Use of genotyping-by-sequencing to determine the genetic structure in the medicinal plant chamomile, and to identify flowering time and alpha-bisabolol associated SNP-loci by genome-wide association mapping.

BACKGROUND: Chamomile (Matricaria recutita L.) has a long history of use in herbal medicine with various applications, and the flower heads contain numerous secondary metabolites which are medicinally active. In the major crop plants, next generation sequencing (NGS) approaches are intensely applied to exploit genetic resources, to develop genomic resources and to enhance breeding. Here, genotyping-by-sequencing (GBS) has been used in the non-model medicinal plant chamomile to evaluate the genetic structure of the cultivated varieties/populations, and to perform genome wide association study (GWAS) focusing on genes with large effect on flowering time and the medicinally important alpha-bisabolol content. RESULTS: GBS analysis allowed the identification of 6495 high-quality SNP-markers in our panel of 91 M. recutita plants from 33 origins (2-4 genotypes each) and 4 M. discoidea plants as outgroup, grown in the greenhouse in Gatersleben, Germany. M. recutita proved to be clearly distinct from the outgroup, as was demonstrated by different cluster and principal coordinate analyses using the SNP-markers. Chamomile genotypes from the same origin were mostly genetically similar. Model-based cluster analysis revealed one large group of tetraploid genotypes with low genetic differentiation including 39 plants from 14 origins. Tetraploids tended to display lower genetic diversity than diploids, probably reflecting their origin by artificial polyploidisation from only a limited set of genetic backgrounds. Analyses of flowering time demonstrated that diploids generally flowered earlier than tetraploids, and the analysis of alpha-bisabolol identified several tetraploid genotypes with a high content. GWAS identified highly significant (P < 0.01) SNPs for flowering time (9) and alpha-bisabolol (71). One sequence harbouring SNPs associated with flowering time was described to play a role in self-pollination in Arabidopsis thaliana, whereas four sequences harbouring SNPs associated with alpha-bisabolol were identified to be involved in plant biotic and abiotic stress response in various plants species. CONCLUSIONS: The first genomic resource for future applications to enhance breeding in chamomile was created, andanalyses of diversity will facilitate the exploitation of these genetic resources. The GWAS data pave the way for future research towards the genetics underlying important traits in chamomile, the identification of marker-trait associations, and development of reliable markers for practical breeding.

Induction of phenolic metabolites and physiological changes in chamomile plants in relation to nitrogen nutrition.

Alternative tools, such as the manipulation of mineral nutrition, may affect secondary metabolite production and thus the nutritional value of food/medicinal plants. We studied the impact of nitrogen (N) nutrition (nitrate/NO3(-) or ammonium/NH4(+) nitrogen) and subsequent nitrogen deficit on phenolic metabolites and physiology in Matricaria chamomilla plants. NH4(+)-fed plants revealed a strong induction of selected phenolic metabolites but, at the same time, growth, Fv/Fm, tissue water content and soluble protein depletion occurred in comparison with NO3(-)-fed ones. On the other hand, NO3(-)-deficient plants also revealed an increase in phenolic metabolites but growth depression was not observed after the given exposure period. Free amino acids were more accumulated in NH4(+)-fed shoots (strong increase in arginine and proline mainly), while the pattern of roots' accumulation was independent of N form. Among phenolic acids, NH4(+) strongly elevated mainly the accumulation of chlorogenic acid. Within flavonoids, flavonols decreased while flavones strongly increased in response to N deficiency. Coumarin-related metabolites revealed a similar increase in herniarin glucosidic precursor in response to N deficiency, while herniarin was more accumulated in NO3(-)- and umbelliferone in NH4(+)-cultured plants. These data indicate a negative impact of NH4(+) as the only source of N on physiology, but also a higher stimulation of some valuable phenols. Nitrogen-induced changes in comparison with other food/crop plants are discussed.

Desenvolvimento da camomila, rendimento e qualidade do óleo essencial em diferentes idades de colheita / Development of chamomile and essential oil yield and quality at different harvesting times

A camomila é uma planta herbácea, anual e aromática com várias propriedades medicinais. No Brasil, vem sendo cultivada em área representativa na região Sul devido, principalmente, às exigências climáticas da espécie, e por ter sido introduzida por imigrantes europeus nesta região. A colheita da camomila é realizada em várias colheitas, contudo, a idade adequada da planta para o início desta atividade, objetivado a maior produção de capítulos florais, bem como o rendimento e qualidade de óleos essenciais, ainda não foi determinada para a espécie. Neste trabalho, investigou-se o efeito de diferentes épocas de colheita no desenvolvimento de capítulos florais e no acúmulo e qualidade do óleo essencial da camomila. O experimento foi conduzido no município de Piraquara (PR). O delineamento experimental foi inteiramente casualizado, com cinco tratamentos (85, 92, 99, 106 e 113 dias após a emergência das plântulas) e cinco repetições. Após cada colheita, avaliou-se o acúmulo de massa seca de capítulos, rendimento, produção e composição do óleo essencial. A extração de óleo foi realizada por hidrodestilação e as amostras analisadas por cromatografia gasosa. Concluiu-se que aos 85 dias após a emergência (DAE) os capítulos apresentaram maior rendimento de óleo. Porém, após três semanas ocorreu a maior produção de massa de capítulos, resultando em maiores médias de produtividade de óleo essencial. A partir de 113 DAE houve redução do rendimento de óleo essencial e no teor de alguns constituintes.

Chamomile is an aromatic herb with many medicinal properties. In Brazil, it is cultivated on an extensive area in the Southern region mainly because of its climatic conditions for this culture. Chamomile harvesting is done by several cuttings of the entire plant, but the ideal starting point of this activity to obtain high production of flower heads has not been determined. This work investigated the effect of the harvesting period of chamomile on flower head development and essential oil accumulation. The experiment was carried out at field conditions in Piraquara, Paraná, Brazil. The experimental design was completely randomized with five treatments (85, 92, 99, 106 and 113 days after emergency) and five replications. After each harvest, dry mass accumulation and essential oil yield and quality were evaluated. Essential oil extraction was performed by hydrodistillation, and the sample was analyzed by gas chromatography. When harvested 85 days after emergency, the flower heads presented the highest essential oil content, but three weeks later the highest flower head dry mass was obtained, which resulted on great essential oil productivity. From 113 days after emergency, there is a reduction in essential oil yield and production.