Recent Advances in Annona Breeding: a Review

Available online at www.ijpab.com Vinay et al Int. J. Pure App. Biosci. 5 (2): 1168-1181 (2017) ISSN: 2320 – 7051 DOI: http://dx.doi.org/10.18782/2320-7051.2722 ISSN: 2320 – 7051 Int. J. Pure App. Biosci. 5 (2): 1168-1181 (2017) Research Article

Recent Advances in Annona Breeding: A Review

Vinay, G. M.*, Sakthivel, T. and Priyanka, H. L. Division of Fruit Crops, Indian Institute of Horticultural Science, Hesaraghatta lake Post, Bengaluru-89 *Corresponding Author E-mail: [email protected] Received: 19.03.2017 | Revised: 30.03.2017 | Accepted: 2.04.2017

ABSTRACT The Annonaceae family comprises about 120 genera and more than 2000 species. The genus Annona is the most economically important one, containing 120 species. The opportunity for breeding improvement in the annona is significant and challenging. There is a lot more species are available but certain inherent constraints are involved like: long juvenility, higher levels of heterozygosity and large area requirement for assessment of hybrids. On the other hand, wide range of diversity and ease of vegetative hybrid propagation are the advantages for the breeders. There are very few man made commercially important hybrids. Its development is mostly based on the selection of clones were made for fruit quality only. Requirements of a good cultivar involve: dwarfness, precocity and profuse bearing, attractive, good sized and quality fruit, absence of physiological disorders, disease and pest resistance and improved shelf life etc. Comprehensive knowledge about the phenology and advanced techniques for hybridization have been quite helpful to overcome the problems like poor eating and keeping quality. The

development of the genetic markers has further reduced the uncertainty in breeding of annona and maintaining the hybrid populations in a better way.

Key words: Annona, Heterozygosity, Breeding, Juvenility.

INTRODUCTION significant40 and economically important fruits The Annonaceae family comprises about 120 in several tropical and sub-tropical countries. genera and more than 2000 species. The genus And also some other species are ramphal Annona is the most economically important (Annona reticulata) soursop (Annona one, containing 120 species19. The relatively muricata) and pond apple (Annona glabra). wider adaptability of the trees and a good The sugar apple species name ‘squamosa’ consumer preference for their fruits make the refers to the knobby appearance of the fruit. commercial cultivation of these species Sugar apple is a small tropical tree originating attractive. Among the edible annonas, in the New World tropics, probably Central cherimoya (Annona cherimola Mill.), sugar America. Sugar apple is the mostly grown apple (A. squamosa L.) and the hybrid annona species. between the two, atemoya, are commercially

Cite this article: Vinay, G.M., Sakthivel, T. and Priyanka, H.L., Recent Advances in Annona Breeding: A Review, Int. J. Pure App. Biosci. 5(2): 1168-1181 (2017). doi: http://dx.doi.org/10.18782/2320-7051.2722

Vinay et al Int. J. Pure App. Biosci. 5 (2): 1168-1181 (2017) ISSN: 2320 – 7051 The fruit is frequently found in village markets spring. The single, protogynous, mild fragrant but has not shown much potential for large flower emerges from the leaf axils, and commercial cultivation due to the small fruit possesses a short peduncle. Flowering occurs size, frequent cracking at maturity and poor once in a year and it starts when the tree is 3 to shelf life. The perishable nature and supply 4 years old. Anthesis starts in the early shortages make marketing localized or air morning and it takes 8h to attain complete shipment essential. Commercial cultivation of opening. The fruit is normally heart-shaped, cherimoya occurs in Chile, Ecuador, conical, oval or somewhat irregular in form Columbia, Australia, Spain and California28, due to irregular pollination. Fruits measure 7.5 However, the cultivation of atemoya is mostly to 12.5 cm in length and weigh from 200 to limited to Australia and Florida12. Another 700 g. The fruit surface is smooth in some allied species bullock’s heart, which possesses varieties; in others, it is covered with small a number of traits that would be desirable in conical protuberances over the carpels. The other edible annonas, is grown on a limited fruit rind is delicate and thin, and is greenish- scale in Australia, Africa, India, Peru and yellow when ripe29. The white, subacid flesh South-East Asia19. has a fragrant, delicate flavor. The fruit has The improvement of annona rather numerous seeds (21 to 41 seeds/fruit), which any crop needs to explore new recombinants are 1.5 to 2.0 cm in length and approximately primarily by means of exploiting the breeding 1.0 cm in width42. methodologies. Diversity or heterogeneity is 2. Sugar apple – Annona squamosa L. the main character desired for breeding either The sugar apple tree is deciduous and much natural or manmade. It is required to have vast smaller than the soursop, reaching a maximum genetic pool to get new combinations of of 6.0 m in height, with many lateral branches. desired nature and developing new hybrids. The stems present lenticels, while the young Breeding has yet to play its role in the shoots are pubescent and the oldest are development of this crop as it has not been smooth. The leaves are oblong-elliptical in effectively manipulated in the distant past. form, measuring 5 to 17 cm in length and 2 to Now, the scientists have developed certain 7 cm in width, with an obtuse or acuminate hybrids of annona. To go for such strategies, a apex35. The flowers measure 2.0 to 2.5 cm in comprehensive knowledge of the different length and are much smaller than soursop annona species, physiology of the reproductive flowers, being similar in size and form to those parts, their breeding behaviour and cytological of cherimoya. Pollen germination is low and information of the crop is needed. This will may influence final fruit set, which varies from eventually help to improve annona production 5.4% to 5.6%51. The fruit is rounded, heart- status. shaped, ovate or conical, 5 to 7.5 cm in Description of the important Annona diameter, 6 to 10 cm in length and weighing species 120 to 330 g. The white, custard-like pulp has 1. Cherimola - Annona cherimola P. Mill. a pleasant sweet-sour flavour. The fruit The name cherimoya derives from the contains 35 to 45 black seeds, each 1.5 to 2.0 Quechua name "chirimuya", which means cm in length and 0.6 to 0.8 cm in width50. "cold seeds"27. It is a small, erect, deciduous 3. Atemoya – Annona atemoya L. tree. It has simple, alternate, leaves are ovate- The tree is highly vigorous, fast-growing, 25 lanceolate to elliptical in shape, 10 to 25 cm to 30 feet tall. The branches typically drooping long, glabrous on the ventral surface and and the lowest touching the ground. The leaves pubescent dorsally, with leaf shedding in the are deciduous, alternate, elliptical, leathery and

