ABSTRACT The hybridization technique through bridging plants uses indirect or intermediate crosses as an instrument to cross parents with different levels of ploidy. The objective of this work is to obtain hybrids that can be used as bridges in crosses with ornamental species of the genus Petunia and/or Calibrachoa. This can overcome the incompatibility barriers that arise when hybridization is direct between species. To obtain hybrids, directed crosses were carried out between: P. integrifolia, P. inflata, P. interior and P. axillaris subspecies axillaris; with C. linoides, C. ovalifolia, C. thymifolia, C. caesia, C. x hybrida var. Kabloom blue and C. x hybrida var. Kabloom yellow. The first step was the detection of putative combinations. In order to do so, the fruiting rate, which was defined as the formation of fruits every 10 intergeneric crosses, was compared, and variable results were found for each combination. A hybrid plant was obtained between P. inflata x C. x hybrida 'kabloom yellow', where morphological, pollen viability and meiotic studies confirm its nature. With the bridge hybrid obtained, reciprocal crosses were carried out with all of the Petunia and Calibrachoa species involved, obtaining 31 hybrid plants sharing morphological characteristics of both genera in combinations with P. inflata and P. integrifolia. The identification of intergenetic crosses with a high fruiting rate opens doors for future studies on the development of hybrid embryos. This would allow determining the optimum time to apply embryo rescue techniques in order to increase the number of hybrid bridging plants.

RESUMEN La técnica de hibridación a través de plantas puentes, utiliza cruzamientos indirectos o intermedios como instrumento para cruzar parentales con diferente nivel de ploidía. El objetivo de este trabajo fue obtener híbridos que puedan ser utilizados como puentes en cruzamientos con especies ornamentales del género Petunia y/o Calibrachoa. Lo anterior permite sortear las barreras de incompatibilidad que surgen cuando la hibridación es directa entre especies. Para la obtención de híbridos se realizaron cruzamientos dirigidos entre: P. integrifolia, P. inflata, P. interior y P. axillaris subespecie axillaris; con C. linoides, C. ovalifolia, C. thymifolia, C. caesia, C. x hybrida var. Kabloom blue y C. x hybrida var. Kabloom yellow. El primer paso fue la detección de combinaciones putativas, para lo cual se comparó la tasa de fructificación definida como la formación de frutos cada 10 cruzamientos intergenéricos, encontrando resultados variables para cada combinación. Se obtuvo una planta híbrida entre P. inflata x C. x hybrida 'Kabloom yellow', en que los estudios morfológicos, de viabilidad del polen y meióticos confirman su naturaleza. Con el híbrido puente obtenido se realizaron cruzamientos recíprocos con todas de las especies de Petunia y Calibrachoa involucradas, obteniendo 31 plantas híbridas que comparten características morfológicas de ambos géneros en las combinaciones con P. inflata y P. integrifolia. La identificación de cruzamientos intergénericos con alto porcentaje de fructificación abre una ventana para futuros estudios sobre el desarrollo de los embriones híbridos. Esto permitirán conocer el momento oportuno para aplicar técnicas de rescate embrionario con el objeto de aumentar el número de plantas híbridas puentes.

Resumo Calibrachoa hybrida (calibrachoa ou falsa-petúnia) é uma planta ornamental com importância crescente no mercado devido a existência de inúmeras variedades que podem ser cultivadas em vasos, jardins e paisagismo. A característica marcante é sua floração abundante e de cores vibrantes. Em outubro de 2019, foi obtido um isolado de fungo em folhas basais de calibrachoa, com manchas foliares marrons e irregulares, em plantas cultivadas em Buenos Aires, Argentina. O objetivo do presente estudo foi identificar o agente causal das manchas foliares em calibrachoa para expandir o conhecimento sobre as doenças prevalentes nesta importante planta ornamental. O isolado foi identificado por meio de marcadores morfológicos e moleculares, bem como a sua patogenicidade foi comprovada por meio do postulado de Koch. O fungo foi identificado como Nigrospora oryzae, sendo que presente artigo, além de relatar a sua patogenicidade, parece ser o primeiro registro desta doença em calibrachoa no mundo.

Abstract Calibrachoa hybrida (calibrachoa, million bells) is a flowering ornamental with increasing importance due to the existence of many successful cultivars for growing indoors in containers and planting in the garden and landscape. The outstanding characteristic is a profuse flowering and intense colour. In October 2019, a fungal isolate was obtained from basal calibrachoa leaves with irregular brown leaf spots, in plants cultivated in Buenos Aires, Argentina. The aim of the present study was to identify the cause of the disease in this ornamental genus, to expand knowledge about prevalent pathologies. The isolate was identified using morphological and molecular markers, and the pathogenicity tests were fulfilled. This paper reports that Nigrospora oryzae is pathogenic to calibrachoa, which seems to be the first record of this leaf spot disease in the world.

