Studies On The Life Cycle And Classification Of Agrocybe Salicacola

In this study, the life cycle and classification of Agrocybe salicacola was deeply researched from four parts:mating type, relationship between mating type factors and mycelial growth rate, interspecific hybridization and molecular systematics.Basidiospores of A. salicacola YAASM0711 were collected from mature fruiting body of by ejection.224 single-spore strains were isolated respectively by dilution separation method,210 were identified as monocaryon. The result of mating test suggested that the sex-inheritance of A. salicacola is tetrapolar heterothallism. Four groups of strains belonging to different mating-types were obtained by mating tests, and the ratio of monocaryotic strains corresponding to A1B2, A1B2, A2B1 and A2B2 was 47:59:53:51. The results showed that the number and distribution of its mating types was related with spore germination and growth rate. In all strains, the growth rate of one mating type strains was more slow than other three, and only there were 3 and 5 slow-growing strains in other two mating type strains. Especially, although the growth rates were identical between the Fast-Fast mating heterocaryotic strains and Fast-Slow mating heterocaryotic strains in YPD plate, there were visible difference in polypropylene mushroom cultivation bags. We concluded that the growth rate genes of mycelia may be linkaged with mating factor A and B, and genetic recombination occurred in the basidiospores formation.Because Fast-Slow mating heterocaryotic strains had advantages in the growth, most scientists may select Fast-Slow mating heterocaryon in breeding of mushroom, so distorted segregation of mating types may happen.To further test the genetic model of A. salicacola and the relationship between the different strains of Agrocybe, the mating types of strain YAASM0962, YAASM0969 and cultivation strain CS45 of A. aegerita were studied according to the method of upper research in YAASM0711, and the result showed the sex-inheritance of three strains was tetrapolar heterothallism. The ratio of monocaryotic strains corresponding to A3B3, A3B4, A4B3, A4B4 was 6:10:14:7 in YAASM0962, and there was only two mating-types in YAASM0969 with a ratio of 38:35 for A5B5:A6B6. Similarly, only two mating-types occurred in CS45, and the ratio of A7B7:A8B8 was 51:55. The hybridization result showed YAASM0711 could form dicaryon with tested strains YAASM0962, YAASM0969, YAASM0967 and CS45, and produce fruiting bodies except CS45, which further A. salicacola was a independent species in morphological and biological senses. Because there were multiple alleles of mating factor caused by the geography differences, any monokaryotic strains of YAASM0711 could produce fruit bodies after mating with any monokaryotic strains of YAASM0962, YAASM0969 and YAASM0967. In the progress of natural mates and cultivation, some recombination, insertion or deletion events constantly occurred in the locus of mating gene, which caused segregation distortion. So reduction of subunit of mating gene could lead to the ration imbalance of mating types of strain (for example YAASM0962), even the lost of some mating types (for example YAASM0969 and CS45) and artificial selection by mushroom cultivation may enhance this phenomena.As one of delicious edible mushroom, A. aegerita was widely cultivated in China. Its diversified names, and the complexity as probable multispecies complex (A. aegerita, A. cylindraceae, A. chaxingu, A. salicacola etc), brought many troubles to the researchers. On the basis of wide collection of test strains in this study, their internal transcribed spacer (ITS) and 18S rDNA were sequenced, and analyzed systematically combined with the existing GenBank sequences to establish the taxonomy of them. The results showed the test strains were divided into two parts, and congregated together compared with other species of Agrocybe geneus in the phylogenetic tree. The hybridization result further confirmed that A. salicacola was an independent biological species different from A. aegerita. In addition, high diversity happened in the A. aegerita strains, which implied A. aegerita was the multispecies complex. In conclusion, we could distinguish the mixed A. aegerita based on the rDNA sequences with the hybridization analysis, which provide a convenient and accurate way for the resource collection and breeding.Totally, A. salicacola was an independent species from A.aegerita, and the sex-inheritance was tetrapolar heterothallism. We speculated that there are some new species, sub-species or variation in the genus Agrocybe, and further researches needed to test this conclusion.