Female and male sterility cause low fruit set in a clone
of the `Trevatt' variety of apricot (
Prunus armeniaca
)
A.M. Lillecrapp, M.A. Wallwork, M. Sedgley
*Department of Horticulture, Viticulture and Oenology, Waite Agricultural Research Institute, The University of Adelaide, Glen Osmond, S.A. 5064, Australia
Accepted 9 April 1999
Abstract
This study investigated ovule and anther structure of the `Trevatt Blue' variety of apricot (Prunus armeniaca) using bright field microscopy following reports of low fruit set. Ovules and anthers from fertile `Moorpark' and `Trevatt Knight' flowers were compared with those of the `Trevatt Blue'. In the `Moorpark' and `Trevatt Knight' ovules, all reproductive structures were present including embryo sacs with a complete set of eight nuclei and the anthers contained mature pollen grains. Multiple ovules which were small and retarded in development were present in the `Trevatt Blue' apricot flowers and the anthers contained degenerated microspores, with some failure in tapetal breakdown. This is the first report of a simultaneous mutation in both female and male function in apricot.#1999 Elsevier Science B.V. All rights reserved.
Keywords: Apricot; Fertility;Prunus; Microscopy; Ovule; Embryo sac; Anther; Pollen grain
1. Introduction
`Trevatt' is an old self-fertile cultivar of apricot (Prunus armeniaca) and high yielding clones have been developed to produce large fruit with desirable characteristics. The clone, `Blue' was planted over wide areas by many growers because the original selection was high yielding with fruit of excellent canning quality, but the trees failed to set fruit for several seasons after the juvenile phase had ended.
* Corresponding author. Tel.: +61-8-8303-7401; +61-8-8303-7249; fax: +61-8-8303-7116
E-mail address:[email protected] (M. Sedgley)
Some varieties of apricot have been found to be self-sterile or have pollination problems (McLaren and Fraser, 1996), but it seemed that in `Blue' female sterility may have been a possible contributor to low fruit set. Very low levels of set were achieved even when pollen from a different cultivar was introduced to the trees via bouquets from fertile orchards. It was also possible that male sterility contributed to the low fruit set. Female and male sterility have been reported independently in apricot (Eaton and Jamont, 1964; Nakanishi, 1983; Medeira and Guedes, 1991; Burgos and Egea, 1994) as well as in many other tree crops (Sedgley and Griffin, 1989), but have been attributed to adverse environmental conditions.
In this study, the female and male structures of `Trevatt Blue' were observed microscopically and compared with those of `Trevatt Knight' which is known to be fertile. A comparison was also made with the cultivar `Moorpark' in case there was a problem inherent in the `Trevatt' cultivar.
2. Materials and methods
Flower samples of the low yielding `Trevatt Blue' were randomly collected from a single variety apricot orchard in Griffith, New South Wales. Fertile `Trevatt Knight' flowers were obtained from a mixed planting at Loxton, South Australia, and fertile `Moorpark' flowers were obtained from the Waite orchard of the University of Adelaide, South Australia.
Flowers were collected at the late balloon stage just prior to anthesis and fixed in FPA50 (90% ethanol at 50%, 5% propionic acid and 5% formaldehyde). Ovules and anthers were dissected from each flower under a microscope and the samples dehydrated using a tertiary-butyl alcohol series and embedded in GMA (glycol methacrylate). Serial longitudinal 4.0mm sections through each ovule (up
to 150 sections per ovule) and48 sections of 4.0mm through each anther were
collected onto a microscope slide, stained with periodic acid-Schiffs reagent (PAS) and toluidine blue O (TBO) stain and mounted in methyl methacrylate in xylene (O'Brien and McCully, 1981). The sections were observed under a Zeiss Axiophot photomicroscope with bright field illumination. Ovules and anthers from 36 `Trevatt Blue', 15 `Trevatt Knight' and 9 `Moorpark' flowers were examined. The structure of male and female reproductive structures was recorded for each flower.
3. Results
smaller in size than the fertile `Moorpark' and `Trevatt Knight' ovules but all similar in size to each other. One hundred and four ovules were obtained from 36 `Trevatt Blue' flowers; 2.8% of flowers had one ovule, 27.8% had two, 47.2% had three and 22.2% had four (Table 1).
Of the ovules examined of the `Moorpark' and `Trevatt Knight', all had an outer integument, inner integument, nucellus, micropyle and embryo sac (Fig. 1(A)) with an egg cell (Fig. 1(B)), two synergids (Fig. 1(C)), two polar nuclei (Fig. 1(D)) and three antipodals (Fig. 1(E)), which had degenerated in some ovules.
All `Trevatt Blue' ovules had an outer integument, inner integument and nucellus, and 88.5% had a micropyle (Fig. 2(A)). Some had mis-shapen nucellus (Fig. 2(B)), and embryo sacs were degenerated or not present in 50% of ovules (Table 1). The other 50% contained embryo sacs with 0, 1 (Fig. 2(C)), 2 (Fig. 2(D)) or 4 (Fig. 2(E)) nuclei. No `Trevatt Blue' apricot ovules observed contained eight nuclei, and 19.2% of ovules were at the megaspore mother cell stage. Other abnormalities included small, spherical ovules, ovules joined together and ovules with underdeveloped or abnormal nucellus (Fig. 2(B)) or nucellus incompletely surrounded by the integuments.