Vinay et al Int. J. Pure App. Biosci. 5 (2): 1168-1181 (2017) ISSN: 2320 – 7051 less hairy than those of the cherimoya. The considered the least tasty of the cultivated flowers are long-stalked, triangular, yellow, 6 annonas. There are commonly more than 40 cm long and 4-5 cm wide15. The fruit is oblong, dark coffee-coloured seeds per fruit. conical or heart-shaped, weighs more than 5. Soursop - Annona muricata L. 1.00kg. The rind is thick and composed of The soursop has an erect growth habit with a fused areoles more prominent and angular than high canopy height-to-diameter ratio, although those of the sugar apple, with tips that are it tends to be low-branching and bushy, with rounded or slightly upturned; firm, pliable, and upturned limbs. It is a small, slender, indehiscent. Pulp is fragrant, snowy-white, evergreen tree, 4 to 8 m tall when fully mature. fine texture, almost solid, not conspicuously The stems are rounded, rough and not divided into segments, with fewer seeds than pubescent, with a dark-brown colour. The the sugar apple; sweet and subacid at the same leaves have short petioles, and are oblong- time and resembling the cherimoya in flavor. ovate to cylindrical, 14 to 16 cm in length and The seeds are cylindrical, dark brown or black 5 to 7 cm in width42. The flowers of soursop in colour13. are much larger than the other species, being 4. Bullock's heart / Ramphal - Annona 3.2 to 3.8 cm in length. The flowers start to reticulata L. open in the early morning and complete The tree reaches 6.0 to 7.5 m in height, with anthesis takes approximately 6h, depending on many lateral branches; stems are cylindrical, the climate. Soursop produces an ovate, with lenticels and very short coffee-coloured conical or heart-shaped fruit, that is dark green hairs. It is considered the most vigorous of the when unripe and a slightly lighter green when annonas. The fruit is considered to be of ripe. The rind has many short, fleshy, pointed inferior quality. It can be distinguished from carpel protuberances and is popularly regarded cherimoya by its long, narrow, glabrous as 'spiny'. The soursop has the largest fruit in leaves, from sugar apple by its solid, compact the genus, weighing from 0.9 to 10 kg, and fruit, as well as its larger leaves, and from A. averaging 4 kg41. The flavour is more acid and glabra by its small, dark brown seeds42. The less sweet .The fruit has 127 to 170 seeds, leaves are oblong-lanceolate and dark-green, scattered throughout the pulp. They are toxic. measuring 25 to 30 cm in length and 7 cm Seed size varies from 1 to 2 cm in length and wide, with 10 to 20 vein pairs and a pubescent from 0.33 to 0.59 g in weight, with a black petiole. Flowers are similar in form to those of colour soon after harvest, but becoming dark- sugar apple, except that they are grouped in a brown later42. short inflorescence with 2 to 10 flowers, with 6. Pond apple - Annona glabra L pedicels measuring 1.5 to 3.0 cm in length. The tree vigorous, commonly known by This species also presents inefficient natural Alligator apple, Monkey apple and originated pollination and poor fruit set13. in south eastern America and Mexico, the Fruits weigh from 0.1 to 1.0 kg and flowers are born in current season growth and are commonly heart-shaped, but may be flower length measures about 2-3 cm, the conical, ovate or irregular in form, and 10 to fruits are oblong or spherical in shape and 12 cm in length. They are coriaceous and have weight about 300-700 gm, the peculiar a reddish-yellow surface colour, with characters of pond apple are the flesh is sweet- impressed lines (around 5 to 6 angled areoles) scented flavour; resemble like papaya and above the carpels42. The flesh is milk-white pineapple flavour. This species is mainly used and sweet, although insipid in flavour, being for the drought tolerance and coloured flesh42. Copyright © April, 2017; IJPAB 1170

