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AGRARIAN SCIENCES • An. Acad. Bras. Ciênc. 92 (suppl 1) • 2020 • https://doi.org/10.1590/0001-3765202020180062   copy

Compatibility in pollen-pistil interaction of interspecific crossings with Passiflora spp.

Authorship SCIMAGO INSTITUTIONS RANKINGS

Abstract

Intraspecies or interspecies crossings transfer relevant alleles between plants. However, some interspecies crossings involving Passiflora species impede ovule fertilization and the viable development of seeds. Thus, the purpose of this study was to verify the viability of interspecific crossings and monitor pollen tube development. The experiment had six species of Passiflora in the reciprocal crossings. Histochemical tests aimed to evaluate the percentage of intraspecies or interspecies crossings that resulted in fruit development and pollen tube development. Ovule fertilization and fruit development occurred in determined directions of crossings when controlling the female or male genitor, but only one case of reciprocal crossing had success. In crossings with no fruit development, histological analysis showed that some callus developed in the stigma and style, confirming unilateral and interspecies incompatibility in the genus Passiflora to some species and some directions of crossings.

Key words
interspecific hybridization; incompatibility; plant crossings; genetic breedings

INTRODUCTION

The contribution of the genus Passiflora for plant diversity outstands in the family Passifloraceae with an expressive number of species. Only in Brazil, there are more than 135 species, from which there are 81 species and eight taxonomic varieties that are endemic in tropical and subtropical areas (Cervi & Imig 2013CERVI AC & IMIG DC. 2013. A new species of Passiflora (Passifloraceae) from Mato Grosso do Sul, Brazil. Phytotaxa 103: 46-50., Bernacci et al. 2013BERNACCI LC, CERVI AC, MILWARD DE AZEVEDO MA, NUNES TS, IMIG DC & MEZZONATO AC. 2013. Passifloraceae In: Lista de Espécies da Flora do Brasil. Jardim Botânico do Rio de Janeiro.). In Brazil, Passiflora species have an economical relevance to both food production and medical drugs as well as some have the potential to be ornamental plants (Cervi et al. 2013CERVI AC, MILWARD DE AZEVEDO MA & BERNACCI LC. 2013. Passifloraceae. In: Lista de Espécies da Flora do Brasil, Jardim Botânico do Rio de Janeiro.). The ornamental potential of plants is still in process to be explored (Santos et al. 2012SANTOS EA, SOUZA MM, ABREU PP, CONCEIÇÃO LDHCS, ARAÚJO IS, VIANA AP, ALMEIDA AAF & FREITAS JCO. 2012. Confirmation and characterization of interspecific hybrids of Passiflora L. (Passifloraceae) for ornamental use. Euphytica 184: 389-399.).

In addition, it has a high potential for the production of new varieties of Passiflora by the formation of hybrid plants with abundant size and flowering varieties (Abreu et al. 2009ABREU PP, SOUZA MM, SANTOS EA, PIRES MV, PIRES MM & ALMEIDA AF. 2009. Passion flower hybrids and their use in the ornamental plant market:perspectives for sustainable development with emphasis on Brazil. Euphytica 166: 307-315.). With the aim of producing new varieties, some breeding programs can use wild Passiflora species in crossings, since they have genes that promote adaptation to several natural areas (Debouck & Libreros 1995DEBOUCK DG & LIBREROS D. 1995. Neotropical montane forests: A fragile home of genetic resources of wild relatives of New World crops. In: Churchill SP, Balslev H, Forero E and Luteyn JL (Eds). Biodiversity and conservation of Neotropical montane forests, proceedings of a symposium, New York Botanical Garden 1993, p. 561-577., Bruckner 1997BRUCKNER CH. 1997. Perspectivas do melhoramento do maracujazeiro. In:Maracujá temas selecionados. Porto Alegre: Cinco Continentes, 70 p.).

Breeding programs have the purpose to create a new variety using the germplasm of individual species from populations or genotypes. It is a challenge to generate a new variety of cultivar with the maximum possible number of favorable alleles in only one individual, with the alleles originating from different parental genotypes. In order to develop new cultivars, most plant breeding programs use a set of genotypes for developing favorable genetic combinations (Peel & Rasmusson 2000PEEL MD & RASMUSSON DC. 2000. Improvement strategy for mature plant breeding programs. Crop Science (CS), Madison 40(5): 1241-1246., Reddy & Comstock 1970REDDY BVS & COMSTOCK RS. 1970. Simulation of the backcross method: I. Effects of heritability and gene number on fixation of desired alleles. Crop Science (CS), Madison 16(6): 825-830.).

