, two homologs, each found in different clades, are upregulated similarly to SlMLO1. Using an RNAi approach, we silenced the additional clade V-SlMLO homologs, namely SlMLO3, SlMLO5 and SlMLO8, to investigate their possible role in PM resistance. We observed that none of these homologs if individually silenced, leads to PM resistance. However, if SlMLO5 and SlMLO8 are silenced together with SlMLO1, a significantly higher level of resistance is achieved compared to plants carrying the ol-2 allele. The role of SlMLO3 could not be verified. We, therefore, concluded that there are three SlMLO genes in tomato unevenly contributing to the PM disease, of which SlMLO1 has a major role. Chapter 3 focuses on the components of the tomato mlo-based resistance. In Arabidopsis, it is known that four members of the SNARE protein family, involved in membrane fusion, are involved in mlo-based resistance. In this chapter, we focused on the identification of tomato homologs of the Arabidopsis syntaxin PEN1 (AtSYP121). Among the group of syntaxins identified in tomato, two were closely related to each other and also to AtPEN1, denominated SlPEN1a and SlPEN1b. Another Arabidopsis syntaxin that shows a high level of homology with PEN1, called SYP122, was also found to group together with the newly identified SlPEN1 genes. However, the role of SYP122 in plant immunity was not shown in literature. After obtaining individual silencing RNAi constructs, we transformed the resistant ol-2 line, and we challenged the obtained transformants with the adapted PM On, and the non-adapted Bgh. Interestingly, we observed a significant On growth and an enhanced Bgh cell entry only in SlPEN1a silenced plants but not in SlPEN1b silenced ones. We performed a protein alignment of tomato and Arabidopsis functional and non-functional PEN sequences. The presence of three differently conserved non-synonymous amino-acid substitutions is hypothesised to be responsible for the specialization in plant immune function. In Chapter 4 and Chapter 5, we build up a body of evidence pointing to the fact that the function of the MLO susceptibility genes is highly conserved between monocot and dicot plant species. In Chapter 4 we started by identifying and functionally characterizing two new MLO genes of Solanaceous crops affected by the PM disease, tobacco (Nicotiana tabacum) and eggplant (Solanum melongena). We named them NtMLO1 and SmMLO1 in the respective species, as they are the closest homologs to tomato SlMLO1. By overexpressing these genes in the resistant ol-2 line, we obtained transgenic plants that were susceptible to the PM pathogen On. This finding demonstrates that both heterologous MLO proteins can rescue the function of the impaired ol-2 allele in tomato. In addition, we found in tobacco NtMLO1 an amino acid (Q198) of critical importance for the susceptibility function of this protein. In Chapter 5 , we used the same approach adopted in Chapter 4 to show that other MLO proteins of more distant dicot species, like pea PsMLO1, can rescue the loss-of-function of the tomato ol-2 allele. And finally, we stretched this concept also to monocot MLO proteins, using barley HvMlo. While performing these experiments, we could verify that the function of the monocot and dicot susceptibility MLO proteins does not rely on the presence of class-specific conservation. The latter can be the reason for the phylogenetic divergence, placing monocot MLO proteins in clade IV and dicot MLO proteins in clade V of the phylogenetic MLO tree. However, functional conservation might depend on crucial shared amino acids of clade IV and V MLO proteins. Therefore, we also conducted a codon-based evolutionary analysis that resulted in the identification of 130 codons under negative selection, thus strongly maintained during evolution. In Chapter 6 we introduce the PM disease in cucumber caused by Podosphaera xanthii (Px). We cloned the candidate susceptibility gene for PM in cucumber, CsaMLO8, from susceptible and resistant genotypes. The latter was described as an advanced cucumber breeding line characterized by hypocotyl resistance. In this line, we found the presence of aberrant splicing variants of the CsaMLO8 mRNA due to the insertion in its corresponding genomic region of a Class LTR retrotransposon. Heterologous expression of the wild-type cucumber allele in the tomato ol-2 line restored its PM susceptibility, while the heterologous expression of the aberrant protein variant failed to do so. This finding confirms that the resistance of the advanced cucumber breeding line is due to the disruption of the coding region of this gene. We also showed that the expression of CsaMLO8 in the susceptible genotype is induced by Px in hypocotyl tissue, but not in cotyledon or leaf. Finally, by examination of the resequencing data of a collection of 115 cucumber accessions, we found the presence of the TE-containing allele in 31 of them among which a wild cucumber accession that might have been used in breeding programs to obtain resistance to the PM disease in cucumber. In Chapter 7 a novel loss-of-function allele of the SlMLO1 gene is described, designated m200. This allele was found in a resistant plant (M200) from a mutagenized tomato Micro-Tom (MT) population obtained with the chemical mutagen ethyl methanesulfonate (EMS). The m200 mutation corresponds to a nucleotide transversion (T a A) which results in a premature stop codon. The length of the predicted SlMLO1 protein in the M200 plant is only 21 amino acids, thus much shorter than the predicted protein of the previously described ol-2 allele, consisting of 200 amino acids. Thanks to the development of a High-Resolution Melting (HRM) marker designed to detect the m200 mutation, we observed that this allele confers recessively inherited resistance in backcross populations of the resistant M200 plant with MT and Moneymaker. Histological study showed that the resistance of the m200 mutant is associated with papilla formation. Finally, we compared the rate of On penetration in epidermal cells of m200 plants with the one of plants carrying the ol-2 allele and the transgenic plants in which multiple SlMLO homologs were silenced, generated in Chapter 2. Ultimately, in Chapter 8 the results of the previous chapters are discussed in the context of 1) practical applications in breeding programs aimed at introducing the mlo-based resistance in new crops, 2) possible research aimed at unraveling the function of the MLO protein and 3) the role of other SNARE proteins.