Vinay et al Int. J. Pure App. Biosci. 5 (2): 1168-1181 (2017) ISSN: 2320 – 7051 Reprodcutive Physiology phase. In glabra the anthesis takes place at 7 Floral biology: am to 9 am in the morning, anther dehiscence Flowering in most of the annona species occurs at 12 pm to 4 pm of next day of the appeared in different flushes although April anthesis and maximum receptivity found at the and September were observed to be major day of anthesis34. bloom periods under tropical humid eastern Pollination: coastal region of India. Flowers were terminal The flowers are hermaphrodite and or axillary and borne solitary, paired or in dichogamous in nature, both the male and multiflowered fascicles either on new female flowers are borne in the same flower25, branches, older branches or on the main trunk but the female part matures before the male as cauliflorous nature of bearing25. The flowers part (protogyny) in most of the species except are hermaphrodite and dichogamous in nature. pond apple (protoandry), therefore the annonas The soursop flowers are found to be possess highly cross pollination47. Natural protoandrous and other species are pollination in annona mainly carried by only protogynous in nature34. through insects, such as beetles (canthrophily), The anthesis takes place in sugar apple nitidulid beetles (Carpophilos domidiatus and flowers at morning 5:30 am to 6:30 am and C. hemipterous) play a major role in anther dehiscence at 12 pm to 2 pm of next pollination of sugar apple25, insects rarely day of the anthesis and stigma receptivity start visits the annona flowers because the flowers a day prior to anthesis and continue upto day are not attractive and have low mild fragrance after anthesis but found maximum receptivity to attracts the insects, which suggests that at the day of anthesis25. In cherimoya the canthorophylous pollination plays only a anthesis takes place at 7 am to 9 am in the secondary role in the annona species. Because morning, anther dehiscence at 12 pm to 4 pm pollen grains occur in clumps and are of next day of the anthesis and maximum somewhat sticky, wind pollination is not receptivity found at the day of anthesis. common13. Although dichogamy and the low Atemoya flowers are borne on current season population density of pollinators insects are growth, the peak anthesis of flowers at important limiting factors to successful natural afternoon 2 pm to 4 pm then the anther pollination, the effect of these factors may dehiscence accurse after a day of anthesis at 12 result in fertilization failure of all or several pm to 2 pm and the stigma receptivity start a ovules, resulting in small or asymmetrical day prior to anthesis and continue upto day fruits, which obviously affects the yield, after anthesis but found maximum receptivity quality and commercialization of annona at the day of anthesis. The anthesis takes place fruits49. in reticulata flowers at morning 6:00 am to Hand pollination is the only certain 8:00 am and anther dehiscence at 12 pm to 2 strategy to ensure commercial production, pm of next day of the anthesis and stigma since it guarantees significantly higher receptivity start a day prior to anthesis and production and better fruit quality than continue upto day after anthesis but found open/natural pollination in annona18,47. Hand maximum receptivity at the day of anthesis34. pollination improves fruit set and substantially In muricata the flowers exhibited protoandrous boosts fruit yield per tree by increasing the dichogamy, the peak anthesis occurs in the fruit size and number, the major yield afternoon 2:00 pm to evening 8:00 pm, anther components43. Hand pollination is normally dehiscence at 4 am – 8 am and stigma carried out before 8.00 am using a small brush. receptivity start after completion of male Pollen can be collected in the morning

Vinay et al Int. J. Pure App. Biosci. 5 (2): 1168-1181 (2017) ISSN: 2320 – 7051 between 5.00 and 8.00 am from fully open of clones were made for fruit quality only. flowers, when the sacs have turned from white Seedling screening from known mother plants to cream. The collected pollen is used to is another way of selection for better lines. pollinate half-open flowers whose pistils are Modern age requirements of a good cultivar already receptive18. involve: dwarfness, precocity, profuse and Pollinizers regular bearing, attractive, good sized and Several agents have been credited as quality fruit, absence of physiological pollinators of annona. Nitidulid beetles, disorders, disease and pest resistance and Carpophilos domidiahus and Carpophilos improved shelf life etc. Now, more hemipterous were identified as pollinators in comprehensive knowledge about the terms of visitation frequency, pollination phenology, advanced techniques for potential score and pollination efficiency. hybridization is available. Many However, Carpophilos hemipterous was environmental and physiological factors relatively effective pollinator than Carpophilos related to the undesirable character of annona domidiahus. Sugar apple clearly exhibit varieties (susceptibility to diseases and pests, obligate specialization by filtering only poor eating and keeping quality, etc., are Carpophilos hemipterous and Carpophilos closely controlled by genes. To overcome domidiahus as pollinators and thereby these, plant breeding can play an important explicitly indicating canthrophilous pollination role and work should be done in three 25 syndrome . directions as introduction, selection and Cytogenetics of annona hybridization. Hybrid populations can be Understanding cytology of a crop is useful in managed in a better way and the development planning breeding programme. The of the genetic markers has further reduced the chromosome numbers of cherimoya, reticulata, uncertainty in breeding of annona. soursop and sugar apple are 2n = 14 to 1610,33. Breeding objectives Although there is some variation in The first objective is the determination of the chromosome number, they are all diploids, 2n specific characteristics that are important for = 2x. This slight variation in chromosome the new cultivar - the ideo-type42. General and number may explain the ease or difficulty of very important objectives of breeding in interspecific hybridisation and warrants further annona are; Precocity in bearing, higher in work to determine if intra-specific variation percentage of flower and fruit set, have good also exists. A. glabra, are found to be 3 shelf life of fruits and yield, prefer tetraploid . symmetrical shape and good size fruits, prefer Perspective of breeding in Annona fruits without carpel projections that withstand The opportunity for breeding improvement in bruising during transportation, resistance to the annona is significant and challenging. common pests and diseases of annona and There is a lot more species wealth available but certain inherent constraints are involved other traits includes attractive skin colour, high like: long juvenility, high heterozygosity and pulp recovery, low number of seeds and good flavour. These characteristics may help a large area requirement for assessment of breeder to select parental groups to be used in hybrids. On the other hand, wide range of diversity and ease of vegetative hybrid an annona breeding programme, in order to propagation are the advantages for the obtain desirable progenies. These progenies may not have all desirable characteristics, but breeders. There are very few man made at least will have those most important to commercially important hybrids. Annona development is mostly based on the selection growers, retailers and consumers. Copyright © April, 2017; IJPAB 1172

Vinay et al Int. J. Pure App. Biosci. 5 (2): 1168-1181 (2017) ISSN: 2320 – 7051 Table 1: Main characteristics of cherimoya, soursop and custard apple ideo-types (adult plants)42 Characteristics Cherimoya Soursop Custard apple Vegetative vigor Medium – low Medium – low Medium – low

Yield > 30kg/tree/yr > 60kg/tree/yr > 20kg/tree/yr Bearing Regular Regular Regular PLANT Rootstock/scion High High High compatibility Flower number Abundant Abundant Abundant (>150 (>100 (>180 flowers/tree) flowers/tree) flowers/tree) Pollen fertility High (> 76%) High (> 76%) High (> 76%)

Size/ Industry Large/>650g Large/>2.5kg Large/>400g weight fresh Medium/300- Small/800g Medium/300- consumption 600g 400g Shape Heart Conical Round to Heart