The majority of results for interspecific crossings in a F1 generation contains intermediate phenotypes for some traits such as: stipule size, morphology and position of nectaries; variation on series number, structure and coloring of corona filaments on flowers and variation on lobules number and leaf formats (Ulmer & Macdougal 2004ULMER T & MACDOUGAL JM. 2004. Passiflora: passion flowers of the world. Timber Press Portland, Oregon.). When the purposes are to transfer favorable traits from native species to commercial species of Passiflora edulis Sims., such as genes of resistance to premature death and nematodes tolerance, more than 500 hybrids was obtained in Passiflora (Ataíde et al. 2012ATAÍDE EM, OLIVEIRA JC & RUGGIERO C. 2012. Florescimento e frutificação do maracujazeiro silvestre Passiflora setacea DC. Rev Bras Frutic 34: 377-381., Fischer 2004FISCHER R. 2004. Hybrids and hybridization. In: Ulmer T and MacDougal JM (Eds) Passiflora: passion flowers of the world. Timber Press Portland, Oregon, p. 362-376.). However, interspecific crossings had other purposes such as coloring matches on flowers and cold tolerance between native species (Bugallo et al. 2011BUGALLO V, CARDONE S, JULIA M & GABRIELA P. 2011. Breeding advances in Passiflora spp. (Passionflower) native to Argentina. Floriculture and Ornamental Biotechnology 5(1): 23-34.).

However, sometimes interspecific crossings can present incompatibility barriers that impede ovule fertilization and, therefore, it is impossible to a F1 generation with hybrid plants to develop. There is a need to know about reproductive systems and chromosomal homology between species that will be used in crossings (Meletti et al. 2005MELETTI LMM, SOARES-SCOTT MD, BERNACCI LC & PASSOS IR. 2005. Melhoramento genético do maracujá: passado e futuro. In: Faleiro FG, Junqueira NTV and Braga MF (Org). Maracujá: germoplasma e melhoramento genético. Planaltina: EMBRAPA CERRADOS 1: 55-78., Santos et al. 2012SANTOS EA, SOUZA MM, ABREU PP, CONCEIÇÃO LDHCS, ARAÚJO IS, VIANA AP, ALMEIDA AAF & FREITAS JCO. 2012. Confirmation and characterization of interspecific hybrids of Passiflora L. (Passifloraceae) for ornamental use. Euphytica 184: 389-399.). In the literature, there are some studies related to reproductive biology, nevertheless their focus are on morphology, flowering biology and agents of pollination. There is also a lack in study approaches about reproductive systems, pollination efficiency and pollen viability (Amorim et al. 2011AMORIM JS, SOUZA MM, VIANA AJC & FREITAS JCO. 2011. Self-, cross- and interspecific pollinations in Passiflora capsularis and P. rubra. Rev Bras Bot 34: 537-544., Shivanna 2012SHIVANNA KR. 2012. Reproductive assurance through unusual autogamy in theabsence of pollinators in Passiflora edulis (passion fruit). Curr Sci 103: 1091-1096.).

Researches on pollen viability and in vivo fertilization are fundamental to studies with the purpose of reproductive biology and development of hybrid plants, since they guide the researcher to plan the crossings, which can evidently favor the identification of promising cross-breeding. Strategies for plans and crossings improve the experiment with reduction in time for work execution. Therefore, the purpose of this study was to evaluate the viability of interspecific reproduction among six species of Passiflora, so that it is possible to collect information about the potential use of species in crossings and their future use in breeding programs.

MATERIALS AND METHODS

The experiment was conducted from April 2014 to July 2015 in the experimental field from the Department of Plant Breeding of Universidade Federal de Viçosa (UFV), localized in the municipality of Viçosa, Minas Gerais, Brazil (20°45’52.3”S 42°51’09.5”W). The city has 680 meters of altitude, a climate predominantly of tropical altitude, dry and cold winter, hot and humid summer and an average of annual rain precipitation of 1200 mm.

