Cryopreservation and germinative behavior of Passiflora spp. seeds.

Cryopreservation is an interesting strategy for the long-term conservation of the seeds of the majority of cultivated plants as well as many of the corresponding wild parents. The objective of this study was to evaluate the response of the seeds of different passion fruit species to cryopreservation, as well as to apply multivariate tools to better understand the germinative behavior after thawing, to support the application and repeatability of these methods with other Passiflora species. Seeds of P. coccinea, P. edulis, P. gibertii, P. maliformis, P. morifolia, P. setacea, P. suberosa and P. tenuifila were submitted to cryopreservation, and after thawing the following variables were analyzed: emergence percentage (%); mean emergence time (days); mean emergence speed (plantlets.day-1); uncertainty (bit); and synchrony (bit). We applied multivariate analysis of variance (MANOVA) and canonical discriminant analysis (CDA) to the data. The seeds of all the species withstood the freezing well, indicating that cryopreservation is a viable alternative for long-term conservation of Passiflora genetic material. The emergence percentage, average emergence speed and average emergence time were the variables that contributed the most to separation of the species in the first canonical discriminant function (Can1). The seeds of the species studied were found to be tolerant to desiccation because they remained viable even with low water content. The results showed that seeds could be cryopreserved successfully both with and without prior desiccation.

Reproductive barriers in cassava: Factors and implications for genetic improvement.

Cassava breeding is hampered by high flower abortion rates that prevent efficient recombination among promising clones. To better understand the factors causing flower abortion and propose strategies to overcome them, we 1) analyzed the reproductive barriers to intraspecific crossing, 2) evaluated pollen-pistil interactions to maximize hand pollination efficiency, and 3) identified the population structure of elite parental clones. From 2016 to 2018, the abortion and fertilization rates of 5,748 hand crossings involving 91 parents and 157 progenies were estimated. We used 16,300 single nucleotide polymorphism markers to study the parents' population structure via discriminant analysis of principal components, and three clusters were identified. To test for male and female effects, we used a mixed model in which the environment (month and year) was fixed, while female and male (nested to female) were random effects. Regardless of the population structure, significant parental effects were identified for abortion and fertilization rates, suggesting the existence of reproductive barriers among certain cassava clones. Matching ability between cassava parents was significant for pollen grains that adhered to the stigma surface, germinated pollen grains, and the number of fertilized ovules. Non-additive genetic effects were important to the inheritance of these traits. Pollen viability and pollen-pistil interactions in cross- and self-pollination were also investigated to characterize pollen-stigma compatibility. Various events related to pollen tube growth dynamics indicated fertilization abnormalities. These abnormalities included the reticulated deposition of callose in the pollen tube, pollen tube growth cessation in a specific region of the stylet, and low pollen grain germination rate. Generally, pollen viability and stigma receptivity varied depending on the clone and flowering stage and were lost during flowering. This study provides novel insights into cassava reproduction that can assist in practical crossing and maximize the recombination of contrasting clones.

Strategies for vegetative propagation and viral cleaning of a miniature ornamental pineapple hybrid

This study assessed and compared different methods for vegetative propagation of a miniature ornamental pineapple hybrid (ORN-MUT), seeking to determine the best method for production of plantlets, as well as for removal of the PMWaV viral complex from plants cultured in vitro, for production of healthy parent plants. Pineapple wilt is a disease that can cause large economic and is caused by a viral complex called Pineapple mealybug wilt-associated virus (PMWaV). For this, four propagation methods were evaluated (conventional, stem sectioning, micropropagation and etiolation of nodal segments). The time necessary for each method and the number of plants formed were assessed. Stem tips (0.5 mm) were cultured and indexed for three PMWaV types. Conventional propagation produced 17 plantlets per plant in 566 days, stem sectioning produced 2.3 plantlets per stem in 591 days, while the conventional micropropagation technique produced 1,284 plants after four subcultures in 778 days. Stems etiolated for 60 days showed peak production in the second subculture, with 1,224 plants. This method required 883 days to obtain plants with ideal size for transplantation to the field. In turn, stems etiolated for 120 days produced 935 plants at the end of four subcultures, with peak output in the third subculture, in which the plants could be cultivated in the field after 943 days. Conventional micropropagation and etiolation for 60 days were the best methods for production of plantlets of the ORN-MUT hybrid. The results of this work showed that the cultivation of shoot tips is an efficient strategy to remove the PMWaV complex and obtain healthy mother plants and can be a useful tool for other varieties of pineapple.