Fruit set (Natural) High (>27%) High (>26%) High (>25%)

Skin Smooth Short Tuberculate Fruit protuberances Pulp Sweet, Sub-acid, low Sweet, low fiberless fibre fibre

Flavor Delicate Sub-acid flavor Pleasant acidulous

Seed number/ 100g of pulp Low/<6 seeds Low/10-30 Low/<10 seeds seeds

Shelf life Long Long Long (> 10 days) (> 5 days) (> 5 days)

Breeding Strategies between its parents44,46. A. reticulata flourishes According to Jordan and Botti21, there is great well in costal lowlands. This adaptation scope in edible annonas for gene transfer suggests that it may be possible to break the across species boundaries, for widening the geographical barriers of annonas through genetic base and for developing strategic interspecific hybridization and construct new breeding reserves. Gene exchange among annonas suitable to wider or new climatic annonas should result in interesting novel zones. recombinants, with the popular atemoya being Most Annona species and cultivars the most esteemed one currently available. differ in environmental adaptation, Additionally, annona species vary in their productivity and fruit quality. Therefore, climatic adaptation. In the tropics, cherimoya different conventional methods can be used in grows well at higher altitudes and sugar apple their breeding. According to Fehr9, there are at lower altitudes, while the interspecific three requirements for the development of an hybrid, atemoya adaptation is intermediate asexually propagated cultivar: a) a suitable Copyright © April, 2017; IJPAB 1173

Vinay et al Int. J. Pure App. Biosci. 5 (2): 1168-1181 (2017) ISSN: 2320 – 7051 source of genetic variability b) evaluation of characterization and documentation, followed individuals from the population c) asexual by evaluation and selection. Several types of multiplication of a new cultivar for populations can be developed by commercial use. hybridization, from which superior clones are Selection and cultivar development selected42. Introduction of superior genotypes and/or India and Taiwan have selected a few cultivars to establish a germplasm collection named cultivars of sugar apple from seedling is, basically, the first requirement of any population that are propagated vegetatively. In breeding programme15. This can be Cuba, researchers developed ‘Cuban complemented by the introduction into the Seedless’, which is a seedless cultivar with collection of some wild Annona species with medium-sized fruits, and another cultivar with useful genes, mainly for resistance to diseases. low fibre content that is very important for the All accessions require comprehensive commercial market33.

Table 2: Some selections and cultivars of sugar apple that are currently grown in various countries42 Cultivar/Selection Country of origin Cultivar/Selection Country of origin IPA selection Brazil Leone Italy Libby Madeira Portugal Lisa Mateus 1 USA Ott Negrito Spain Red sugar apple Reretai New Zealand Noi Thailand Abd El-Razik Egypt Molate Cuban Seedless Cuba Philippines Lobo Balanagar Mossman Mammoth India Australia Whaley Red Sitaphal

Table 3: Selected cultivars of atemoya42 Cultivars Country of origin Maroochy Gold, KJ Pinks, Australia Pink’s Mammoth African Pride Southern Africa Bradley, Page Florida, USA Gefner, Kabri, Malalai Israel

Table 4: Selected cultivars of cherimoya42 Cultivars Country of origin

Andrews, Kempsey, Mossman Australia

Bays, Booth, Libby, Lisa, White California, USA

Bronceada, Concha Lisa Chile

Burton’s Wonder New Zealand

Campa, Cristalino Spain E-8 Ecuador

Vinay et al Int. J. Pure App. Biosci. 5 (2): 1168-1181 (2017) ISSN: 2320 – 7051 Some of the new selections of sugar apple are ha, in adjoining districts during last 8-10 years developed by different institutions in India; (IHC, 2014). Central Horticultural Experimental Station Some cultivars were identified in (ICAR-IIHR), Bhubaneshwar, develop a new India based on colour of the fruit , its origin clonal selection and named as Arka Neelchal and plant part: Yellow custard apple named Vikram, which bears good quality fruits with after the colour of the fruit; Cultivars like high yield (>15 kg/tree/year), the shelf life Kakarlapahad, Mahaboobnagar, Local upto 5-6 days after maturity and have good Hyderabad and Saharanpur Local named after taste with high TSS (24OB). Regional the place of origin; White stemmed custard Research Station (TNAU) – Aruppukottai, apple named after the part of plant. Tamilnadu, develop another sugar apple Hybridization selection; APK-1, which gives more yield, Wester52 was the first scientist to realize the bears good quality fruits and more suitable for possibilities for genetic improvement of South India. Another selection release from annonas and initiated breeding programmes in Australian custard apple growers association Florida and in the Philippines, due to some (ACAGA) in Australia, the variety belong to problems during the breeding programme no atemoya, which is named as Tropic Sun, hybrids has been developed on that time42. which is semi dwarf and good for high density Globally, interspecific hybridization in planting, more suitable for tropical and sub- annonas is limited to four research groups, one tropical regions of the world with sweet and each in Australia, America, Israel and good quality fruits. India24,54. Although institutions did work on the Several types of populations can be release of varieties in annona also some developed by hybridization, from which farmers in the India has been developed some superior clones are selected and released as varieties of the sugar apple. Shri Navnath new hybrid after evaluation11, breeding in Malhari Kaspate, he is the farmer from annona has been neglected and only a few Solapur district of Maharastra, he develop four hybrids have been developed in the past, due varieties; Annona-2, NMK-1, NMK-2 and to lack of breeding programmes and clear NMK-3 (IHC, 2014), among these NMK-1 strategies. Australian Custard Apple Growers gain lot of importance in south India and Association started breeding programme with commonly called as golden, because of its the objective of developing high quality colour and quality of the fruits. Impact of these seedless hybrids of atemoya14,39. In this varieties; The productivity of existing varieties breeding programme, first they produce with traditional cultivation is about 10 to 12 tetraploids, either through gamma irradiation ton / ha. Whereas the productivity of new or colchicine, and crossed these to diploids to selections is about 15-19 ton/ha, with 70-80% produce seedless triploids. Other potentially Grade-1 fruits. Due to good quality fruits useful methods of obtaining seedlessness in production the market rates are 120-150% annonas, a) progeny from cross between more over existing varieties. The benefit-cost diploid x diploid with small size seeds, b) Ratio is 4.6. Near about 700-800 farmers from producing triploids from diploid x tetraploid 9-10 district of Maharashtra cultivators got the and tetraploid x diploid crossing, c) protoplast benefit of Custard Apple production fusion of diploids and haploids, d) crossing of technology & new cultivars and diversified low seeded parents, e) development of self- their cropping pattern towards dry land fruit incompatible parthenocarpic hybrids and f) farming. endosperm culture to develop triploids42. Due to the efforts of Mr. N.M. Girwani et al16., also developed certain Kaspate and Krishi Vigyan Kendra, Solapur sugar apple hybrids at Sangareddy, India, and the area under Custard Apple increased to 800 revealed that All the hybrids varied in fruit ha. In Solapur district and approximately 450 shape (round, conical and cordate), fruit colour Copyright © April, 2017; IJPAB 1175