The species used in this study were: Passiflora cincinnata Mast.; Passiflora gibertii N. E. Brown; Passiflora mucronata Lam.; Passiflora alata Curtis; and Passiflora edulis Sims. The plants were established in vineyards, drip irrigated and subjected to pruning, fertilization and spraying. The laboratory analyzes were performed at the Laboratory of Plant Anatomy of the Department of Plant Biology.

The crossings from the experiment had seven plant genotypes for each species that were randomly chosen in the field. Species phenology was observed to make viable crossings under the flowering coincidence. The flower-bud in pre-anthesis used in the breedings were filled into a paper bag on the day before anthesis, in order to avoid exogenous pollen contamination, with the exception of P. edulis, that had their flower-bud filled into paper bags in the morning of the breeding day.

Controlled crossings followed these steps: the female genitors were unsacked and emasculated, meanwhile the pollen was collected from the male genitor with the use of swabs and transferred to the stigma of the emasculated flower. Hybridizations under a reciprocal mode in each pair of plants had five flowers being used in each combination of crosses.

The verification of crossing success occurred after seven days, by the evaluation of fruit formation. When haversting fruits, they were bagged with a nylon net until their complete ripening. Then, seeds were extracted from fruits, dried at room temperature and maintained in paper bags in a refrigerator. The interpretation of data was performed by descriptive statistics, with the observation of the percentage of crossings between plants within each species and between species.

In order to verify the formation of the pollen tube, a histochemical test was performed, with the evaluation of ten flowers per plant that were recipients of pollen in reciprocal crossings. In a period of 24 hours after pollination, all flowers were collected and submitted to a fixation solution composed by Formaline at 37%, Acetic acid and Alcohol at 70% (FAA) in the proportion of 5:5:90 for 48 hours. Then, pollen tubes were maintained in Alcohol at 70% until slide preparation for analysis. Slide preparation consisted in soften biological material with sodium hydroxide (NaOH) at 10% and clearing with sodium hypochlorite at 1% and, afterwards, biological material on the slide was stained with Aniline Blue. Then, the laminule was positioned in the slide and a pressure was applied on the stigma making possible the subsequent examination of the biological material under Ultraviolet Light Microscope, in the blue hue. Images were captured in the Olympus Optical AX70TRF Microscope, with a digital camera connected to it (Spot Insightcolour 3.2.0, Diagnostic Instruments Inc., USA), in the Plant Anatomy Laboratory at UFV.

RESULTS

Breedings between Passiflora gibertii N. E. Brown and Passiflora mucronata Lam.

There were 5% of fruit production, when the female genitor was P. gibertii and the male genitor was P. mucronata. On the other hand, there was no fruit development in the reciprocal crossing (Table I). In some cases, the genotypes in the crossings influenced fruit development. Four out of seven genotypes of P. gibertii were female genotypes and had fruit production ranging from 3% to 17%. Seven genotypes of P. mucronata were pollen donors in the experiment, but only five genotypes fertilized the ovules of P. gibertii. At these crossings, with P. mucronata as the male genitor and P. gibertii as female genitor, there was no visualization of the pollen tube development by histochemical analysis. The average number of seeds per fruit from viable crossings was of 27.18 seeds per fruit, considering that P. gibertii plants produce small fruits (Table II).

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Table I
Percentage of fruit development in reciprocal crossings between Passiflora gibertii N. E. Brown and Passiflora mucronata Lam. UFV, Viçosa-MG, 2016.
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Table II
Average of seeds per fruit, obtained in open and controlled crosses with Passiflora spp. UFV, Viçosa-MG, 2016.

Crossings between Passiflora alata Curtis and Passiflora mucronata Lam.

In the crossings between Passiflora alata as the female genitor and Passiflora mucronata as male genitor, the percentage of fruit production was of 1%. On the other hand, no fruit production occurred in the reciprocal crossing (Table III). Among the seven genotypes of P. alata used as female genitor in the crossings, only two cases of crossings had fruit development, which corresponds to 3%. On the other hand, from the genotypes of P. mucronata used as pollen donors, only one genotype fertilized the ovule of the female genitor P. alata and produced fruits (Table III). No cellular structures indicated pollen grains germination in the histochemical evaluation. Finally, the crossings with fruit production had an average of 28 seeds per fruit (Table II).

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Table III
Percentage of fruit production in reciprocal crossings between Passiflora alata Curtis and Passiflora mucronata Lam. UFV, Viçosa - MG, 2016.