Powdery mildew (PM) is a major fungal disease that threatens thousands of plant species. PM is caused by Podosphaera leucotricha in apple and Erysiphe necator in grapevine. Powdery mildew is controlled by frequent applications of fungicides, having negative effects on the environment, and leading to additional costs for growers. To reduce the amount of chemicals required to control this pathogen, the development of resistant apple and grapevine varieties should become a priority. PM pathogenesis is associated with up-regulation of specific MLO genes during early stages of infection, causing down-regulation of plant defense pathways. These up-regulated genes are responsible for PM susceptibility (S-genes) and their knock-out causes durable and broad-spectrum resistance. All MLO S-genes of dicots belong to the phylogenetic clade V. In grapevine, four genes belong to clade V. VvMLO7, 11 and 13 are up-regulated during PM infection, while VvMLO6 is not. Chapter 2 reports the genome-wide characterization and sequence analysis of the MLO gene family in apple, peach and woodland strawberry, and the isolation of apricot MLO homologs. Twenty-one homologues were found in apple, 19 in peach and 17 in woodland strawberry. Evolutionary relationships between MLO homologs were studied and syntenic blocks constructed. Candidate genes for causing PM susceptibility were inferred by phylogenetic relationships with functionally characterized MLO genes and, in apple, by monitoring their expression following inoculation with the PM causal pathogen P. leucotricha. In apple, clade V genes MdMLO11 and 19 were up-regulated, whereas the two other members of clade V, MdMLO5 and 7, were not up-regulated. The clade VII gene MdMLO18 was also up-regulated upon P. leucotricha infection. Chapter 3 reports the knock-down, through RNA interference, of MdMLO11 and 19, as well as complementation of the mutant phenotype by expression of the MdMLO18 gene in the Arabidopsis thaliana triple mlo mutant Atmlo2/6/12. The knock-down of MdMLO19 resulted in a reduction of PM disease severity up to 75%, whereas the knock-down of MdMLO11, alone or combined with MdMLO19, did not cause any reduction or additional reduction of susceptibility compared to MdMLO19 alone. Complementation by MdMLO18 did not restore susceptibility. Cell wall appositions (papillae), a response to PM infection, were found in both susceptible plants and PM resistant plants where MdMLO19 was knocked-down, but were larger in resistant lines. The expression analysis of 17 genes related to plant defense, and quantification of phenolic metabolites in resistant lines revealed line-specific changes compared to the control. Chapter 4 evaluates the presence of non-functional alleles of the MdMLO19 S-gene in apple germplasm. The screening of the re-sequencing data of 63 apple genotypes led to the identification of 627 SNP in five MLO genes (MdMLO5, MdMLO7, MdMLO11, MdMLO18 and MdMLO19). Insertion T-1201 in MdMLO19 caused the formation of an early stop codon, resulting in a truncated protein lacking 185 amino-acids and the calmodulin-binding domain. The presence of the insertion was evaluated in a collection of 159 apple genotypes: it was homozygous in 53 genotypes, 45 of which were resistant or very resistant to PM, four partially susceptible and four not assessed. These results strongly suggest that this insertion is causative for the observed PM resistance. The absence of a clear fitness cost associated to the loss-of-function of MdMLO19, might have contributed to the high frequency of the mutation in breeding germplasm and cultivars. Among the genotypes containing the homozygous insertion, ‘McIntosh’ and ‘Fuji’ are commonly used in apple breeding. After barley and tomato, apple is the third species with a reported natural non-functional mlo allele in its germplasm, with the important difference that the allele is present in a relatively large number of apple genotypes, most of which not related to each other. Chapter 5 reports the knock-down through RNA interference of four grapevine MLO genes, all members of clade V. VvMLO7, 11 and 13 are up-regulated in early stages of infection, whereas VvMLO6 is not responsive to the pathogen. Knock-down of VvMLO6, 11 and 13, alone or combined, did not decrease PM severity, whereas the knock-down of VvMLO7, alone or in combination with VvMLO6 and VvMLO11, caused a reduction of severity of 77%. Cell wall appositions (papillae), a response to PM attack, were present in both resistant and susceptible lines, but were larger in resistant lines. Thirteen genes involved in defense were less up-regulated in resistant plants, highlighting the reduction of PM disease severity. In Chapter 6 we discuss the results presented in this thesis. The pivotal role of MLO genes in the interaction of PM pathogens with apple and grapevine is described and further experiments aimed at addressing open questions are proposed. The results described in this thesis open interesting avenues in MLO genes research, particularly the finding that a natural mlo mutation in apple appeared to be more common than expected. This mutation is directly applicable in marker assisted breeding for durable PM resistance in apple.

