In this article we will discuss about the cytology of ascus development, explained with the help of suitable diagrams.
Opinions and interpretation are so conflicting that it is rather difficult to take a clear view with regard to the cytology of ascus development. In spite of everything in attempt is being made to give a picture of the whole situation as clearly as possible asking into consideration of the viewpoints of most of the workers.
The phenomenon of nuclear fusion in the Ascomycetes was reported for the first time by Dangeard in 1894. He observed the formation of a crozier at the tip of ascogenous hypha in Peziza Hsiculosa.
The two nuclei of the subterminal cell or loop cell of the crozier fuse and the subterminal cell elongates forming an ascus. Dangeard’s findings have been widely confirmed by many investigators.
In 1895 Harper reported that in Sphaerotheca humuli exual reproduction is initiated by gametangial contact when a passage is developed it the point of contact of the gametangia through which the male nucleus passes into the ascogonium where it fuses with the female nucleus. Immediately after nuclear fusion septa appear in the ascogonium resulting in the development of a row of cells.
The binucleate penultimate cell of which develops into an ascus. Harper stated that there are two nuclear fusions, one between the male and female nuclei in the ascogoaium and the other in the ascus, hence the ascus nucleus is tetraploid. Harper’s account indicates the two chromosome reductions are necessary during the nuclear divisions in the ascus (Fig. 201).
In 1906 Blackman and Fraser (nee Gwynne-Vaughan) described a case of reduced sexual process in Humaria granulata, a self-sterile species. Here antheridium is not developed. A multinucleate ascogonium without any trichogyne is present. The female nuclei fuse in pairs in the ascogonium and the diploid nuclei so formed enter in the ascogenous hyphae that are developed from the ascogonium.
A second nuclear fusion takes place in the ascus.
In 1907 Dangeard and his followers working on Pyronema confluens reported that there is development of antheridium and ascogonium bearing trichogyne. But antheridium and trichogyne do not fuse and the septum at the base of the trichogyne remains intact as a result of which the antheridial nuclei fail to reach ascogonium.
The trichogyne and the antheridium ultimately disintegrate. The ascogonial nuclei arrange themselves in pairs in the ascogonium. The paired nuclei then pass into the ascogenous hyphae and eventually ascus and ascospores are formed by crozier formation. Dangeard denied the occurrence of a nuclear fusion in the ascogonium, but admitted one nuclear fusion and one reduction division that too are in the ascus (Fig. 202).
Gwynne-Vaughan (nee Fraser) in 1908 stated that in Humaria rutilans there are two nuclear fusions and two reduction divisions in the ascus.
Claussen working on Pyronema confluens in 1912 said that the male and the female nuclei pair closely in the ascogonium but they do not fuse. The paired nuclei then enter in then ascogenous hyphae where the ascus is developed by crozier formation, and nuclear fusion takes place in the ascus.
Meiosis of the diploid nucleus takes place immediately after nuclear fusion. According to him, there is one nuclear fusion and only one meiosis in the entire process of sexual reproduction leading to the development of ascospores and that too in the ascus (Fig. 203).
Dodge in 1920 described the life history of Ascobolus magnificus. According to him, a large septate ascogonium wraps about a club-shaped antheridium. The antheridial tip fuses with the tip of the trichogyne. Ascogenous hyphae develop from the ascogonium. Simultaneously numerous sterile hyphae grow from the neighbouring cells producing a closed structure. Dodge did not enter into nuclear details.
Bagchee (1925) working on Pustularia bolarioides reported that the number of chromosomes in the third telophase in the ascus is the same as the first.
Schweizer in 1931 described the development of ascus and ascospores in Ascobolus strobilinus. Here each fructification has sex organs. The male nuclei pass through the unicellular trichogyne to the ascogonium, where they pair with the female nuclei, but do not fuse. The male nuclei can be recognized by their small size. He said that the ascogenous hyphae also contain paired nuclei, one small male nucleus and one large female nucleus. But he did not go any further.
Gwynne-Vaughan and Williamson in 1931 found that in Pyronema confluens the trichogyne and the antheridium make contact. The nuclei of the trichogyne degenerate and the wall at the base of the trichogyne ruptures. Hence a continuous passage is developed from antheridium to ascogonium through trichogyne by the dissolution of the walls at the point of contact of the trichogyne and the antheridium.
The male nuclei pass from the antheridium through the trichogyne to the ascogonium and mix with the female nuclei there. The male and female nuclei fuse in pairs in the ascogonium and the diploid nuclei so formed then pass into the ascogenous hyphae. Asci are formed by typical crozier formation and a second nuclear fusion takes place in the ascus resulting a tetraploid nucleus.
During the development of ascospores the tetraploid nucleus undergoes three successive divisions to produce eight haploid nuclei. The first and the third divisions are both reductional and the second one is mitotic (Fig. 204).
The second reduction division that takes place in the ascus has been termed brachymeiosis. Their statements were supported by chromosomal counts at every stage of nuclear division. They reported the chromosome number of the tetraploid nucleus as 24 which reduces to 12 immediately after first division.
Whereas, in the second division the chromosome number remains 12 which again is reduced to a haploid number of 6 as a result of the third nuclear division which is also reductional.