Crossings between Passiflora edulis Sims. and Passiflora mucronata Lam.

The crossing with P. edulis as the female genitor and P. mucronata as male genitor had an average of 1% of fruit production. However, the crossing with P. mucronata as the female genitor had no fruit. In the crossings with fruit production, one out of seven genotypes of P. edulis used as female genitor was responsible for 6% of fruit development. Similarly, P. mucronata as the male genitor (pollen donor) had only one out of seven genotypes being responsible for 6% of fruit development (Table IV).

No cellular structures indicated pollen grains germination in the histochemical analysis. The crossings between the female genitor P. edulis and the male genitor P. mucronata produced 44 seeds at total, which is a small value in relation to crossings with P. edulis that had an average of 330 seeds per fruit. It is worth to mention that most of the 44 seeds resulted from the crossings between the female genitor P. edulis and the male genitor P. mucronata (Table II).

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Table IV
Percentage of fruit production in reciprocal crossings between Passiflora edulis Sims. and Passiflora mucronata Lam. UFV, Viçosa-MG, 2016.

Crossings between Passiflora gibertii N.E. Brown and Passiflora alata Curtis

In crossings with P. gibertii as the female genitor and P. alata as male genitor, there was 1% of fruit production and in the reciprocal crossing there was 0.41% of fruit production. Additionally, only one out of seven genotypes of P. gibertti showed to be responsive to crossings, when it was female and male genitors. A similar behavior occurred with P. alata (Table V). Additionally, these crossings had no visualization of the pollen tube by histochemical analysis. The average number of seeds per fruit was of nine seeds, which is a lower value than the expected value, since P. gibertii had an average of 25 seeds per fruit (Table II), as mentioned before.

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Table V
Percentage of fruit production in reciprocal crossings between Passiflora gibertii N. E. Brown and Passiflora alata Curtis. UFV, Viçosa- MG, 2016.

Crossings between Passiflora gibertii N.E. Brown and Passiflora cincinnata Mast.

There were 9% of fruit production, when P. gibertii was the female genitor and P. cincinnata Mast. was the male genitor. The reciprocal crossing had no fruit. In the direction of the crossing with fruit production, six out of the seven genotypes of P. gibertii used as female genitor had fruit development varying from 3% to 20%. Among P. cincinnata genotypes used as the male genitor only four produced fruits, ranging from 6% to 20% of fruit development. In crossings with fruit production, P. gibertii had an average of 17.10 seeds per fruit (Table VI).

In the crossings with fruit development, histochemical analysis of stigmas can confirm pollen germination under the stigma surface and pollen tube formation. On top of the developed callus in the stigma occurred the development of pollen tube (Figure 1a). Fruit development occurs after egg fertilization, but histochemical analysis confirms that not all the pollen tubes formed can reach the ovary of the flower. In the reciprocal crossings, germination was evidenced in pollen grains (Figure 1b), with occurrence of the same phenomenon of callus deposition in the pollen tube, with no fruit development.

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Table VI
Percentage of fruit production in reciprocal crossings between Passiflora gibertii N. E. Brown and Passiflora cincinnata Mast. UFV, Viçosa – MG, 2016.
Figure 1
Fluorescence microscopy in pistil of P. gibertti pollination with incompatible pollen. The pistils were stained with Aniline Blue. a) P. gibertii and P. cincinnata crossing. Bar = 150 µm; arrows indicate callus; b) P. cincinnata and P. gibertii crossing. Pollen grains germinating under the stigma and tubes with posterior callus development during pollen tube formation; star indicates pollen grain germination. Bar = 300 µm.

Crossings between Passiflora alata and Passiflora cincinnata

The crossings with the female genitor P. alata and the male genitor P. cincinnata had 0.41% of fruit production. The reciprocal crossing had no fruit development (Table VII). Among the genotypes of P. alata, only one had 3% of fruit development. This result also occurred with the male genitor P. cincinnata. Crossings between P. alata as the female genitor and P. cincinnata as the male genitor had an average of 122 seeds per fruit, which is a median value, in relation to crossings with P. alata that presented an average of 191 seeds (Table II). Histochemical analysis of stigmas in the direction of fertilization showed a low percentage of fruit development due to the development of callus that blocked the development of the pollen tube (Figure 2).