Mahlet Teka Anche. (2016). Estimating host genetic effects on susceptibility and infectivity to infectious diseases and their contribution to response to selection. PhD thesis, Wageningen University, the Netherlands Genetic approaches aiming to reduce the prevalence of an infection in a population usually focus on improving host susceptibility to an infection. The prevalence of an infection, however, is also affected by the infectivity of individuals. Studies reported that there exists among host (genetic/phenotypic) variation in susceptibility and infectivity to infectious diseases. The effect of host genetic variation in susceptibility and infectivity on the prevalence and risk of an infection is usually measured by the value of the basic reproduction ratio, R0. R0 is an important epidemiological parameter that determines the risk and prevalence of an infection. It has a threshold value of 1, where major disease outbreak can occur when R0 > 1 and the disease will die out when R0 < 1. Due to this threshold property, genetic improvements aiming to reduce the prevalence of an infection should focus on reducing R0 to a value below 1. The overall aim of this thesis was to develop methodologies that allow us to investigate the genetic effects of host susceptibility and infectivity on the prevalence of an infection, which is measured by the value of R0. Moreover, we also aim to investigating the effect of relatedness among groupmates on the utilization of among host genetic variation in susceptibility and infectivity so as to reduce the prevalence of infectious diseases. The theory of direct-indirect genetic effects and epidemiological concepts were combined to develop methodologies. In addition, a simulation study was performed to validate the methodologies developed and examine the effect of relatedness on the utilization of genetic variation in susceptibility and infectivity. It was shown that an individual’s genetic effect on its susceptibility and infectivity affect the prevalence of an infection and that an individual’s breeding value for R0 can be defined as a function of its own allele frequencies for susceptibility and infectivity and of population average susceptibility and infectivity. Moreover, simulation results show that, not only an individual’s infectivity but also an individual’s susceptibility represents an indirect genetic effect on the disease status of individuals and on the prevalence of an infection in a population. It was shown that having related groupmates allows breeders to utilize the genetic variation in susceptibility and infectivity, so as to reduce the prevalence of an infection.

Overzicht van resultaten van onderzoek bij PPO, PRI en IRS naar mogelijke oplossingen voor de bestrijding van de bodemschimmel Rhizoctonia solani in de suikerbietenteelt. Gewassen en onkruiden die als waardplanten fungeren; gevoeligheid van gewassen en het effect op een volggewas suikerbieten (rot; wegval; suikeropbrengst); effect van verschillende voorvruchten op Rhizoctonia-rot en suikeropbrengst; effecten van bladrammenas en gele mosterd in de rotatie (als braakgewas en/of groenbemester); effect van gewasresten (bietenblad) in de grond; resistentiekarakteristieken van Rhizoctonia-resistente rassen (ziekte-index; bietenaantasting)

Uitleg over ABS (Aaltjes Beheersings Strategie). De basis vormt het bouwplan. Vandaar een tabel met de schadegevoeligheid en waardplantgeschiktheid van akker- en tuinbouwgewassen en groenbemesters voor de aaltjes Meloidogyne hapla (noordelijk wortelknobbelaaltje), M. naasi (graswortelknobbelaaltje), M. chitwoodi (maïswortelknobbelaaltje) en M. fallax (maïswortelknobbelaaltje). Als de grond eenmaal besmet is zijn aanvullende maatregelen mogelijk (zwarte braak; anaërobie in de bouwvoor door inundatie of afdekken; natte grondontsmetting eens in de vijf jaar; toepassing van granulaire nematiciden)

PPO onderzocht op drie locaties (zand, klei, veen) de gebruikswaarde van een groot aantal Monarda-cultivars (bergamotplant), waarbij de nadruk lag op de vatbaarheid voor meeldauw. In een tabel het aanbevolen sortiment met gegevens over bloemkleur, bloeitijd en hoogte, een beoordeling van groeikracht, stevigheid en gezondheid, en de waardering door de KVBC (sterrenkeuring voor het gebruik als tuinplant)

De aardappel is waardplant voor de essenverwelkingsziekte (Verticillium dahliae Klehbahn); dit is van belang bij het bebossen van landbouwgronden. Door IBN-DLO is onderzocht hoe plantmateriaal van verschillende herkomsten van Fraxinus excelsior (twee inheems en twee formeel goedgekeurde rassen) reageert op inoculatie met isolaten van de schimmel (twee aardappel- en twee essenisolaten)