Assessment of in vitro anthelmintic activity and bio-guided chemical analysis of BRS Boyrá pineapple leaf extracts.

Species of the Bromeliaceae are known for their pharmacological actions, including anthelmintic effects. The aim of this study was to investigate the in vitro anthelmintic activity of extracts and fractions of BRS Boyrá pineapple leaf against the eggs and infective larvae of gastrointestinal nematodes (Trichostrongylidae) of goats and to identify the compounds involved in this activity. Crude methanol, hexane, dichloromethane, ethyl acetate and residual hydromethanol extracts were investigated by quantitative analysis of phenolic and flavonoid contents, antioxidant activity, anthelmintic activity against gastrointestinal nematodes of goats. The extracts were submitted to chromatographic methods for substance isolation and spectrometric techniques to identify their structures. The anthelmintic activity was performed by in vitro assays with eggs and larvae of nematodes obtained from naturally infected donor goats. All extracts contained phenolic (2.22-14.12 g of gallic acid equivalent per 100 g of dry extract) and flavonoid compounds (0.13-1.45 g of quercetin equivalent per 100 g of dry extract). Bio-guided fractionation of the BRS Boyrá pineapple leaves showed high antioxidant activity (EC50 for DPPH of 2.16-21.38 mg mL-1 and inhibition of co-oxidation of ß-carotene of 36.40-74.86%) and anthelmintic activity (15.69-100% inhibition of egg hatching). The ethyl acetate extract exhibited greatest activity in all assays. Through chromatographic column analysis it was possible to isolate three substances: ß-sitosterol and stigmasterol mixture in dichloromethane and hexane extracts, identified by NMR and p-coumaric acid in the ethyl acetate extract, identified by HPLC-DAD. The isolated p-coumaric acid exhibited high ovicidal effect against goat gastrointestinal nematodes (IC50: 0.12 mg mL-1) and can be considered the active substance of the ethyl acetate extract. This study revealed for the first time that the pineapple BRS Boyrá possesses inhibitory activity against gastrointestinal nematodes (Haemonchus spp., Oesophagostomum spp. and Trichostrongylus spp.), and that p-coumaric acid is an important bioactive.

Micropropagation and in vitroconservation of Alcantarea nahoumii (Bromeliaceae), an endemic and endangered species of the Brazilian Atlantic Forest

Alcantarea nahoumii(Leme) J. R. Grant is a species native to theAtlantic Forest that stands out for ornamental purposes. The objective of this work was to evaluate the in vitrogerminationof A. nahoumiiseeds and establish a micropropagation protocol for production of seedlings so as to minimize the effects of predatory extractivism and develop an in vitroconservation system. Mature seeds were disinfested, established in three culture media (MS, MS½ and MS⅓) and incubated at four temperatures (20, 25, 30 and 35ºC)in a germination chamber. In the micropropagation experiment, stem segments were introduced in MS medium supplemented with 0.5 μM of 1-naphthaleneacetic acid (NAA) and 0.0, 2.2, 4.4 and 6.6 μM of 6-benzylaminopurine (BAP). For the in vitroconservation, plantlets were established in MS or MS½ medium supplemented with 15 g L-1or 30 g L-1of sucrose. The plants were acclimated with commercial substrate. The highest seed germination percentages were promoted by temperature conditions of 20 and 25ºC, with MS culture medium. The highest multiplication rate of shoots was obtained from the treatment without addition of the growth regulator or when combined with 2.2 μM of BAP + 0.5 μM of NAA. The acclimation of the plants occurred with high survival rate. The species can be conserved in vitrounder slow growth condition for 24 months when incubated in MS medium supplemented with 30 g L-1of sucrose.