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Table VII
Percentage of fruit production in reciprocal crossings between Passiflora alata and Passiflora cincinnata Mast. UFV, Viçosa – MG, 2016.
Figure 2
Fluorescence microscope of pistils in the crossing between P. alata and P. cincinnata, stained with Aniline Blue. a) Deformed pollen grain by callus excessive deposition is still inhibited on the stigma; b, c) Top of pollen tubes after going through the stigma, being blocked by callus synthesis after 24 hours of pollination. Bar = 150 µm respectively.

Crossings between Passiflora edulis Sims. and Passiflora cincinnata Mast.

In crossings between the female genitor Passiflora edulis Sims. and male genitor Passiflora cincinnata Mast., there were 5% of fruit production. On the other hand, the reciprocal crossing had 0.41% of fruit development.

Six plants of P. edulis were used in the crossings, resulted in well-succeeded fruit production, ranging from 3% to 14% of fruit development. Five plants of the male genitor P. cincinnata had 3% to 20% of fruit production (Table VIII).

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Table VIII
Percentage of pollinations with fruit production in reciprocal crossings between Passiflora edulis Sims. and Passiflora cincinnata Mast. UFV, Viçosa - MG, 2016.

In the reciprocal crossing with the male genitor P. edulis, only one genotype had 3% of fruit production. On the other hand, the reciprocal crossing with the female genitor P. cincinnata had only one out of seven genotypes with 3% of fruit production. On these crossings, there was no visualization of the pollen tube by histochemical analysis. In crossings between the female genitor P. edulis and the male genitor P. cincinnata, the average number of seeds per fruit was of 180.12 seeds being an approximate value to the species that is 330 seeds (Table II).

Crossings between Passiflora gibertii N.E. Brown and Passiflora edulis Sims.

The crossings with the female genitor P. gibertii and the male genitor P. edulis had 0.41% of fruit development. The reciprocal crossing had no fruit development (Table IX). In the crossing with fruit development, the female genitor P. gibertii had one out of the seven genotypes with approximately 3% of fruit development. Similarly, the male genitor P. edulis had only one genotype being responsible for 3% of fruit production.

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Table IX
Percentage of fruit development in reciprocal crossings between Passiflora gibertii N. E. Brown and Passiflora edulis Sims. UFV, Viçosa – MG, 2016.

On these breedings, there was no visualization of the pollen tube by histochemical analysis. Crossings with the female genitor P. gibertii and the male genitor P. edulis had a production of nine seeds per fruit, which is a relatively low value, when compared to the expected value of 25 seeds (Table II).

Crossings between Passiflora alata Curtis and Passiflora edulis Sims.

The crossing with the female genitor P. alata and the male genitor P. edulis had 3% of fruit development, but the reciprocal crossing had no fruit development (Table X). In crossings with fruit production, there are three genotypes of P. alata with fruit development, ranging from 6% to 11%. From P. edulis genotypes, only one genotype was a pollen donor and had 23% of fruit production (Table X). Histochemical analysis did not show any pollen tube formation. And, finally, the average number of seeds per fruit was of 120 seeds in crossings with fruit production (Table II).

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Table X
Percentage of pollination with fruit production in reciprocal crossings between Passiflora alata Curtis and Passiflora edulis Sims. UFV, Viçosa - MG, 2016.

DISCUSSION

It was observed a difference in the rate of crossings in hybridizations between plants within each species. It reinforces the idea that the success of hybridizations does not only depend on the species chosen, but also on the plants selected.

The compatibility in the crosses between the species evaluated in this work was only possible due to the phylogenetic proximity between the species, although in some cases compatibility occurred only in one direction of the crossings. The species are included in the same subgenus Dysosmia and have the same chromosome number (2n=18) in P. cincinnata (Guerra 1986GUERRA MS. 1986. Citogenética de angiospermas coletadas em Pernambuco. Rev Bras Genét 9: 21-40.), P. alata (Meletti et al. 2003MELETTI LMM, BERNACCI LC, SOARES-SCOTT MD, AZEVEDO FILHO JA & MARTINS ALM. 2003. Variabilidade genética em caracteres morfológicos, agronômicos e citogenéticos de populações de maracujazeiro-doce (Passiflora alata Curtis). Rev Bras Frutic 25: 275-278.), P. mucronata (Souza et al. 2003SOUZA MM, PEREIRA TNS, SILVA LC, REIS DSS & SUDRÉ, CP. 2003. Karyotype of six Passiflora species in the State of Rio de Janeiro. Cytologia 68: 165-171.), P. gibertii (Mayeda & Vieira 1995MAYEDA LY & VIEIRA MLC. 1995. Estudo cariotípico de três espécies do gênero Passiflora (Passifloraceae). Genet Mol Biol 18: 426.) and P. edulis (Soares-Scott et al. 2003SOARES-SCOTT MD, MELETTI LMM & RECCO-PIMENTEL SM. 2003. Meiotic behavior and pollen fertility in sexual and somatic hybrids of Passiflora species. Caryologi 56: 129-138.).