Summary Late blight, caused by the oomycete Phytophthora infestans, is one of the most devastating potato diseases worldwide. To successfully colonize its host, P. infestans secretes a plethora of RXLR effectors that translocate into host cells to modulate plant defense. The RXLR effectors form the largest and most diverse effector family in oomycete plant pathogens, and include several that were demonstrated to trigger host resistance mediated by intracellular host immune receptors. Chapter 1 is a summary focussing on the molecular mechanisms underlying host–pathogen interactions. It introduces the multi-layered innate immune system of plants, as well as the strategies that pathogens exploit to circumvent and suppress host defense. Furthermore, it highlights the importance of vesicle-trafficking during plant defense. The central subject of this thesis is AVR1, one of the race-specific avirulence (AVR) factors of P. infestans. AVR1 triggers plant resistance mediated by its corresponding potato Nucleotide-binding Leucine-rich repeat (NLR) resistance protein R1. P. infestans isolates that are avirulent on R1-containing potato cultivars always contain AVR1, while virulent isolates lack AVR1 but contain a related gene that we baptized as AVR1-like. AVR1 has all hallmarks of a typical RXLR effector; it contains a signal peptide, an RXLR domain and a C-terminal effector domain that contains two W motifs and one Y motif. In addition, it has, at the very end a stretch of 38 amino acids in length that we named the Tail (T)-region. AVR1-like, or in short A-L, shares high sequence similarity with AVR1. However, due to a premature stop codon the 38 amino acid T-region is missing. Chapter 2 explores the conserved motifs and regions in the C-terminal effector domain of AVR1 that are required to trigger R1-mediated hypersensitive response (HR). Various truncated and chimeric constructs of AVR1 and A-L were generated and assayed for their ability to elicit R1-mediated HR. Results show that the T-region of AVR1 plays an important role in HR activation. Furthermore, we revealed that R1 recognizes two epitopes in AVR1, one located in the C-terminal region containing the conserved W and Y motifs, and one comprised by the T region. In Chapter 3 the subcellular localization of AVR1 and R1 was investigated. Both were demonstrated to be nucleocytoplasmic proteins. We artificially modified the nucleocytoplasmic partitioning of AVR1 and R1 using nuclear localization and export signals (NLS/NES), and studied the effect on R1-AVR1 recognition. This revealed that nuclear localization of both AVR1 and R1 is important to induce R1-mediated immunity. In addition, we showed that AVR1-mediated suppression of CRN2-induced cell death is dependent on cytosolic localization of AVR1. In Chapter 4, we investigated how AVR1 modulates host defense. In a yeast two-hybrid screening we identified the exocyst subunit Sec5 as a host target for AVR1. Interaction between AVR1 and Sec5 was confirmed in planta by co-immunoprecipitation and bimolecular fluorescent complementation. Although A-L shares high sequence similarity with AVR1, we found that it is not able to interact with Sec5. Sec5 was shown to be required for proper plant defense against P. infestans. The role of Sec5 in plant response upon pathogen attack was further supported by its role in callose deposition and in secretion of the pathogenesis-related protein PR-1, which indicates that Sec5 plays a crucial role in vesicle trafficking during host defense. AVR1 is able to suppress callose deposition while A-L is not, which suggests that P. infestans manipulates host vesicle trafficking by secretion of AVR1 to target Sec5. Overall, our findings unravelled a novel strategy that oomycete pathogens exploit in order to modulate host defense. In Chapter 5 we further analysed the potential virulence activities of AVR1 and A-L. Both AVR1 and A-L were able to promote P. infestans colonization, indicating that both are genuine P. infestans virulence factors. Moreover, AVR1 was found to suppress not only callose deposition, but also Sec5-dependent cell death induced by the P. infestans elicitors INF1 and CRN2. In contrast, A-L was neither able to suppress Sec5-dependent nor Sec5-independent cell death. The conserved C-terminal motifs and regions required for virulence activity of AVR1 were investigated using AVR1 truncated constructs. In addition, the conserved C-terminal motifs and regions of AVR1 required for Sec5 interaction were studied by Y2H assays. Although the T-region of AVR1 was found to be sufficient to facilitate P. infestans colonization and suppression of CRN2-induced cell death, it could not fully accommodate the interaction of AVR1 with Sec5. Instead, both the Y motif and the T-region of AVR1 appear to be required for Sec5 targeting. Next to Sec5, the role of other exocyst subunits in Phytophthora resistance was studied (Chapter 6). The evolutionary relationships of exocyst subunits from three Solanaceous plants, i.e. Nicotiana benthamiana, tomato and potato, were investigated in comparison to their Arabidopsis orthologs. Virus-induced gene silencing in N. benthamiana of the majority of the exocyst subunit genes (exo84s were not yet included) showed that, except for some Exo70 members, all other tested exocyst subunits are required for plant defense against P. infestans and callose deposition. In addition, all of the analysed exocyst subunit gene-silenced tomato plants showed gain of susceptibility to both P. infestans and Phytophthora capsici. In Chapter 7, our findings obtained in this thesis on the mechanisms of AVR1-triggered host immunity and susceptibility are discussed in a broader perspective with emphasis on the current developments in the field of effector biology.

