# Dynamics of Microbial Community Structure and Metabolites during Mulberry Ripening
## 3 Results and Discussion
### Dynamic Changes in Community Structures in WHDS during Ripening  
There is a close correlation between microbial community structure and plant growth. The high-throughput sequencing of micro-organisms at different maturation stages of WHDS was performed using the Illumina MiSeq PE300 microbial detection platform to determine the succession of micro-organisms on its surface. The test results yielded a total of 521,485 valid sequences, with an average of 104,297 sequences per sample and a total of 1647 OTUs divided by 97% similarity, which were mainly concentrated in the interval length of 361-380 bp. The statistics of the number of taxonomic units after OTU clustering were 1 domain, 1 kingdom, 12 phyla, 19 orders, 56 orders, 101 families, 190 genera, and 242 species. As shown in Figure 6a, the bacterial communities in the different maturation stages of WHDS consisted of _Proteobacteria_, _Actinobacteria_, _Firmicutes_, _unclassified norank Bacteria_, _Bacteroidota_, _Myxxoccoccta_, _Deinococcota_, _Chlorofexi_, _Verrucomicrobiota_, _Abbitibacteri_, and Others. Among them, _Proteobacteria_, _Actinobacteria_, _Firmicutes_, and _unclassified norank bacteria_ were the dominant genera and equally present in other fruits [59; 60], while the relative abundance of the other 12 phyla was below 1%. It can also be seen in Figure 6a that the relative abundance of _Proteobacteria_ showed a significant increasing trend with increasing maturity, from 10.82% to 65.89% and, finally, 75.51%. The relative abundance of _Actinobacteria_ decreased from 52.14% to 11.78% and that of _Firmicutes_ from 35.68% to 11.30%, with both abundances decreasing significantly. _Unclassified norank bacteria_ varied less throughout the stage, at around 1%. It is worth noting that _unclassified norank bacteria_ have been less studied in previous articles, and their specific functional properties and metabolic mechanisms need to be further examined. In addition, as can be seen in Figures 6b,c, S3 and S5 clustered into one group, and the remaining three stages clustered into one group. A total of 123 bacterial genera were present in S1, 105 bacterial genera were present in S2, 104 bacterial genera were present in S3, 112 bacterial genera were present in S4, and 118 bacterial genera were present in S5, with no regular trend in the number of genera. The common dominant bacterial genera were _Bacillus_ (9.1%-34.95%), _Brewhibacterium_ (6.46%-38.94%), _Pseudomonas_ (1.25%-42.16%), _Tatumella_ (0.18%-37.15%), _Corynebacterium_ (2.21%-11.19%), _Pantoea_ (0.69%-12.12%), _Ammonophilus_ (1.22%-5.29%), _Unclassified enterobacterales_ (0.66%-7.07%), _Unclassified gammaproteobacteria_ (0.49%-4.44%), and _Brachybacterium_ (1.07%-2.13%). _Bacillus_ changes in relative abundance as it increases in maturity, first increasing and then decreasing. Some species of _Bacillus_, like _Bacillus cereus NRKT_, are important biological control bacteria to reduce the fruit ripe rot disease caused by _Colletotrichum gloosporoides_[61], and these genera have been  
Figure 6: Microbial community (**a**), heat map (**b**), and Venn diagram (**c**) at different maturity stages of WHDS. Note: the numbers in (**c**) indicate the species of strains common to certain stages. detected in soil where fruits are grown [62]. The trend of _Bacillus subtilis_ was similar to that of _Bacillus_. _Bacillus subtilis_ is an effective genus of biocontrol organisms [63], and spraying it as a solution on fruit vines was effective in increasing the alcohol and ester content of the corresponding fruit wines [64]. The abundance of _Pseudomonas_ showed an increasing trend, reaching a maximum at S5. _Pseudomonas_ can have beneficial effects on plants, such as nitrogen fixation and phosphorus solubilisation, as well as in other diseases, such as spotting [65; 66]. _Corynebacterium_ uses glucose to produce methyl anthranilate, which significantly enhances the flavour of fruits [67].