Similar results were found by Soares et. al. (2015). The authors describe about the experiment success, when using the following species with the same chromosome number (2n=18) in the crossings: P. racemosa, P. gibertii, P. edmundoi, P. mucronata, P. edulis f. flavicarpa, P. galbana and P. tenuifila. Souza et al. (2008)SOUZA MM, PEREIRA TNS & VIEIRA MLC. 2008. Cytogenetic studies in some species of Passiflora L. (Passifloraceae): a review emphasizing Brazilian species. Braz Arch Biol Technol 51: 247-258. describe the positive results in interspecies crossings with the same chromosome number (2n=18) in the following crossings: between P. edulis f. flavicarpa Deg. and P. setacea; P. coccinea and P. glandulosa Cav. According to Pereira et al. (2005)PEREIRA TNA, NICOLI RG, MADUREIRA HC, JÚNIOR PCD, GABURRO NOP & COUTINHO K. 2005. Caracterização morfológica e reprodutiva de espécies silvestres do gênero Passiflora. In: Faleiro FG, Junqueira NTV, Braga MF, Pinto ACQ and Sousa ES (Eds). In: IV Reunião Técnica de Pesquisas em Maracujazeiro. Planaltina, DF: Embrapa Cerrados, p. 29-34. the expected chromosomal homology between genetically similar species can induce viable interspecific crossings and, then, reduce the incongruity, allowing the development of hybrid plants.

It is interesting to note that in the present study some reciprocal crossings were not successful and had no fruits. This phenomenon is referred in the literature as incongruity or unilateral incompatibility. Although there are some exceptions of successful crossings such as the hybridizations between Passiflora gibertii and Passiflora alata, and also between Passiflora edulis and Passiflora cincinnata, which had the development of fruits in reciprocal crossings. In these cases, reciprocal crossings had differences in results with no fruit production in some of them.

This unilateral incongruity occurs with a certain stability in the genus Passiflora, as already mentioned by Bugallo et al. (2011)BUGALLO V, CARDONE S, JULIA M & GABRIELA P. 2011. Breeding advances in Passiflora spp. (Passionflower) native to Argentina. Floriculture and Ornamental Biotechnology 5(1): 23-34.. These authors performed interspecific hybridizations with P. alata, P. caerulea, P. amethystina, P. edulis and P. violacea. They discovered that P. alata and P. caerulea had success in crosses in both directions, though the other interspecific combinations in crossings had one unilateral incongruity.

Soares et al. (2015)SOARES TL, JESUS ON, SOUZA EH & OLIVEIRA EJ. 2015. Reproductive biology and pollen–pistil interactions in Passifloraspecies with ornamental potential. Scientia Horticulturae 197: 339-349. verified that in the reciprocal crossing between P. racemosa and P. capsularis, fruit development only occurred in the crossing with P. racemosa as the male genitor. Junqueira et al. (2005)JUNQUEIRA NTV, BRAGA MF, FALEIRO FG, PEIXOTO JR & BERNACCI LC. 2005. Potencial de espécies silvestres de maracujazeiro como fonte de resistência a doenças. In: Faleiro FG, Junqueira NTV and Braga MF (Eds). Maracujá: germoplasma e melhoramento genético. Planaltina, Embrapa Cerrados, p. 1-106. verified that P. setacea, P. coccinea and P. glandulosa are responsive in the crossings in which they were the female genitors; Also, Conceição et al. (2011)CONCEIÇÃO LDHCS, BELO GO, SOUZA MM, SANTOS SF, CERQUEIRA-SILVA CBM & CORRÊA RX. 2011. Confirmation of cross-fertilization using molecular markers in ornamental passion flower hybrids. Genet Mol Res 10: 47-52. described that the reciprocal crossings between P. watsoniana x P. gardneri and between P. gardneri x P. gibertii had unilateral incompatibility. Additionally, Santos (2013)SANTOS EA. 2013. Melhoramento do maracujazeiro azedo (Passiflora edulis Sims.) visando à resistência ao Cowpea aphid borne mosaic vírus. Master Thesis. UENF, Campos dos Goytacazes, RJ. described that the percentage of fruit development doubled when P. edulis was used as the female genitor (100%) or as the male genitor (50%) in crossings with P. setacea. Finally, OCampo et al. (2016)OCAMPO J, ARIAS JC & URREA R. 2016. Interspecific hybridization between cultivated and wild species of genus Passiflora L. Euphytica 209: 395-408. described fruit production ranging from 2.1% to 60% in crossings with P. vitifolia, P. mucronata, P. edulis f. edulis and P. edulis f. flavicarpa as female genitors. On the other hand, no fruit development occurred in the reciprocal crossing, which confirms the occurrence of unilateral and interspecific incompatibility.