Summary Late blight, caused by the oomycete Phytophthora infestans, is one of the most devastating potato diseases worldwide. To successfully colonize its host, P. infestans secretes a plethora of RXLR effectors that translocate into host cells to modulate plant defense. The RXLR effectors form the largest and most diverse effector family in oomycete plant pathogens, and include several that were demonstrated to trigger host resistance mediated by intracellular host immune receptors. Chapter 1 is a summary focussing on the molecular mechanisms underlying host–pathogen interactions. It introduces the multi-layered innate immune system of plants, as well as the strategies that pathogens exploit to circumvent and suppress host defense. Furthermore, it highlights the importance of vesicle-trafficking during plant defense. The central subject of this thesis is AVR1, one of the race-specific avirulence (AVR) factors of P. infestans. AVR1 triggers plant resistance mediated by its corresponding potato Nucleotide-binding Leucine-rich repeat (NLR) resistance protein R1. P. infestans isolates that are avirulent on R1-containing potato cultivars always contain AVR1, while virulent isolates lack AVR1 but contain a related gene that we baptized as AVR1-like. AVR1 has all hallmarks of a typical RXLR effector; it contains a signal peptide, an RXLR domain and a C-terminal effector domain that contains two W motifs and one Y motif. In addition, it has, at the very end a stretch of 38 amino acids in length that we named the Tail (T)-region. AVR1-like, or in short A-L, shares high sequence similarity with AVR1. However, due to a premature stop codon the 38 amino acid T-region is missing. Chapter 2 explores the conserved motifs and regions in the C-terminal effector domain of AVR1 that are required to trigger R1-mediated hypersensitive response (HR). Various truncated and chimeric constructs of AVR1 and A-L were generated and assayed for their ability to elicit R1-mediated HR. Results show that the T-region of AVR1 plays an important role in HR activation. Furthermore, we revealed that R1 recognizes two epitopes in AVR1, one located in the C-terminal region containing the conserved W and Y motifs, and one comprised by the T region. In Chapter 3 the subcellular localization of AVR1 and R1 was investigated. Both were demonstrated to be nucleocytoplasmic proteins. We artificially modified the nucleocytoplasmic partitioning of AVR1 and R1 using nuclear localization and export signals (NLS/NES), and studied the effect on R1-AVR1 recognition. This revealed that nuclear localization of both AVR1 and R1 is important to induce R1-mediated immunity. In addition, we showed that AVR1-mediated suppression of CRN2-induced cell death is dependent on cytosolic localization of AVR1. In Chapter 4, we investigated how AVR1 modulates host defense. In a yeast two-hybrid screening we identified the exocyst subunit Sec5 as a host target for AVR1. Interaction between AVR1 and Sec5 was confirmed in planta by co-immunoprecipitation and bimolecular fluorescent complementation. Although A-L shares high sequence similarity with AVR1, we found that it is not able to interact with Sec5. Sec5 was shown to be required for proper plant defense against P. infestans. The role of Sec5 in plant response upon pathogen attack was further supported by its role in callose deposition and in secretion of the pathogenesis-related protein PR-1, which indicates that Sec5 plays a crucial role in vesicle trafficking during host defense. AVR1 is able to suppress callose deposition while A-L is not, which suggests that P. infestans manipulates host vesicle trafficking by secretion of AVR1 to target Sec5. Overall, our findings unravelled a novel strategy that oomycete pathogens exploit in order to modulate host defense. In Chapter 5 we further analysed the potential virulence activities of AVR1 and A-L. Both AVR1 and A-L were able to promote P. infestans colonization, indicating that both are genuine P. infestans virulence factors. Moreover, AVR1 was found to suppress not only callose deposition, but also Sec5-dependent cell death induced by the P. infestans elicitors INF1 and CRN2. In contrast, A-L was neither able to suppress Sec5-dependent nor Sec5-independent cell death. The conserved C-terminal motifs and regions required for virulence activity of AVR1 were investigated using AVR1 truncated constructs. In addition, the conserved C-terminal motifs and regions of AVR1 required for Sec5 interaction were studied by Y2H assays. Although the T-region of AVR1 was found to be sufficient to facilitate P. infestans colonization and suppression of CRN2-induced cell death, it could not fully accommodate the interaction of AVR1 with Sec5. Instead, both the Y motif and the T-region of AVR1 appear to be required for Sec5 targeting. Next to Sec5, the role of other exocyst subunits in Phytophthora resistance was studied (Chapter 6). The evolutionary relationships of exocyst subunits from three Solanaceous plants, i.e. Nicotiana benthamiana, tomato and potato, were investigated in comparison to their Arabidopsis orthologs. Virus-induced gene silencing in N. benthamiana of the majority of the exocyst subunit genes (exo84s were not yet included) showed that, except for some Exo70 members, all other tested exocyst subunits are required for plant defense against P. infestans and callose deposition. In addition, all of the analysed exocyst subunit gene-silenced tomato plants showed gain of susceptibility to both P. infestans and Phytophthora capsici. In Chapter 7, our findings obtained in this thesis on the mechanisms of AVR1-triggered host immunity and susceptibility are discussed in a broader perspective with emphasis on the current developments in the field of effector biology.

Als we deskundigen de laatste twee jaar vroegen naar hun mening over kastanjebloedingsziekte was het antwoord meestal; “niet opgelost, maar duidelijk minder actueel en agressief.” In Ridderkerk en Utrecht lijkt de situatie inmiddels heel anders. Van de ongeveer 550 kastanjes die in Ridderkerk begin 2010 nog stonden zijn er al 150 gekapt. Een inboet van 30 procent dus. Tot de slachtoffers behoort ook de cv Aesculus x carnea “Plantierensis” . Uit eerder onderzoek bleek deze soort juist minder gevoelig. In Utrecht is de situatie vergelijkbaar dramatisch.

Als we deskundigen de laatste twee jaar vroegen naar hun mening over kastanjebloedingsziekte was het antwoord meestal; “niet opgelost, maar duidelijk minder actueel en agressief.” In Ridderkerk en Utrecht lijkt de situatie inmiddels heel anders. Van de ongeveer 550 kastanjes die in Ridderkerk begin 2010 nog stonden zijn er al 150 gekapt. Een inboet van 30 procent dus. Tot de slachtoffers behoort ook de cv Aesculus x carnea “Plantierensis” . Uit eerder onderzoek bleek deze soort juist minder gevoelig. In Utrecht is de situatie vergelijkbaar dramatisch.

Soortgevoeligheid van Gerbera voor Botrytis bepaalt voor een deel of er problemen met Botrytis in de keten kunnen optreden. In dit project is nagegaan of een betrouwbare toets voor voorspelling van botrytisgevoeligheid ontwikkeld kan worden. In dit rapport staat een protocol beschreven dat vervolgens getoetst is. Het protocol gaat uit van oogst van 20 oogstbare bloemen die voor gewaterd worden, besmet worden met inoculum, gedurende 2 dagen bij hoge RV gezet worden en tenslotte in een uitbloeiruimte tot uitbloei kunnen komen. Om een uitspraak te kunnen doen over de gevoeligheid van een ras moeten bloemen van minimaal 3 bedrijven getest worden op minimaal 3 dagen verspreid over het jaar. Het is daarmee voor veredelaars een weinig toepasbare toets.