Vinay et al Int. J. Pure App. Biosci. 5 (2): 1168-1181 (2017) ISSN: 2320 – 7051 (yellowish green, grayish green, light green (female) x Annona muricata (male). The and red), pulp colour (creamy white, light pink former F1 hybrids produced plants that and white), areole shape (tuberculate, flowered but set no fruit. The latter F1 hybrids hexagonal) and texture (course, soft and produced fruit similar to Rollinia31. Out of 35 meaty). Time of fruit maturity varied from germplasm of custard apple evaluated the September 2nd fortnight and lasted till the end varieties viz; Atemoya X Balanagar (highest of December. Hybrid-1 (17/4 Atemoya × fruit weight - 278.9g), Y. Palli – 12 (highest Balanagar) was earliest to mature. Maximum pulp weight – 148g), NLD – 8 (maximum fruit weight (250 g) was recorded in Hybrid-1 T.S.S – 25.9), Balanagar SR (more number of (17/4 Atemoya × Balanagar) followed by fruits/tree – 51.4) and Ramaphal (highest Hybrid-6 (15/3 Red Sitaphal × Atemoya) (225 yield/tree – 15.7) can be recommendable for g) and Hybrid-4 (1/6 British Guinea × scarce rainfall zone under rainfed conditions in Atemoya) (220 g). Among all the hybrids, poor fertile soils4. Among the 22 accessions, maximum number of fruits per tree (94) was notable variations were observed in tree shape, recorded in Hybrid-1 (17/4 Atemoya × trunk colour, trunk ramification, colour of Balanagar) followed by Hybrid-6 (15/3 Red young branches, leaf blade shape, pulp colour Sitaphal × Atemoya) (67). The TSS ranged and taste. Generally, soursop genotypes from 22 to 28°B and was found to be showed good adaptation to the specific soil maximum in Hybrid-2 (15/2 Red Sitaphal × and climate conditions of these regions and Pond apple), while the seed content per fruit great variability was found among them30. was also minimum (20) in this hybrid. Based There is a growing awareness particularly on overall performance the Hybrid-1 (17/4 about the anti cancer properties of this plant Atemoya × Balanagar) and Hybrid-3 (15/3 among the people and the consumption of this Red Sitaphal × Atemoya) and Hybrid-2 (15/2 fruit and leaves by the cancer-affected patients Red Sitaphal × Pond apple) were found to be is on the increase. So, it has become necessary excellent in quality with pleasant aroma, to document the trees available in Tamil Nadu smooth pulp texture with less seed, good sugar and variations them to exploit the potential of acid blend and shelf life. the trees30. Crosses have also been made between In India breeding work was initiated in four different species; A. cherimola, A. 1989 at the Indian Institute of Horticultural squamosa, A. reticulata, A. diversifolia and A. Research, Bangalore on evolving a less seeded atemoya. To date none of the progeny of these custard apple cultivar with higher proportion crosses has produced commercial cultivars. In of good quality edible pulp and better shelf florida approximately 3000 seedling life20. More than 2000 inter and intra-species 14 progeny , mainly interspecific crosses, have Annona hybrids F2 backcrosses were been planted out in Zill orchards near Boynton developed and evaluated which resulted in the beach. To date, none of the progeny of these release of a hybrid by the name ‘Arka Sahan’. crosses has produced commercial cultivars. It is an outstanding recombinant isolated from The studies conducted in Serdang, Selangor, the family of ‘Island Gem’ (atemoya) and Malaysia, involved six selected species of ‘Mammoth’ (custard apple). However, for Annonaceae family, namely: Annona improving the fruit-set, fruit-size and fruit- muricata, A. squamosa, A. glabra, A. montana, shape of this rare interspecific recombinant A. rollinia, and A. reticulata. Hybridizations pollination with Annona squamosa pollen was were attempted between the species and also found to be most effective18. Fruits of ‘Arka their reciprocal crosses. Although fruits are Sahan’ are remarkable for scarce small seeds formed between some species, seeds were not (9/100 g fruit weight), sweetness (>32°B) and viable except for those produced in two slow ripening (5-6 days). Nutritionally, 100 g hybridizations: Annona muricata (female) x pulp of ‘Arka Sahan’ contains 2.49 g crude Annona montana (male) and Rollinia mucosa protein, 42.29 mg phosphorus and 225 mg Copyright © April, 2017; IJPAB 1176