Evaluation of the micropropagation potential of curauá pineapple hybrids for fiber production / Avaliação do potencial de micropropagação de híbridos do abacaxi curauá para a produção de fibras

Plant fiber is a renewable and biodegradable material that can be used effectively to reinforce various composites. Pineapple hybrids selected for their fiber quality are in the phase of agronomic validation in Brazil by the Embrapa Cassava and Fruits research unit. The selection of a hybrid for large-scale fiber production depends on obtaining a large number of seedlings. This study evaluated the morphogenetic response and propagation potential of eight hybrids of Ananas comosus var. erectifolius, for the purpose of producing high-quality seedlings on a large scale. Stem and crown buds were reduced and placed in MS nutritive medium supplemented with BAP at 0.5 mg L-1, NAA at 0.01 mg L-1 and Phytagel® at 2.5 g L-1. After 45 days, the number of oxidized, contaminated and surviving buds was determined. Swollen buds and plantlets were transferred to a multiplication medium containing MS sucrose, salts and vitamins. The propagation potential was evaluated based on the geometric growth rate among sub-cultures. The FIB-NEG hybrid presented the best results for the establishment phase (40.28%). The best propagative potential was obtained from crown buds with the highest values for FIB-EST (3.93), FIB-MIN (3.91) and FIB-BOY (3.91) hybrids.

A fibra vegetal é uma fonte renovável, biodegradável e de excelente desempenho como reforço em compósitos variados. Híbridos selecionados pela qualidade de suas fibras estão em fase de validação agronômica na Embrapa Mandioca e Fruticultura e sua adoção para produção de fibra em larga escala depende de um elevado número de mudas. Este trabalho teve como objetivo avaliar a resposta morfogenética e o potencial propagativo de oito híbridos de Ananas comosus var. erectifolius, com a finalidade de produzir mudas de qualidade em larga escala. Gemas do caule e coroa foram reduzidas, introduzidas em meio nutritivo MS suplementado com BAP a 0,5 mg L-1, ANA a 0,01 mg L-1 e Phytagel® a 2,5 g L-1. Aos 45 dias foram avaliados o número de gemas oxidadas, contaminadas e sobreviventes. Gemas intumescidas e plantas formadas foram transferidas para o meio de multiplicação contendo sacarose, sais e vitaminas MS. Avaliou-se o potencial propagativo a partir de uma taxa de crescimento geométrico entre subcultivos. O híbrido FIB-NEG (40.28%) apresentou os melhores resultados em porcentagem para a fase de estabelecimento. O melhor potencial propagativo foi obtido a partir de gemas de coroa, com os valores mais elevados registrados para os híbridos FIB-EST (3.93), FIB-MIN (3.91) e FIB-BOY (3.91).

In vivo fertilization of banana / Fertilização in vivo de bananeira

The aim of this research was to study the in vivo fertilization process of banana cultivars. The diploid hybrid (AA) 091087-01 was the male progenitor. Flower samples were checked for fertilization from the first to the twentieth day after pollination. The size of the diploid ovules increased gradually at the beginning of the seed formation process. On the other hand, in the AAA triploids (Cavendish subgroup), the not fertilized ovules were aborted. In the AAB triploids (Prata subgroup) some ovules were fertilized. The flowers of Grand Naine, Nanicão and 'Pacovan' cultivars presented necrosis in the distal part of the ovary on the first day after pollination. Necrosis can hinder pollen tube growth towards the ovule, which might be related to the low seed yield in 'Pacovan' cultivars and to the absence of seeds in the Cavendish subgroup cultivars.