Soortgevoeligheid van Gerbera voor Botrytis bepaalt voor een deel of er problemen met Botrytis in de keten kunnen optreden. In dit project is nagegaan of een betrouwbare toets voor voorspelling van botrytisgevoeligheid ontwikkeld kan worden. In dit rapport staat een protocol beschreven dat vervolgens getoetst is. Het protocol gaat uit van oogst van 20 oogstbare bloemen die voor gewaterd worden, besmet worden met inoculum, gedurende 2 dagen bij hoge RV gezet worden en tenslotte in een uitbloeiruimte tot uitbloei kunnen komen. Om een uitspraak te kunnen doen over de gevoeligheid van een ras moeten bloemen van minimaal 3 bedrijven getest worden op minimaal 3 dagen verspreid over het jaar. Het is daarmee voor veredelaars een weinig toepasbare toets.

Botryotinia fuckeliana of grauwe schimmel (Botrytis) is een algemeen voorkomende schimmelziekte, die schade kan aanrichten in een breed scala aan tuinbouwgewassen. Botrytis is een zwakteparasiet. Daarom zijn verzwakt, afstervend weefsel en wonden gemakkelijke invalspoorten. De aanpak bestaat uit een combinatie van maatregelen om de sporendruk laag te houden, (klimaat)maatregelen om infectie te voorkomen en overige maatregelen, zoals de keuze voor minder gevoelige rassen, meer belichting in de ochtend en avond en weerstandsverhogende micro-nutriënten.

Botryotinia fuckeliana of grauwe schimmel (Botrytis) is een algemeen voorkomende schimmelziekte, die schade kan aanrichten in een breed scala aan tuinbouwgewassen. Botrytis is een zwakteparasiet. Daarom zijn verzwakt, afstervend weefsel en wonden gemakkelijke invalspoorten. De aanpak bestaat uit een combinatie van maatregelen om de sporendruk laag te houden, (klimaat)maatregelen om infectie te voorkomen en overige maatregelen, zoals de keuze voor minder gevoelige rassen, meer belichting in de ochtend en avond en weerstandsverhogende micro-nutriënten.

Op basis van onderzoek in 2008 naar de gevoeligheid van onderstammen voor wortelknobbelaaltjes (onderzoek in voor- en najaar) bij Wageningen UR Glastuinbouw, zijn er een aantal onderstammen uitgelicht die relatief weinig wortelknobbels vertoonden. Deze zijn in 2009 op enkele biologische bedrijven getoetst op hun gebruikswaarde. Het doel was om na te gaan wat de productie en kwaliteit is van enkele onderstammen die zich positief onderscheiden voor wat betreft gevoeligheid voor wortelknobbelaaltjes ofwel die relatief weinig wortelknobbels vertonen

Op basis van onderzoek in 2008 naar de gevoeligheid van onderstammen voor wortelknobbelaaltjes (onderzoek in voor- en najaar) bij Wageningen UR Glastuinbouw, zijn er een aantal onderstammen uitgelicht die relatief weinig wortelknobbels vertoonden. Deze zijn in 2009 op enkele biologische bedrijven getoetst op hun gebruikswaarde. Het doel was om na te gaan wat de productie en kwaliteit is van enkele onderstammen die zich positief onderscheiden voor wat betreft gevoeligheid voor wortelknobbelaaltjes ofwel die relatief weinig wortelknobbels vertonen

This research project aims to understand the factors that influence susceptibility to HIV for female and male students in LAMS (Lycee Agricole Meidji o Sekou in Benin), in order to contribute to the reduction of the phenomenon. The results of questionnaires, group discussions and interviews show the perceptions, knowledge, behaviour and risk factors that influence susceptibility to HIV for female and male students in LAMS.

Kun je zien of koeien voldoende weerstand hebben, zodat ze minder snel ziek worden? In een proef van de Animal Sciences Group van Wageningen UR en het Louis Bolk Instituut is getracht parameters te vinden om de weerstand van een koe te bepalen. Komen waarnemingen over de koe (glanzende vacht, alertheid, bloedanalyse, maar ook energiemetingen) overeen met de gezondheid van de dieren? Vooral waarnemingen in combinatie met bloedanalyses lijkt een redelijk beeld te geven over de gezondheissituatie van het dier. Meer onderzoek is nodig.

Single-species acute toxicity data and (micro)mesocosm data were collated for 16 insecticides. These data were used to investigate the importance of test-species selection in constructing species sensitivity distributions (SSDs) and the ability of estimated hazardous concentrations (HCs) to protect freshwater aquatic ecosystems. A log-normal model was fitted to a minimum of six data points, and the resulting distribution was used to estimate lower (95% confidence), median (50% confidence), and upper (5% confidence) 5% HC (HC5) values. Species sensitivity distributions for specific taxonomic groups (vertebrates, arthropods, nonarthropod invertebrates), habitats (saltwater, freshwater, lentic, lotic), and geographical regions (Palaearctic, Nearctic, temperate, tropical) were compared. The taxonomic composition of the species assemblage used to construct the SSD does have a significant influence on the assessment of hazard, but the habitat and geographical distribution of the species do not. Moreover, SSDs constructed using species recommended in test guidelines did not differ significantly from those constructed using nonrecommended species. Hazardous concentrations estimated using laboratory-derived acute toxicity data for freshwater arthropods (i.e., the most sensitive taxonomic group) were compared to the response of freshwater ecosystems exposed to insecticides. The sensitivity distributions of freshwater arthropods were similar for both field and laboratory exposure, and the lower HC5 (95% protection with 95% confidence) estimate was protective of adverse ecological effects in freshwater ecosystems. The corresponding median HC5 (95% protection level with 50% confidence) was generally protective of single applications of insecticide but not of continuous or multiple applications. In the latter cases, a safety factor of at least five should be applied to the median HC5.