The anthers of the `Moorpark' and `Trevatt Knight' flowers were bright yellow in colour and plump compared to those of the `Trevatt Blue' flowers which were red-brown and shrunken. Anthers from `Moorpark' and `Trevatt Knight' apricot flowers had an endothecium and degenerated tapetum (Fig. 3(A)). Fully developed pollen grains with an exine, intine, germination pores, vegetative Table 1
and generative nuclei were present in anthers from all flowers (Fig. 3(B) and (C)). `Trevatt Blue' anthers were shrunken (Fig. 4(A)) with an endothecium (Fig. 4(B)), 72.2% with degenerated tapetum (Fig. 4(C)) but the remainder without tapetal degeneration (Fig. 4(B)). No pollen grains were observed in the anthers and only degenerated lumen contents were visible (Table 2).
Fig. 1. (A±E) Longitudinal sections of `Moorpark' (figures A, B, D and E) and `Trevatt Knight' (figure C) apricot ovules stained with periodic acid-Schiff's reagent (PAS) and toluidine blue O (TBO) and photographed using bright field optics. (A) Ovule showing embryo sac (es), nucellus (n), outer integument (oi), inner integument (ii) and micropyle (m). (B) Embryo sac with egg cell (e). A polar nucleus is also visible. (C) Embryo sac showing two synergids (s). (D) Embryo sac showing two polar nuclei (pn). An egg cell and antipodal are also visible. (E) Embryo sac showing three antipodals (a). A polar nucleus and a synergid are also visible. Bar represents 200mm in A, 700mm
4. Discussion
The results show that both female and male sterility contributed to low fruit set in `Trevatt Blue' apricot trees. `Trevatt Knight' was both female and male fertile and there are no other reports of sterility in the cultivar indicating a simultaneous mutation in female and male function. `Trevatt Knight' was structurally similar to Fig. 2. (A±E) Longitudinal sections of `Trevatt Blue' apricot ovules stained with PAS and TBO and photographed using bright field optics. (A) Ovule showing nucellus (n), outer integument (oi), inner integument (ii) and micropyle (m). (B) Ovule showing an abnormal and underdeveloped nucellus (n). (C) Embryo sac with one nucleus. (D) Embryo sac with two nuclei. (E) Embryo sac with four nuclei. Bar represents 200mm in A, 100mm in B, 450mm in C, 600mm in D and
Fig. 3. (A±C) Transverse sections of `Moorpark' (figures A and B) and `Trevatt Knight' (figure C) apricot anthers stained with PAS and TBO and photographed using bright field optics. (A) Anther lobe showing endothecium (en), degenerated tapetum (dt) and pollen grains (pg). (B) Pollen grain showing intine (i), exine (ex) and nuclei (nc). (C) Pollen grain showing germinaton pores (gp). Bar represents 200mm in A and 750mm in B and C.
Table 2
Anther structure for fertile `Moorpark' and `Trevatt Knight', and sterile `Trevatt Blue' apricot flowers at anthesis
`Moorpark' `Trevatt Knight' `Trevatt Blue'
Percentage anthers with
endothecium 100 100 100
degenerated tapetum 100 100 72.2
degenerated lumen contents 0 0 100
pollen grains 100 100 0
intine 100 100 0
bexine 100 100 0
germination pore 100 100 0
`Moorpark' showing no inherent problem in the `Trevatt' cultivar. Female sterility was due to multiple ovules and retarded development, and the clone was male sterile due to microspore degeneration and some failure in tapetal breakdown. The findings explain the lack of success achieved using pollinator bouquets and pollen mixtures, although occasional fruit were produced. This suggests that some fertile embryo sacs were formed despite none being observed by microscopy.
`Trevatt Blue' tree from which the budwood was taken and from which the trees were clonally propagated. Trees maintained for budwood are generally prevented from flowering to limit spread of pollen-borne viruses, so a mutation would not have been recognised until the progeny flowered. The magnitude of the problem suggests that a single mutation or mutational event affected both the female and male fertility of the clone.
Sterile plants can show both male and female sterility as many processes during microsporogenesis and megasporogenesis are under combined genetic control (Chaudhury, 1993). In particular, disruptions during meiosis result in mutations that can cause sterility of both male and female gametophytes (Reiser and Fischer, 1993). This research into the `Trevatt Blue' apricot is the only report of both male and female sterility occurring in the same variety of apricot but the phenomenon has been reported in other crops. Abnormal embryo sacs of the female gametophytes and failure in gametogenesis of both male and female gametophytes was the cause of unfruitfulness in the `Swan Hill' variety of olive (Rallo et al., 1981), and a gene has been identified in the male gametophyte of soybean which reduces male and female fertility due to failure of cytokinesis following meiosis (Kennell and Horner, 1985). Failure in meiosis is suggested in this study by the low number of embryo sacs with nuclei, and the lack of pollen grains.
Sterility of apricots has previously been attributed to factors other than a mutation, such as adverse environmental conditions (Eaton and Jamont, 1964). Multiple ovules, as described here, have been described in apricot (Egea and Burgos, 1995), and it was suggested that there may be a relationship between the number of ovules per ovary and temperature (Egea and Burgos, 1998). In other studies, abnormalities such as degeneration of ovules which had developed normally, ovules with twin nucellus and shortened integuments were reported (Burgos and Egea, 1994; Egea and Burgos, 1994). Some authors have found that male sterile apricot trees have shrunken anthers with little or no pollen (Nakanishi, 1983; Medeira and Guedes, 1991), but there are no previous microscopy studies.
particular cultivar or clone, so that a mutation in one will not result in widespread loss of yield.
Acknowledgements
The authors acknowledge Peter Burn for reporting the problem, John Zirilli for supply of the `Trevatt Blue' flowers, Jenny Witherspoon for comments on the problem and supply of the `Trevatt Knight' flowers and Mike Harms for assistance.
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