O objetivo do trabalho foi investigar o processo de fertilização in vivo em cultivares de bananeira. Utilizou-se como genitor masculino o híbrido diploide (AA) 091087-01. Do primeiro até o vigésimo dia após a polinização, foram retiradas amostras de flores para comprovação da fertilização. Verificou-se que os óvulos de diploides aumentaram de tamanho gradualmente, iniciando o processo de formação de sementes, enquanto, nos triploides AAA (subgrupo Cavendish), ocorreu o aborto dos óvulos não fertilizados. Nos triploides AAB (subgrupo Prata), alguns óvulos são fertilizados. As flores das cultivares 'Grande Naine', 'Nanicão' e 'Pacovan' apresentaram uma necrose na região distal do ovário, detectada desde o primeiro dia após a polinização, a qual pode se constituir em uma barreira para o crescimento do tubo polínico em direção ao óvulo, o que provavelmente pode estar relacionado à baixa produção de sementes em Pacovan e à ausência de sementes em cultivares do subgrupo Cavendish.

Morfologia e viabilidade de grãos de pólen de acessos silvestres de abacaxi / Morphology and viability of pollen grains from wild varieties of pineapple

O objetivo do trabalho foi avaliar a viabilidade de grãos de pólen de acessos silvestres de abacaxi mediante a percentagem de germinação in vitro, crescimento do tubo polínico em duas temperaturas de incubação e uso de carmim acético. Além disso, o trabalho busca reconhecer os padrões da morfologia polínica de algumas variedades de abacaxi. Foram utilizadas amostras de pólen coletadas de seis acessos de abacaxi: Silvestre-25, I-26/803 (Ananas macrodontes), Igor (A. comosus var. erectifolius), Ananás Santo Amaro, FRF-22 e FRF-32 (A. comosus var. bracteatus). Os grãos de pólen foram inoculados em meio de cultura contendo 10 por cento de sacarose, 0,01 por cento H3BO3, 0,01 por cento KNO3, 0,03 por cento Ca(NO3)2.4H2O, solidificado com 1 por cento de ágar e pH ajustado para 6,5. Foram testadas as temperaturas de incubação de 25°C e 30°C. A viabilidade do pólen foi avaliada pela coloração com carmim acético a 2 por cento. Todas as variedades de A. comosus apresentaram grãos de pólen de tamanho médio, simetria radial, âmbito ovalado a esferoidal, 2-porados, subprolatos, exina eutectada, reticulada, heterobrocada, lumens variando de arredondados a poligonais, sem granulações, com exceção dos dois acessos de A. macrodontes que apresentaram tamanho grande. Melhores taxas de germinação foram obtidas para A. macrodontes em comparação com o erectifolius e bracteatus. A maioria dos acessos apresentou viabilidade com carmim acético acima de 76 por cento.

The aim of this research was to evaluate the pollen grains viability from accessions of wild pineapple using in vitro pollen germination and the growth of the polinic tube in two different temperatures and acetic carmine. In addition, the study aimed to recognize the pattern of pollen morphology of some pineapple varieties. Samples from six pineapple's accessions: Silvestre-25, I-26/803 (Ananas macrodontes), Igor (A. comosus var. erectifolius), Ananás Santo Amaro, FRF-22 and FRF-32 (A. comosus var. bracteatus) were used. The pollen grains were inoculated onto medium culture containing 10 percent of sucrose, 0.01 percent H3BO3, 0.01 percent KNO3, 0.03 percent Ca(NO3)2.4H2O, solidified with 1 percent agar and pH adjusted to 6.5. Two incubation temperatures, 25°C e 30°C were evaluated. Pollen viability was evaluated by staining with 2 percent acetic carmine. All varieties of A. comosus showed pollen grains of medium size, radial symmetry, oval to spheroidal, ambit 2-porate, subprolate, exine eutectic, reticulate, heterobrochate, lumens ranging from polygonal to rounded, grain-free, except for two accessions of A. macrodontes that showed big size. The best germination rates were obtained for A. macrodontes when compared to erectifolius and bracteatus. The majority of accessions presented pollen viability higher than 76 percent using the acetic carmine technique.