Literatuurstudie naar weerstandsbevorderende middelen voor de productie van landbouwgewassen in het kader van het onderzoeksprogramma 'Verbetering van de kwaliteit van plantaardig uitgangsmateriaal voor biologische en duurzame landbouw'. Er wordt ingegaan op de achtergrond van weerstandsbevordering, verschillende categorieën natuurlijke middelen die de weerstand kunnen bevorderen met de nadruk op middelen die geschikt zijn voor zaadbehandeling in de biologische landbouw, en de regelgeving in Europa

Literatuurstudie naar weerstandsbevorderende middelen voor de productie van landbouwgewassen in het kader van het onderzoeksprogramma 'Verbetering van de kwaliteit van plantaardig uitgangsmateriaal voor biologische en duurzame landbouw'. Er wordt ingegaan op de achtergrond van weerstandsbevordering, verschillende categorieën natuurlijke middelen die de weerstand kunnen bevorderen met de nadruk op middelen die geschikt zijn voor zaadbehandeling in de biologische landbouw, en de regelgeving in Europa

Tweede artikel van een serie van drie over een vierjarig onderzoek van het Louis Bolk Instituut en Plant Research International naar Phytophthora infestans. In veldproeven werden een groot aantal Phytophthora-resistente rassen geteeld onder sterke infectiedruk en beoordeeld op opbrengst en kwaliteit, en de relatie tussen groeiduur en opbrengst. Ook werd gekeken naar de mogelijkheden om de resistentie te versterken door sturing van het stikstofaanbod en door toepassing van endofytische bacterieën die de schimmel kunnen remmen

Colorectal cancer is the second most common cancer in the Western world and is thought to arise mainly from colorectal adenomas. Red meat and alcohol intake and (long-term) cigarette smoking probably increase colorectal tumor risk. Although risk increase was found to be weak, certain subgroups might be more susceptible to these carcinogens because of inherited polymorphisms resulting in increased activation of potential carcinogens. In this thesis, we investigated whether meat consumption, cigarette smoking, and alcohol intake, in combination with such genetic polymorphisms, increase the risk of colorectal adenomas and colorectal cancer.For this purpose, we used two different study populations. One was an adenoma case-control study with 440 adenoma cases and 447 polyp-free controls recruited among those undergoing endoscopy at eight Dutch outpatient clinics between June 1997 and June 2000. The other was a Dutch prospective cohort enrolled between January 1987 and December 1991. Follow-up for 8.5 years resulted in 102 colorectal cancer cases. We compared these cases with a random sample of 537 controls frequency-matched on age, sex and town. Information on dietary and lifestyle habits was collected through self-administered questionnaires. DNA was isolated from whole blood and genetic polymorphisms were subsequently determined by use of standardized methods.We found that colorectal adenoma risk was not increased with high meat consumption (OR 1.2, 95% CI 0.8-1.9) or unfavorable meat preparation methods. These null-associations were not modified by genetic polymorphisms affecting metabolism of heterocyclic amines that may be formed during preparation of meat at high temperatures ( N -acetyltransferases ( NAT ) 1 and 2, sulfotransferase ( SULT ) 1A1 , and glutathione S- transferases ( GST ) M1 and T1 ). Long-term cigarette smoking increased adenoma risk (OR 2.4, 95% CI 1.4-4.1 for smoking for more than 25 years compared to never smokers). Although most pronounced in those with fast SULT1A1 (OR 4.3, 95% CI 1.6-11.8) and slow NAT2 variants (OR 3.5, 95% CI 1.9-6.4), there was no statistically significant effect modification by genetic polymorphisms involved in metabolism of arylamines and polycyclic aromatic hydrocarbons from cigarette smoke ( NAT1 , NAT2 , SULT1A1 , GSTM1 , GSTT1 , and epoxide hydrolase). Alcohol consumption increased colorectal adenoma risk especially among women (OR 1.8, 95% CI 1.0-3.2 for 10 or more drinks weekly versus less than one drink per week). Among men, adenoma risk increased only with consumption of more than 21 drinks per week (OR 1.8, 95% CI 0.9-3.8). Alcohol is metabolized to carcinogenic acetaldehyde by alcohol dehydrogenase (ADH3). The association between alcohol and adenomas was weakly - but not statistically significantly - stronger among those with the fast ADH3 variant compared to those with imputed slow phenotypes.Colorectal cancer risk increased slightly with frequent red meat consumption (OR 1.6, 95% CI 0.9-2.9, highest vs. lowest intake). Genetic polymorphisms in NAT1 , NAT2 , and GSTM1 did not importantly modify this association. Risk of colorectal cancer was increased with smoking duration, but only among former smokers having smoked for more than 15 years (OR 2.7, 95% CI 1.0-7.4) compared to former smokers having smoked for shorter time. NAT1 , NAT2 , and GSTM1 polymorphisms did not influence this association.In summary, the results of our studies do not point toward strong modifying effects of genetic polymorphisms of enzymes involved in carcinogen metabolism, which is in accordance with results of similar studies on colorectal tumors. Such effects may however be present, but possibly, we were not able to demonstrate them. To elucidate the potential role of genetic susceptibility in colorectal carcinogenesis, alternative epidemiologic study designs and statistical methods should be considered. These observational studies should be conducted simoultaneously with experimental studies aiming to generate more biological knowledge on the diverse processes leading to colorectal tumorigenesis in humans.