Resumo A petúnia é uma planta ornamental muito importante, com uma ampla gama de cores e tamanhos de flores, e a maioria das cultivares são propagadas através de sementes. A esterilidade citoplasmática masculina (CMS) é um caráter hereditário da mãe, determinado por genes mitocondriais que resultam em prejuízo ao desenvolvimento do pólen. O citoplasma macho estéril único e bem caracterizado em Petúnia é um recurso valioso para a produção de híbridos, pois previne a autofecundação das plantas-mãe e garante a pureza do F1. A introgressão do citoplasma macho estéril nas linhas de elite de Petúnia é alcançada seguindo um esquema de retrocruzamento e pode ser assistida usando marcadores moleculares associados à característica de interesse. O objetivo deste estudo foi desenvolver um marcador molecular para identificar o citoplasma macho estéril de Petunia. Foi desenhado um marcador baseado em PCR que amplificou uma região de urfS associada à CMS mitocondrial apenas em plantas masculinas estéreis. Os resultados mostraram amplificação por PCR diferencial de um produto de ≈ 600 bp em plantas portadoras de citoplasma estéril masculino em quatro espécies de Petúnia e suas gerações F1s e BC1. Uma reação de PCR multiplex foi subsequentemente configurada, adicionando iniciadores específicos amplificando um produto de ≈ 800 bp da região conservada trnT-trn-L do genoma do cloroplasto como um controle positivo, para identificar inequivocamente os tipos de citoplasma como normal ou estéril. Um marcador molecular rápido, simples e preciso está agora disponível para auxiliar no melhoramento genético de híbridos F1 em Petunia.

Abstract Petunia is a very important ornamental plant with a broad range of flower colour and size, and most of the cultivars grown are propagated through seeds. Cytoplasmic male sterility (CMS) is a maternally inherited character determined by mitochondrial genes that results in impaired pollen development. The unique and well characterized male sterile cytoplasm in Petunia is a valuable resource for hybrids production because it prevents self-fertilization of mother plants and ensures the purity of F1s. Introgression of the male sterile cytoplasm in elite lines of Petunia is achieved following a backcross scheme and can be assisted using molecular markers associated to the trait of interest. The objective of this study was to develop a molecular marker to identify the male sterile cytoplasm of Petunia. A PCR-based marker amplifying a region of the mitochondrial CMS-associated urfS only in the male sterile plants was designed. Results showed differential PCR amplification of a ≈ 600 bp product in plants carrying male sterile cytoplasm in four Petunia species and their F1s and BC1 generations. A multiplex PCR reaction was subsequently set up, adding specific primers amplifying a ≈ 800 bp product from the conserved region trnT-trn-L of the chloroplast genome as a positive control in order to unambiguously identify the cytoplasm types as normal or sterile. A rapid, simple and precise molecular marker is now available for assisting breeding of F1 hybrids in Petunia.

The commercial value of Bacopa monnieri, a widespread herbaceous plant in Argentina, can be substantially improved increasing its flower size by chromosome doubling with colchicine. MS supplemented with 0.25 mg/L of 6-benzylaminopurine proved to be an appropriate medium for the in vitro multiplication of nodal segments of B. monnieri. Polyploidization was achieved submerging nodal segments during 24 or 48 hrs in colchicine solution 0.001 and 0.01% P/V, in 1% DMSO. Segments submerged in water and in DMSO 1% aqueous solution were used as controls. DNA contents from recovered plants was measured and characterized and their phenotype described. Two tetraploid plants originated from independent events were detected. These plants showed significant differences in size and colour both in leaves and flowers compared to untreated controls.

The genus Scoparia is native from Argentina. The diversity of colours and shape of their flowers bestows this genus a very interesting ornamental potential. The purpose of the present study is to explore the Scoparia species germplasm by means of in vitro polyploidization in order to improve their ornamental qualities. Accessions of S. montevidiensis var. montevidiensis, S. montevidiensis var. glandulifera, S. nudicaulis, S. hasleriana and S. dulcis were collected and maintained under greenhouse conditions. The Murashige-Skoog medium, supplemented with 0.25 mg/L BAP was used for the nodal segments multiplication of the five Scoparia species. Except for S. hasleriana, the multiplication rate of the other species ranged between 10 and 12 shoots per explant. The colchicine doses tested with S. montevidiensis were: 0.0; 0.1; 0.05; 0.01 and 0.001% (24 and 48 hrs). From a total of 364 recovered plants, 4 solid tetraploid and 16 chimeras were detected. Significant differences were observed for the size of flower, leaves, and the stem diameter among the tetraploid plants and between them and the control. The tissue culture proved to be a powerful tool both to multiply the Scoparia material incorporated to our germplasm collection and to obtain new improved varieties of this beautiful genus