Gwynne-Vaughan and Williamson in 1932 investigated on Ascobolus magnificus and confirmed the findings of Dodge. They were the first to report that a continuous passage is developed from antheridium through trichogyne to the ascogonium along which antheridial nuclei pass into the ascogonium.
The nuclei of the antheridium possess it large chromatin body and a small stainable granule. Whereas, the ascogonial nuclei show a small rounded chromatin body. The nuclei bear 4 chromosomes. Fusion of male and female nuclei takes place in the ascogonium.
The diploid nuclei then pass into ascogenous hyphae that are developed from the ascogonium. They identified the diploid nature of the nuclei by chromosome counts. The chromosome number of the diploid nuclei is 8, along with the chromosomes, the large and the small chromatin bodies were also present in the diploid nuclei.
The binucleate apical cells of the ascogenous hyphae produce asci by crozier formation, when fusion of the two diploid nuclei results a tetraploid nucleus in the ascus having 16 chromosomes.
During the first division of the ascus nucleus 8 bivalents are visible and 8 chromosomes travel to each pole. In the second division also 8 chromosomes are seen. In the third division prophase 4 recurved and elongated bodies are seen. 4 chromosomes pass to each pole, so that the ascospore nuclei are haploid with 4 chromosomes.
The first division in the ascus is reductional, second educational, and the third division is also reductional which they called brachymeiosis.
In 1937 Gwynne-Vaughan reported two nuclear fusions and two reduction divisions in Patella (Lachnea) melaloma (Alb. and Schw.) Seaver.
The whole idea of two nuclear fusions and two reduction divisions was strongly criticized by many workers resulting into difference of opinion. To a large number of investigators it appeared very unusual to have two successive fusions and two reduction divisions in the same life cycle.
From time to time there has been debate concerning the number of nuclear fusions and reductional divisions in the life cycle of certain members of the Ascomycetes. The solution of problem of this kind is necessary before the investigators can arrive at a true insight into the interrelationships and evolutionary tendencies of the group.
Olive in 1949, 1950 using propino-carmine staining technique demonstrated that there was no evidence of brachymeiosis in Patella melaloma which has previously been reported by Gwynne-Vaughan in 1937. He clearly worked out that there is one nuclear fusion in the young ascus giving rise to a diploid nucleus which is followed by a single reduction division.
Hirch in 1950 using techniques especially devised for staining chromosomes stated that there is no evidence that brachymeiosis occurs in Pyronema confluens, as was claimed by Gwynne-Vaughan and Williamson in 1931. He stated that the young ascus nucleus is diploid and not tetraploid with 24 chromosomes as claimed by them.
A single reduction division occurs, in the young ascus, and the haploid number of chromosome is 12.
In 1953, working on the cytology of ascus development in Ascobolus magnificus Wood reported that asci arise in the usual fashion from the binucleate penultimate cells of the croziers. There is a conjugate division in the ascogenous hyphae. The darkly staining spots or granules present in the ascus nucleus may be heterochromatin in nature. The number of bivalents in the first division metaphase is eight.
The diploid number of chromosome in the fusion nucleus of the ascus therefore is sixteen. Following first division metaphase the homologous chromosomes separate. During first division anaphase eight univalent chromosomes pass to each spindle pole two daughter nuclei with their nucleoli are formed. At prophase and metaphase of the second division there appears to be 8 chromosomes in each nucleus.
During second division anaphase 8 chromosomes pass to each pole. At the end of the second division four nuclei with four promonent nucleoli are present in the ascus. It would appear from Wood’s work that the haploid number of chromosomes in Ascobolus magnificent eight. No evidence of brachymeiosis is found.
The evidence contradicts the claims of Gwynne-Vaughan and Williamson that the haploid number is four and that the fusion nucleus is tetraploid. The deeply staining bodies claimed by Gwynne-Vaughan and Williamson in the fusion nucleus of the ascus may be two heteropyknotic regions of a pair of chromosomes.
Wood’s observations were mainly on the nuclear behaviour in the ascus nuclei. In the ascus, the nucleus is a prominently vacuolate body. The number of bivalent chromosomes at metaphase I is eight. The diploid number of chromosomes in the fusion nucleus is sixteen. Following metaphae I the homologous chromosomes separate.
During anaphase I eight univalent chromosomes pass to each pole. The two daughter nuclei with their nucleoli are formed.
Hence no evidence of brachymeiosis has been found in Patella melaloma, Pyronema cdnsfluens, or in Ascobolus magnificus. The whole idea of brachymeiosis would now be unacceptable.
In general, Claussen’s findings have received wider support being more reasonably acceptable than those of other workers.
Those who still hold that there occur two successive nuclear fusions followed by a double reduction division could very well settle this question by a genetic study of the species of Ascobolus. With only a single nuclear fusion and that one in the ascus, there cannot be normally more than four genotypically different kinds of ascospores in an ascus.
If the spores are arranged in one line they will be found to alternate 4 and 4; and 2 or 2, 4, 2 for any pair of genetic factors. With double fusion and double reduction division, one should find many asci with eight different kinds of spores. If the spores are in linear arrangement they should very frequently alternate 1 and 1 for members of a pair of factors, such, for example, as mating type factors.