The aim of the study is: 1) to substantiate the role of uncertainties and assumptions within the SSD concept 2) to derive scientific conclusions on various aspects of the SSD concept 3) to express the limitations and points of concern regarding the use of the SSD-based extrapolation procedure in the derivation of environmentakl quality criteria and ecological risk assessment

Colorectal cancer is the second most common cancer in the Western world and is thought to arise mainly from colorectal adenomas. Red meat and alcohol intake and (long-term) cigarette smoking probably increase colorectal tumor risk. Although risk increase was found to be weak, certain subgroups might be more susceptible to these carcinogens because of inherited polymorphisms resulting in increased activation of potential carcinogens. In this thesis, we investigated whether meat consumption, cigarette smoking, and alcohol intake, in combination with such genetic polymorphisms, increase the risk of colorectal adenomas and colorectal cancer.For this purpose, we used two different study populations. One was an adenoma case-control study with 440 adenoma cases and 447 polyp-free controls recruited among those undergoing endoscopy at eight Dutch outpatient clinics between June 1997 and June 2000. The other was a Dutch prospective cohort enrolled between January 1987 and December 1991. Follow-up for 8.5 years resulted in 102 colorectal cancer cases. We compared these cases with a random sample of 537 controls frequency-matched on age, sex and town. Information on dietary and lifestyle habits was collected through self-administered questionnaires. DNA was isolated from whole blood and genetic polymorphisms were subsequently determined by use of standardized methods.We found that colorectal adenoma risk was not increased with high meat consumption (OR 1.2, 95% CI 0.8-1.9) or unfavorable meat preparation methods. These null-associations were not modified by genetic polymorphisms affecting metabolism of heterocyclic amines that may be formed during preparation of meat at high temperatures ( N -acetyltransferases ( NAT ) 1 and 2, sulfotransferase ( SULT ) 1A1 , and glutathione S- transferases ( GST ) M1 and T1 ). Long-term cigarette smoking increased adenoma risk (OR 2.4, 95% CI 1.4-4.1 for smoking for more than 25 years compared to never smokers). Although most pronounced in those with fast SULT1A1 (OR 4.3, 95% CI 1.6-11.8) and slow NAT2 variants (OR 3.5, 95% CI 1.9-6.4), there was no statistically significant effect modification by genetic polymorphisms involved in metabolism of arylamines and polycyclic aromatic hydrocarbons from cigarette smoke ( NAT1 , NAT2 , SULT1A1 , GSTM1 , GSTT1 , and epoxide hydrolase). Alcohol consumption increased colorectal adenoma risk especially among women (OR 1.8, 95% CI 1.0-3.2 for 10 or more drinks weekly versus less than one drink per week). Among men, adenoma risk increased only with consumption of more than 21 drinks per week (OR 1.8, 95% CI 0.9-3.8). Alcohol is metabolized to carcinogenic acetaldehyde by alcohol dehydrogenase (ADH3). The association between alcohol and adenomas was weakly - but not statistically significantly - stronger among those with the fast ADH3 variant compared to those with imputed slow phenotypes.Colorectal cancer risk increased slightly with frequent red meat consumption (OR 1.6, 95% CI 0.9-2.9, highest vs. lowest intake). Genetic polymorphisms in NAT1 , NAT2 , and GSTM1 did not importantly modify this association. Risk of colorectal cancer was increased with smoking duration, but only among former smokers having smoked for more than 15 years (OR 2.7, 95% CI 1.0-7.4) compared to former smokers having smoked for shorter time. NAT1 , NAT2 , and GSTM1 polymorphisms did not influence this association.In summary, the results of our studies do not point toward strong modifying effects of genetic polymorphisms of enzymes involved in carcinogen metabolism, which is in accordance with results of similar studies on colorectal tumors. Such effects may however be present, but possibly, we were not able to demonstrate them. To elucidate the potential role of genetic susceptibility in colorectal carcinogenesis, alternative epidemiologic study designs and statistical methods should be considered. These observational studies should be conducted simoultaneously with experimental studies aiming to generate more biological knowledge on the diverse processes leading to colorectal tumorigenesis in humans.

The aim of the study is: 1) to substantiate the role of uncertainties and assumptions within the SSD concept 2) to derive scientific conclusions on various aspects of the SSD concept 3) to express the limitations and points of concern regarding the use of the SSD-based extrapolation procedure in the derivation of environmentakl quality criteria and ecological risk assessment

PPO onderzocht op drie locaties (zand, klei, veen) de gebruikswaarde van een groot aantal Monarda-cultivars (bergamotplant), waarbij de nadruk lag op de vatbaarheid voor meeldauw. In een tabel het aanbevolen sortiment met gegevens over bloemkleur, bloeitijd en hoogte, een beoordeling van groeikracht, stevigheid en gezondheid, en de waardering door de KVBC (sterrenkeuring voor het gebruik als tuinplant)