Probably such restrictions to fruit development in some crossings can result in some degree of genetic incompatibility or any zygotic barrier in pre or post fertilization. It can have a rejection mechanism in pollen recognition process under the stigma, style or ovary, which proteins block the germination of pollen grains or inhibit/suspend the development and growth of the pollen tube, besides the ratio pollen:ovary, which can influence on the efficacy of the pollen grain to reach the compatible stigma (Bugallo et al. 2011BUGALLO V, CARDONE S, JULIA M & GABRIELA P. 2011. Breeding advances in Passiflora spp. (Passionflower) native to Argentina. Floriculture and Ornamental Biotechnology 5(1): 23-34., Jones et al. 2012JONES R, OUGHAM H, THOMAS H & WAALAND S. 2012. The molecular life of plants. In: Flowering and sexual reproduction. A J Wiley & Sons, Ltd., Publication, Chichester, p. 605- 619., Ferreira et al. 2014FERREIRA NR, FRANKE LB & BLOCHTEIN B. 2014. Pollen–ovule relation in Adesmiatristis and reflections on the seed–ovule ratio by interaction with pollinators intwo vertical strata. An Acad Bras Cienc 86: 1327-1335.). Also the embrionary sac immaturity, ovule degeneration or lack of synchronism between male and female gametes can directly affect fertilization and, thus, fruit development (Guerra et al. 2011GUERRA ME, WUNSCH A, LOPEZ-CORRALES M & RODRIGO J. 2011. Lack of fruit setcaused by ovule degeneration in japanese plum J Am Soc Hortic Sci 136: 10-15.).

However, this mechanism of unilateral inconsistency, even well frequent in Passiflora, it is not exclusive of this genus, which can also be evidenced in other species of plants such as in the genus Capsicum spp. For example, Martins (2014)MARTINS KC. 2014. Cruzabilidade, tamanho do genoma e viabilidade polínica em Capsicum spp. Master’s Thesis. UENF, Campos dos Goytacazes, RJ. described differences in reciprocal crossings with a better fruit production in the crossing between C. annuum var. annuum and C. baccatum, which had nine fruits, with two fruits in the reciprocal crossing. Nascimento et al. (2012)NASCIMENTO NF, RÊGO ER, REGO MM, NASCIMENTO MF & ALVES LI. 2012. Compatibilidade em cruzamentos intra e interespecíficos em pimenteiras ornamentais. Revista Brasileira de Horticultura Ornamental (RBHO) 18: 57-61. had 50% of fruit development in the crossing between C. chinense and C. baccatum, and the reciprocal crossing showed to be unviable.

The histochemical analysis by fluorescence also evidenced the development and inhibition of pollen tubes due to the development of callus. Such response occurs in a fast manner in the stigma surface, limiting the growth of incompatible pollen tubes. Madureira et al. (2012)MADUREIRA HC, PEREIRA TNS, CUNHA M & KLEIN DE. 2012. Histological analysis of pollen-pistil interactions in sour passion fruit plants (Passiflora edulis Sims). Biocell 36(2): 83-90. states that in cases in which crossings are compatible, callus were not visible due to their association to tube elongation.

