| Myxobacteria have been identified as one of the most promising producers of natural products and have the potential to be widely used in agriculture,biomedicine and environmental protection.Myxococcales,phylogenetically affiliated with the Deltaproteobacteria,are a class of gram-negative bacteria.The new classification standard has classified myxobacteria as phylum Myxococcota.Myxobacteria possess complex multicellular behavior and complex life history,such as density dependence of cell growth,development of multicellular fruiting bodies in the absence of nutrients,the group movement and "wolf pack" predation of cells.In addition,myxobacteria can produce abundant secondary metabolites and are another important bacterial resources for novel antibiotics after actinomycetes.Myxobacteria are widely distributed in a variety of habitats around the world,including terrestrial,marine and freshwater related habitats.In comparison,myxobacteria have the relatively less research in freshwater environment.Previously,we confirmed that freshwater habitats are also important habitats for myxobacteria,and by analyzing the phylogenetic relationship of myxobacteria from marine,terrestrial and freshwater environments,we speculated that salinity had an important influence on myxobacterial phylogeny.However,at present,the diversity analysis of myxobacteria pays more attention to a certain habitat,such as soil,marsh,composting,etc.,and the mining of myxobacterial resources also pays more attention to the terrestrial and marine environments,while the research on myxobacteria in freshwater habitats is less.Furthermore,the diversity and distribution of myxobacteria on a global scale,and the compare between different habitats are still poorly understood.Although myxobacteria are widely distributed,it is difficult to obtain the resources of myxobacteria,which is one of the important bottlenecks restricting its development and application.At present,the enrichment of myxobacteria is mainly based on their feeding properties by Escherichia coli and filter paper with targeted enrichment of bacteriolytic group and cellulolytic group,respectively.The isolation and purification mainly depend on their morphological characteristics to distinguish them from other microorganisms.To date,the myxobacteria described consist of three suborders:Cystobacterineae,Sorangiineae,and Nannocystineae.Many studies have indicated the existence of a large number of uncultured myxobacteria taxa,but the cultured and genome sequenced status of myxobacteria are still unknown.First,for a comprehensive and systematic understanding of myxobacterial diversity and global geographic distribution,as well as the extent of cultured and genome sequenced of myxobacteria.Here,through a comparative analysis of Earth microbiome project(EMP)data and myxobacterial information in public databases,we determined that the myxobacteria was one of the most diverse prokaryotic orders known on Earth,and preferred non-salt soil environments,such as plant rhizosphere and non-salt sediments.The annotation results showed that there were a large number of myxobacteria OTUs that had not been classified at the family(at least 20%)or genus level(at least 50%).Myxobacteria belonged to a group with low cultured rate and genome-sequenced rate,and their cultured rate and genome-sequenced rate varied greatly among different environments and different families/genus.In addition,myxobacteria with a high cell abundance or a wide distribution of samples were more likely to have been cultured or sequenced.However,even for the most environmentally superior myxobacteria OTU,more than 60%of them had not been isolated and cultured,and more than 70%had not yet been sequenced,indicating that there were still a large number of myxobacteria resources that had not yet been discovered.In summary,we demonstrated the global distribution of myxobacteria and evaluated the extent to which myxobacteria had been cultured and their genome sequenced for the first time.In view of the weak research status of myxobacteria in freshwater habitats,we used high-throughput sequencing technology to analyze the microbial diversity in lake surface sediments in Yunnan-Guizhou area to compare the differences in microbial community composition of different lakes and explore bacterial geographical distribution characteristics and its assembly mechanism in lake sedimentary environments.We found that in 10 lakes,the bacterial community structure of Chenghai Lake was significantly different from other lakes.By analyzing the correlation between the occurrence frequency and relative abundance of OTU in the samples,it was found that OTUs with higher occurrence frequency tended to have higher relative abundance,that is,high-abundance taxa had higher dispersion or adaptability.The correlation analysis between geographic distance and Bray-Curtis distance showed that the bacterial community structure in lake sediments did not conform to the distance-decay patterns,implying that even if the two lakes were far apart,due to the similar living environment,there was little difference in bacterial community structure.Through neutral model and null model analysis,we found that stochastic process was not the main ecological process that dominated the assembly of bacterial communities in Yunnan-Guizhou lake sediments,while homogenous selection in the deterministic process was the dominant process in the assembly of bacterial communities in lake sediments.At the same time,we selected the myxobacterial sequence for analysis,and found that the myxobacteria were widely present in all lake sediments,covering the three major suborders of myxobacteria,and there were also a large number of unclassified sequences.Among them,Haliangium and Anaeromyxobacter were the dominant groups in lake sediments.In the current omics era,pure culture of microorganisms is particularly important.Microbial resources can provide reliable genetic