Vinay et al Int. J. Pure App. Biosci. 5 (2): 1168-1181 (2017) ISSN: 2320 – 7051 calcium compared to 1.33 g, 17.05 mg and 159 classic vegetative propagation methods, mg respectively, in common custard apple. because the induction rate of adventitious roots After success of Arka Sahan the A. from scions obtained from adult specimens atemoya x A. reticulata was another cross with agronomic interest is null. This low combination that has attracted interest as A. morphogenetic capacity is the general trend for reticulata carries genes for uniform heart- most of other annona species even if it shaped fruits with a stout stalk and smooth and happens in different degrees depending on the attractive skin colour as well as having a species. In order to fix the Annonaceae robust root system and compact tree habit vegetative propagation problems, in vitro (Council of Scientific and Industrial Research methods have been applied. But unfortunately 1985). Jalikop19 studied that A. reticulata is the limited funds and human resources devoted potentially a valuable source of a variety of to this subject never reach the necessary novel traits. However, the F1 trihybrids volume to obtain quick and substantial exhibited reduced fertility with pronounced advances. undesirable traits of A. reticulata and the The in vitro tissue culture methods of survival of F2 trees was poor, with those that micropropagation can be applied successfully did survive being extremely weak and failing to cherimoya and other annona species to to flower or set fruit. Consequently, overcome these problems. Most of the backcrossing may be more suitable approach protocols of micropropagation and than advancing trihybrid generation for regeneration were developed using the cultivar transferring desirable genes from A. reticulata. Fino de Jete, which is the major cultivar in

Backcrossing F1 progeny of [(A. cherimola x Spain. First it is developed the method to A. squamosa) x A. reticulata] to A. squamosa, micropropagate the juvenile material of a highly fertile species, is expected to yield cherimoya5, and later it was optimized a productive introgressants in which the protocol to micropropagate adult cherimoya contribution of the A. squamosa genome will genotypes selected by outstanding 37 be extended from 25 to 62.5% in the F1BC1 agronomical traits and further it was generation19. The genetics is described herein improved the process through micrografting38. based on the expressions of certain tree and At the present time we are involved in fruit traits in the progeny. inducing and obtaining new elite genotypes, as Biotechnology part of a breeding program for the cherimoya The tissue culture in annona is mainly used for and other annonas, using and optimizing the micropropogation, maintains of germplasm different methodologies in vitro: a) collection, used for the embryo rescue Adventitious organogenesis and regeneration technique, preparation of explants for from cellular cultures6, b) Ploidy manipulation transformation of genes, enhancement of of the cherimoya, to obtain haploid, tetraploid variability by somaclonal variation and and triploid plants (seedless), c) Genetic production of haploid plants42. transformation, for the genes introduction to The vegetative propagation of these control the postharvest processes and the genes species present problems of different degree, introduction to provide resistance to pathogen being its sexual propagation of limited and insects and d) Micropropagation and agronomic value due to the high degree of regeneration of other wild Annona or related heterozygosis of these species and preventing species such as: Annona senegalensis, A. their propagation by seed. scleroderma, A. montana, A. reticulata, A. Among these Annonaceae, in glabra, A. diversifolia and Rollinia sp. cherimoya (Annona cherimola), the most The first haploid plants induced by exploited annona, the morphogenetic capacity anther culture in fruit trees were reported in is extremely low, showing minimal level of sugar apple. Aim of the work is to rooting making almost impossible the use of development of seedless fruits. The Copyright © April, 2017; IJPAB 1177

Vinay et al Int. J. Pure App. Biosci. 5 (2): 1168-1181 (2017) ISSN: 2320 – 7051 researchers obtained callus differentiation, and provides the potential to introduce this feature formation of roots and shoots from sugar apple into annona fruit production. Jorge et al22., endosperm, but a complete plantlet was not determined the developmental cause of obtained32. seedlessness in Ts and, using information from Genetic transformation studies on Arabidopsis, showed that the The preliminary steps in the cherimoya genetic mutant is associated with the deletion of the A. transformation via Agrobacterium have been squamosa INO (inner no outer) gene. A unique done to increases the shelf life, to change post- case of elucidation of the function of a gene harvest characteristics and to provide pest and through forward genetics. A potentially disease resistances. For other Annona species agronomically useful manifestation of the there are no advanced studies7. particular mutation is the production of Molecular markers seedless fruits. The determination of the In annona the molecular markers are mainly molecular basis of the Ts mutant can aid used for germplasm characterization, hybridity introgression of the trait into commercially confirmation, identification of parents of the desirable varieties by providing a method for progeny/hybrid and diversity analysis of identification of homozygous mutant seedless annona. There are limited reports found on plants. The likely conservation of the function exploitation of molecular markers for diversity of ino in the genetic regulatory pathway of analysis in annonas. Few of these markers are ovule development across angiosperms random amplified polymorphic marker broadens the implications of these results22. (RAPD) marker1,48, amplified fragment length polymorphism (AFLP) markers45,53 and simple CONCLUSION sequence repeat (SSR) markers8,26. On the Immense prospects exist to develop desirable other hand, information regarding nutritional cultivars of annonaceous fruits by value is of utmost important to select desired conventional breeding techniques, there is genotype for domestication in area of greater scope to exploit inter-specific variation adaptation. Very little information is available than intra-species variation, edible annona on proximate analysis of annona fruits2,23,36. species exhibit morphological affinity to one During analysis, a set of 11 RAPD primers another but each one is unique and distinct for yielded a total of 152 bands with 80.01 % fruit shape, size, skin surface, pulp colour, polymorphism and 12 SSR primers produced texture, flavour and taste, some species like 39 amplicons. The dendrogram discriminated annona squamosa exhibit narrow genetic all the annona genotypes suggesting that variation. However, several interesting traits significant genetic diversity was present are available in one or the other allied Annona among the genotypes17. Proximate fruit species, because there are no serious barriers composition study of nine fruiting genotypes in inter-specific hybridization between A. of annona revealed that A. squamosa squamosa, A. cherimola, and A. reticulata possessed significantly higher amount of most gene transfer across species boundaries is of the studied biochemicals which gives an possible, gene exchange among them should opportunity to fruit breeders to improve the result in interesting novel recombinants at other annona species. Likewise, A. muricata times most esteemed ones like the popular being rich in seed oil content can be exploited atemoya. in oil industries17. Mutation REFERENCES There is very limited work has been done in 1. Bharad, S.G., Kulwal, P.L. and Bagal, annona for development of mutant varieties S.A., Genetic diversity study in Annona but the natural mutants are obtained; Brazilian squamosa by morphological, biochemical seedless and Thai seedless (Ts), Thai seedless, and RAPD markers. Acta Horticulture., produces fully seedless normal-sized fruits and 839: 615–623 (2009). Copyright © April, 2017; IJPAB 1178