Madureira et al. (2014)MADUREIRA HC, PEREIRA TNS, CUNHA M, KLEIN DE, OLIVEIRA MVV, MATTOS L & SOUZA FILHO G. 2014. Self-incompatibility in passionfruit: cellular responses in incompatible pollinations. Biologia (Section Botany) 69/5: 674-584. reinforces that besides callus development, in cases of incompatibility, there is an occurrence of cellular disorganization, which deforms the anatomy of the pollen tube. This is an effect of the actine reduction, which is related to the action of the cytoskeleton in the pollen tube making them very thick. This fact can be observed in the images 1 and 2 obtained in this study, with visualization of the pollen tubes with thicker or thinner areas.

According to Payán & Martín (1975)PAYÁN FR & MARTÍN FW. 1975. Barriers to the hybridization of Passiflora species. Euphytica 24: 709-716., Bugallo et al. (2011)BUGALLO V, CARDONE S, JULIA M & GABRIELA P. 2011. Breeding advances in Passiflora spp. (Passionflower) native to Argentina. Floriculture and Ornamental Biotechnology 5(1): 23-34. and Conceição et al. (2011)CONCEIÇÃO LDHCS, SOUZA MM, BELO GO, SANTOS SF & FREIRE JCO. 2011. Hybridization among wild passionflower species. Rev Bras Bot 34(2): 237-240., there is a second barrier to the occurrence of crossings in Passiflora species that is the abortion of seeds, even if it is possible some crossings between species. The degree of similarity between the number and size of parental chromosomes is associated to the maternal effects that normally promote the endosperm growth, meanwhile the parental effect suppress the endosperm development (Kinoshita et al. 2008KINOSHITA T, IKEDA Y & ISHIKAWA R. 2008. Genomic imprinting: a balance between antagonistic roles of parental chromosomes. Semin Cell Dev Biol 19(6): 574-579.). The reason is that fertilization can cause negative interactions between nucleus-cytoplasm, resulting in risks for embryo development, due to failure in endosperm development (Nimura et al. 2003NIMURA M, KATO J, MII M & MORIOKA K. 2003. Unilateral compatibility and genotypic difference in crossability in interspecific hybridization between Dianthus caryophyllus L. and Dianthus japonicus Thunb. Theor Appl Genet 106: 1164-1170.). This is the explanation for non-germination of seeds obtained from the crosses between P. alata and P. edulis; P. gibertii and P. edulis; P edulis and P. cincinnata, since there is embryo death and a posterior barrier to hybridization.

Failure in the embryo development was described by Conceição et al. (2011) in the crossing between P. gardneri and P. cincinnata, which fruits had 100% of seeds with no endosperm. Furthermore, the crossings between P. gibertii and P. kermesina and between P. gibertii and P. alata had fruits with no seed production. Similarly, Junqueira et al. (2005)JUNQUEIRA NTV, BRAGA MF, FALEIRO FG, PEIXOTO JR & BERNACCI LC. 2005. Potencial de espécies silvestres de maracujazeiro como fonte de resistência a doenças. In: Faleiro FG, Junqueira NTV and Braga MF (Eds). Maracujá: germoplasma e melhoramento genético. Planaltina, Embrapa Cerrados, p. 1-106. found fruits with a rare production of seeds in crossings between P. caerulea and P. edulis f. flavicarpa. However, when changing the direction of the crossings using the male genitor P. edulis Sims., fruits from F1 generation had viable seeds.

CONCLUSIONS

Thus, the experiment results described in this paper can be helpful to other researchers as an auxiliary tool during the plan of intra-crossings or inter-crossings, improving the establishment of breeding programs with the purpose to use interspecific hybrid plants. Although further studies are necessary in order to increase knowledge about this rejection mechanism.

The analysis of the crossings performed in the experiment showed the following crossings with the highest potential for reproduction: P. gibertii x P. cincinnata; P. gibertii x P. mucronata, P. gibertii x P. alata, P. gibertii x P. edulis, P. alata x P. cincinnata, P. alata x P. mucronata, P. alata x P. edulis, P. edulis x P. cincinnata, P. edulis x P. mucronata.

ACKNOWLEGMENTS

The authors are grateful to Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) and Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) for the fellowship offered to the lead author during his PhD studies.

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Publication Dates

  • Publication in this collection
    27 July 2020
  • Date of issue
    2020

History

  • Received
    19 Jan 2018
  • Accepted
    1 Aug 2019
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