| In today’s growing bacterial resistance, strengthen research and development of new drugs has a very important practical significance. Actinomycetes are an important source of new antimicrobial drugs. For a long time, the research of actinomycetes natural medicine mostly concentrated on soil actinomycetes. However, as exploitation and utilise of actinomycetes resource increasingly reached the limits, the chances of finding new drugs or lead compounds greatly reduced. The active metabolites isolated more than 95% are all known compounds. Therefore, broaden actinomycetes research, looking for new actinomycetes biological resources from different sources is the only way. Since 1960 s, marine microbes and bacteria in medicine plant is becoming a hot spot for raw drug development. So far,20,000 natural products have been found from them. However, research on insect microbial natural products saw sporadic reports. Insects are the largest group of organisms in nature,accounting for more than 85% of the whole species, vitality is very tenacious, and a large number of endophyte insect mutualism, coevolution, to resist the invasion of pathogens.American cockroach is a traditional Chinese herbal medicine, has anti-bacterial, anti-tumor,anti-inflammatory effects and so on. American cockroach exists in the Earth for more than200 million years, still going, has become the dominant populations interior, mainly living in the ditch heating, toilet, shower and other dark and humid environment. An increasing number of reseach on American cockroach pharmacological effects by modern medicine,but research about American cockroach endophytic actinomycetes diversity and applied is still less. This paper intend to study the American cockroach gut actinomycetes diversity, to fully understand the medicinal value and biological characteristics.Obstacle This paper regards the wild American cockroach as the research object, isolated intestinal endophytic actinomycetes. Using morphological and molecular biology methods analysis these actinomycetes’ classifying, identification and phylogenetic. And among them,13 strains of gordonia’s physiological and biochemical were researched. It is desirable to provide a new microorganism resources for drug development through the biological diversity study of American cockroach intestine actinomycetes.Method1. Periplaneta Americana intestinal endogenous actinomyces separation Choosing appropriate antibacterial agent, taking agar plate dilution method, to isolate blatt intestinal actinomycetes. Morphology observation and 16 S r DNA amplification product Blast sequence alignment and DNAman 6.0 sequence alignment were utilized to study their classification and identification, using Mega 5.1 software, the maximum likelihood method, constructing phylogenetic tree to analyze it.2. The physiological and biochemical research of 13 gordonias Morphology observation, 16 S r DNA amplification product Blast sequence alignment and DNAman 6.0 sequence alignment were utilized to study their classification and identification. Through Gen Bank Bank It software, applying 13 gordonias gene sequence number. Then, Staphylococcus aureus, Methicillin-resistant staphylococcus aureus, E. coli,Klebsiella pneumoniae and Bacillus subtilis five kinds of bacteria and Candida albicans,Aspergillus fumigatus, Aspergillus niger, Trichophyton and other fungi were used for bacteria to detect 13 gordonias initial bacteriostatic activity of fermented liquid. And through the carbon source utilization experiment, cellulose, starch, biological macromolecules, the pesticide main active ingredient degradating, hydrolysis experiments and heavy metal ions adsorption experiment to study its physical and chemical properties.Result1. 159 endophytic actinomycetes strains had been isolated from 13 periplaneta Americana intestinal, morphological characteristics conform to the classic characteristics of actinomycetes, 16 S r DNA amplification product Blast sequence alignment showed that they belonged to 10 families, 12 generas. Among which Streptomyces and Gordonia were dominant generas, respectively, 67.3%, 8.2% of the total number of isolates. DNAman 6.0sequence alignment showed that the similarities of 149 strains were lower than 98.2%compared to the known actinomycetes, which showed that they may be new strains,accounting for 93.7% of the total number of isolates.2. DNAman 6.0 sequence alignment results showed that the similarities of 13 gordonias were lower than 98.2% compared to the known strains, which indicated that 13 gordonias are possible the new strains. After application, 8 strains gene sequence numbers were obtained. Physiological and biochemical characteristics experimental results showed that there were 4 stains fermentation liquor had inhibition effect to Staphylococcus aureus,Methicillin-resistant staphylococcus aureus and E. coli, 8 strains to Bacillus subtilis; 3strains to Candida albicans and Trichophyton, only 1 strain to Klebsiella pneumoniae and Aspergillus niger, found no inhibition effect to Aspergillus fumigatus; 13 gordonias had different absorption utilization degree to different carbon source; All could degrade cellulose and hydrolysis starch, but the degradation and hydrolysis ability were not strong;having strong adsorption to copper, lead, mercury and other heavy metal ions, strong degradation to pyrene, anthracene, phenanthrene and other biological macromolecules, and pesticides main active ingredients.Conclusion There are rich actinomycetes resources in periplaneta Americana intestinal, which can provides a new microbial resource for new drug research and environmental governance. |
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