resources for omics research and verify various ecological functions and physiological metabolism predicted by omics research.Myxobacteria,as an important bacterial resources for novel antibiotics,grow slowly and are difficult to isolate and purify,which makes it difficult to obtain myxobacteria.For the exploration of myxobacteria resources,first of all,we summarized all the current cultured myxobacterial type species and their habitat sources and found that most of them came from soil related environment,only 6 species came from marine related habitats,and no one came from freshwater environment.Myxobacterial phylogeny showed that the cultured myxobacteria were divided into 3 suborders,10 families,30 genera and 75 species.Most of these type species were obtained by classical myxobacteria enrichment and isolation methods.This method is both effective and limited,that is,only a few common taxa can be obtained,and the discovery of new taxa is accidental.In order to explore the real enrichment effect of it,we used E.coli and filter paper for enrichment of a campus soil sample,sampled at different time points,and sequenced the microbial diversity.The analysis results showed that a large number of known myxobacteria groups were enriched,especially those that could be observed during isolation and purification,such as Myxococcus and Archangium.At the same time,a variety of uncultured taxa can also be enriched in large quantities,although they have not been cultured for the time being.While enriching myxobacteria,other groups of bacteria were also enriched,such as the predatory Bdellovibrionales and Burkholderiales with high motility and so on.Using the above-mentioned classical enrichment and isolation method for myxobacteria,we obtained more than 450 myxobacterial strains isolated from three habitats including land related environments(campus soil,the desert soil,trees),limnetic environment(lake sediments)and marine related environments(the Yellow River delta saline wetlands and transition zone sediments).The obtained myxobacteria were distributed in the 3 suborders of myxobacteria,including Myxococcus,Corallococcus,Archangium,Cystobacter,Stigmatella,Aggregicoccus,Pyxidicoccus,Vitiosangium,Sorangium,Polyangium,Nannocystis and Chondromyces a total of 12 genera,of which Myxococcus was the most isolates,including the currently known five species M.xanthus,M.virescens,M.fulvus,M.stipitatus and M.macrosporus,followed by the Coralcoccus including C.coralloides and C.exiguous.Six potential new species of myxobacteria were obtained.In addition,while isolating myxobacteria,9 new strains of other non-myxobacteria were obtained.The results of the isolation and culture of myxobacteria in the three major habitats are consistent with the above-mentioned diversity analysis results of myxobacteria enrichment.Myxococcus was the most easily enriched and isolated genus,and some of the taxa were enriched in large quantities but difficult to isolate and cultivate.The reasons for this remained to be further explored.In the process of excavating myxobacterial resources by the classical enrichment and isolation method,a number of potential new strains of bacteria were isolated,including new species of myxobacteria and other new species of non-myxobacteria.We selected one of the potential new species of myxobacteria,designated SDU3-1,isolated from soil of Shandong University campus.The strain belonged to the genus Polyangium of Polyangiaceae family.In addition,three new strains with obvious characteristics were selected,including a new species of Chryseolinea,designated SDU1-6,which is similar in morphology to myxobacteria and capable of gliding movement.Strain SDU3-2 with extreme irradiation resistance represented a new species of Deinococcus.Strain SDU3-3,which has only 84.2%similar to the 16S rRNA gene of type strains,is a new member of the class Chloroflexia in phylum Chloroflexi.From the aspects of morphological characteristics,phylogenetic relationships,physiological and biochemical characteristics,and genome information of these four potential new bacteria,and according to the characteristics of each strain,a new species of Polyangium aurulentus was established through a polyphasic taxonomy method,the representative strain is SDU3-1T.A new species of Chryseolinea flava is established and the representative strain is SDU1-6T.A new species of Deinococcus terrestris is established and the representative strain is SDU3-2T.A new family(Allapsifilum lubricum)of the class Chloroflexus,the representative strain is SDU3-3T.In summary,this study systematically analyzed the geographical distribution and diversity of myxobacteria from the perspective of culture-dependent and culture-independend approaches,and excavated new taxa of myxobacteria from various habitats to enrich the myxobacterial resource library.First,we exhibited the global geographic distribution and diversity of myxobacteria,and assessed the current cultured and genome sequenced status of myxobacteria.Secondly,given the weak research status of myxobacteria in freshwater environment,we analyzed the distribution and diversity of bacteria and myxobacteria in lake sediment environment,and analyzed the community assembly mechanism of bacteria in lake sediment environment.On this basis.we conducted resource mining of myxobacteria from three major habitats.We first summarized the phylogenetic relationships and their environmental sources of all cultured myxobacteria at present,and deeply understood the true enrichment effect of the classical enrichment methods for myxobacteria,and enriched and isolated myxobacteria from terrestrial related habitats,lake sediments and Yellow River Delta wetland related habitats.Finally,we selected one of the new myxobacteria strains and the other three strains with obvious characteristics to confirm their scientific taxonomic status by polyphasic taxonomy. |
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