Vinay et al Int. J. Pure App. Biosci. 5 (2): 1168-1181 (2017) ISSN: 2320 – 7051 2. Boake, A.A., Faustina, D., Jacob, K. and 14. George, A.P., Broadley, R.H., Nissen, Ibok, O., Dietary fibre, ascorbic acid and R.J., Hamil, S.D. and Topp, B.L., proximate composition of tropical Breeding startegies for atemoya and underutilized fruits. African Journal of cherimoya. Acta horticulture., 497: 255- Food Science., 8: 305–310 (2014). 260 (1999). 3. Datta, P.C. and De, B., Karyology of 15. George, A.P., Broadley, R.H., Nissen, Some Indian Annonaceae. Cytologia., 55: R.J., Hamill, S.D. and Drew, R., Breeding 187-196 (1990). new varieties of atemoya. Acta 4. Dhanumjaya Rao, K. and Subramanyam, Horticulture., 575: 323–328 (2002). K., Growth and yield performance of 16. Girwani, A., Madahvi, A., Kumar, T.S. custard apple germplasm under scare and Reddy, G.S., Evaluation of custard rainfall zone. Indian Journal Agricultural apple hybrids for fruit yield and quality Research., 45(2): 156-160 (2011). attributing characters. Acta Horticulture., 5. Encina, C.L., In vitro morphogenesis of 890: 251-254 (2011). juvenile Annona cherimola Mill. bud 17. Hirdayesh Anuragi, L., Haresh, Dhaduk, explants. Journal of Horticultural Sushil Kumar, J., Jitendra, Dhruve, J., Science., 69(6): 1053-1059 (1994). Mithil, A,. Parekh, A., Amar, and Sakure, 6. Encina, C.L., Biotechnology of fruit and Molecular diversity of Annona species and nut crops. Wallingford; CAB proximate fruit composition of selected International., 74-88 (2004). genotypes, Biotechnology., 6: 204 (2016). 7. Encina, C.L., Padilla, I.M.G., Carzola, 18. Jalikop, S.H. and Ravindra Kumar, Psedo- J.M. and Caro, E., Tissue culture in xenic effect of allied Annona spp. Pollen Cherimoya. Acta Horticulture. 497: 289- in hand pollination of Cv.’Arka Sahan’ 294 (1999). [(A.cherimola x A.squamosa) x 8. Escribano, P., Viruel, M.A. and Hormaza, A.squamosa]. Hort Science., 42(7): 1534- J.I., Characterization and cross species 1538 (2007). amplification of microsatellite (SSRs) 19. Jalikop, S.H., Tree and fruit traits of markers in cherimoya (Annona cherimola progenies from the cross between (A. Mill.). Molecular Ecology., 4: 746–748 cherimola Mill. x A. squamosa L.) and A. (2004). reticulata L. and approaches for the 9. Fehr, W.R., Principles of cultivar introgression of valuable genes from development - Theory and Technique, A.reticulata L. Euphytica., 173: 161-171 Macmillan publishing company, New (2010). York. 1: (1987). 20. Jalikop, S.H., Breeding of Pomegranate 10. Folorunso, A.E. and Olorode, O., and Annonaceous Fruits. Acta karyotype studies in some species of the Horticulture., 890: 191-196 (2011). Annonaceae. Ife journal of science., 9(2): 21. Jordan, M. and Botti, C., Tropical and 161-166 (2007). subtropical small fruits. In: Biotechnology 11. Gardiazabal, I.F. and Cano, G.L., of perennial fruit crops. CAB Characterization of Cherimoya Cultivars International., 519-522 (1992). (Annona cherimola Mill.), Acta 22. Jorge Lora, I., Jose, Hormaza, Maria Horticulture., 497: 239-253 (1999). Herrero, S., Charles Gasser, Seedless fruits 12. George, A.P., Nissen, R.J. and Brown, and the disruption of a conserved genetic B.I., The custard apple. Quinceland pathway in angiosperm ovule Agriculture Journal., 113: 287–297 development, Plant Biology., 108(13): (1987). 5461–5465 (2011). 13. George, A.P. and Campbell, J., Hand 23. Kulkarni, S., Joshi, S., Kamthe, P. and pollination in subtropical Australia. Tekale, P., Proximate analysis of peel and California Growers., 15(2): 36–38 (1991). seed of Annona squamosa (Custard apple) Copyright © April, 2017; IJPAB 1179

Vinay et al Int. J. Pure App. Biosci. 5 (2): 1168-1181 (2017) ISSN: 2320 – 7051 fruit. Research Journal of Chemical 34. Nalawadi, U.G., Sulikeri, G.S. and Singh, Science., 3: 92–94 (2013). C.D., Floral biological studies of Annona 24. Kumar, P.S. and Jalikop, S.H., Cross- squamosa L. under Dharwad condition. compatibility among Annona species. Progressive Horticulture., 7(1): 15-24 Indian Journal of Horticulture., 57(4): (1975). 309-313 (2000). 35. Ochse, J.J., Soule, M.J., Dijkmen, M.J. 25. Kundan Kishore., Shukla, A.K., Naresh and Welhburg, C., Outros cultivars Babu., Sarangi, D.N. and Samiksha Frustules, Plants of tropical and Patanayak, Pollination biology of Annona subtropical, Mexico, 587-818 (1974). squamosa L.(Annonaceae): evidence for 36. Onimawo, I.A., Proximate composition pollination syndrome. Scientia and selected physicochemical properties of the seed, pulp and oil of soursop (Annona Horticulture., 144: 212-217 (2012). muricata). Plant Food Human Nutrition., 26. Kwapata, K., Mwase, W.F., Bokosi, J.M., 57: 165–171 (2002). Kwapata, M.B. and Munyenyembe, P., 37. Padilla, I.M.G. and Encina, C.L., Genetic diversity of Annona senegalensis Micropropagation of adult cherimoya Pers. Populations as revealed by simple (Annona cherimola Mill). Cv. Fino de sequence repeats (SSRs). African Journal Jete. In vitro Cellular and Developmental of Biotechnology., 6: 1239–1247 (2007). Biology-Plant., 40: 210-214 (2004). 27. Lizana, L.A. and Reginato, G., Cherimoya. 38. Padilla, I.M.G. and Encina, C.L., The use In: Fruits of Tropical and Subtropical of consecutive micrografting improves Origin: Composition, Properties and Uses. micropropagation of cherimoya (Annona Florida Science Source., 131-148 (1990). cherimola Mill.) cultivars. Scientia 28. Ludders, P., Cherimoya (Annona Horticulturae., 129: 167-169 (2011). cherimola Mill.) Botany, cultivation, 39. Pawshe, Y.H., Patil, B.N. and Patil, L.P., storage and uses of a tropical-subtropical Effect of pre germination seed treatment fruit. Erwerbsobstbau., 44(4): 122–126 on the germination and vigour of seedlings (2002). in custard apple (Annona squamosa L.), 29. Manica, I., Taxonomy, Morphology and Annuals of Plant Physiology., 2(11): 150- Anatomy, Proceedings of Brazilian 154 (1997). symposium on annonaceous, Vitoria da 40. Pena, J.E., Nadel, H., Barbosa Pereira, M. Conauista. 20-35 (1997). and Smith, D., Pollinators and pests of 30. Manigandan, S. and Vijayakumar, R.M., Annona species. Tropical fruit pests and Morphological Characterization of pollinators. CAB International., 197–221 Soursop (Annona muricata L.) Germplasm (2002). 41. Pinto, A.C. and Silva, E.M., Graviola para in Tamil Nadu. Trends in Biosciences., Expartacao, Flores e plantas 7(13): 1458-1460 (2014). Ornamentais., 41 (1994). 31. Mohd Khalid, M.Z. and Drew, R., 42. Pinto, A.C., De, Q., Cordeiro, M.C.R., Hybridization between selected Andrade, S.R.M., De., Ferreira, F.R., Annonaceae species. Acta Horticulture., Filgueiras, H.A., De C., Alves, R.E. and 575: 367–369 (2002). Kinpara, D.I., Annona species. 32. Nair, S., Gupta, P.K. and Mascarenhas, International Centre for Underutilized A.F., Haploid plants from In Vitro anther Crops, University of Southampton, UK culture of Annona squamosa L. Plant Cell (2005). Reports., 2: 198-200 (1983). 43. Pritchand, K.D. and Edwards, W., 33. Nakasone, H.Y. and Paull, R.E., Annonas, Supplementary pollination in the In: Tropical fruits, CAB International., 45- production of custard apple the effect of 75 (1998). pollen source. Journal of Horticulture Copyright © April, 2017; IJPAB 1180

Vinay et al Int. J. Pure App. Biosci. 5 (2): 1168-1181 (2017) ISSN: 2320 – 7051 Science and Biotechnology., 81: 78–83 49. Saavedra, E., Influence of pollen grain (2006). stage at the time of hand pollination as a 44. Purseglove, J.W., Annonaceae. Tropical factor on fruit set of Cherimoya. Hort crops. Dicotyledons. Longman, London. Science., 12: 117-118 (1997). 625–626 (1968). 50. Singh, S.P., Fruit crops for wasteland, 45. Rahman, M., Shimada, T., Yamamoto, T., Scientific publisher., India (1992). Yonemoto, J.Y. and Yoshida, M., 51. Thakur, D.R. and Singh, R.N., Studies on Genetical diversity of cherimoya cultivars floral biology of Annona. Indian Journal revealed by amplified fragment length of Horticulture., 22: 238-253 (1965). polymorphism (AFLP) analysis. Breed 52. Wester, P.J., Annonaceous possibilities Science., 48: 5–10 (1998). for the plant breeder. Philippine 46. Rasai, S., George, A.P. and Kantharajah, Agriculture Review., 6: 312-321 (1913). A.S., Tissue culture of Annona spp. 53. Zhichang, Z., Guibing, H., Ruo, O., (cherimoya, atemoya, sugar apple and Yunchun, L., Yeyuan, C. and Shirong, L., soursop): A review. Scientia Studies of the genetic diversity of seven Horticulturae., 62: 1-14 (1995). sweetsop (Annona squamosa L.) cultivars 47. Richardson, A.C. and Anderson, P., by amplified fragment. African Journal of Flowering date affects pollination of Biotechnology., 10(35): 6711–6715 cherimoya, The Orchadist., 66(6): 28-31 (2011). (1996). 54. Zill, G.E. and Mahdeem, H., General 48. Ronning, C.M., Schnell, R.J. and Gazit, S., observations on hybridizing Annona Using Randomly Amplified Polymorphic species. Proceedings Florida State DNA (RAPD) Markers to Identify Annona Horticulture Society., 111: 314– 315 Cultivars. Journal of American Society of (1998). Horticulture Science., 120(5): 726-729 (1995).