[Analysis of Bacterial Communities and Antibiotic Resistance Genes in the Aquaculture Area of Changli County].

In order to understand the distribution of microorganisms and various antibiotic resistance genes in the aquaculture area of Changli County, Qinhuangdao, high-throughput sequencing technology was used in this study. We utilized 16S rDNA gene sequencing and metagenome sequencing methods to analyze the seawater, sediment, and gut contents of the local fish Synechogobius hasta in the aquaculture area in spring. The results showed that Proteobacteria, Firmicutes, and Bacteroidota were the dominant bacteria in seawater; and Proteobacteria, Crenarchaeota, Acidobacter, and Actinobaciota were rich in the sediment; whereas Proteobacteria, Cyanobacteria, Firmicutes, and Bacteroidota were in relatively high abundance in fish gut contents. The microbial diversity of sediment samples was the most abundant, followed by seawater samples, and the microbial diversity of fish intestinal contents was the lowest. Moreover, the microbial diversity of similar samples was relatively similar, and the microbial diversity of different types of samples was quite different. For samples at different sites, there were significant differences between seawater samples at each site, and there were small differences between sediment samples at each site, and some sediment sample groups did not have significant differences in microbial composition. In all sample groups, five ß-lactam antibiotic resistance genes (blaOXA-325, cepS, blaCARB-20, blaOXA-55, and blaTRU-1) and four aminoglycoside antibiotic resistance genes[aac(6')-IIb, amrA, aac(6')-Ie-aph(2″)-Ia, and aph(3')-Vc] were detected. There was also a certain correlation between antibiotic resistance genes and microbial communities.

Mixed oligosaccharides-induced changes in bacterial assembly during cucumber (Cucumis sativus L.) growth.

The application of oligosaccharides can promote plant growth by increasing photosynthesis or inducing plant innate immunity. However, the mechanisms by which oligosaccharides affect bacterial community diversity and abundance remain unclear. In this study, a mixed oligosaccharide was applied to the growth of cucumbers. The findings of the present study suggest that the application of MixOS has significant effects on the bacterial communities in the phyllosphere, rhizosphere, and bulk soil of cucumber plants. The treatment with MixOS resulted in delayed senescence of leaves, well-developed roots, and higher fruit production. The bacterial diversity and composition varied among the different ecological niches, and MixOS application caused significant shifts in the bacterial microbiome composition, particularly in the phyllosphere. Moreover, mixed oligosaccharides increased the abundance of potential growth-promoting bacteria such as Methylorubrum spp. and Lechevalieria spp., and more zOTUs were shared between the WM and MixOS treatments. Furthermore, the bacterial co-occurrence network analysis suggested that the modularity of the phyllosphere networks was the highest among all samples. The bacterial co-occurrence networks were altered because of the application of MixOS, indicating a greater complexity of the bacterial interactions in the rhizosphere and bulk soil. These findings suggest that mixed oligosaccharides has the potential to improve plant growth and yield by modulating the bacterial communities within and outside the plants and could provide a theoretical basis for future agricultural production.

Generation of Fluorinated Amychelin Siderophores against Pseudomonas aeruginosa Infections by a Combination of Genome Mining and Mutasynthesis.

Pioneering microbial genomic surveys have revealed numerous untapped biosynthetic gene clusters, unveiling the great potential of new natural products. Here, using a combination of genome mining, mutasynthesis, and activity screening in an infection model comprising Caenorhabditis elegans and Pseudomonas aeruginosa, we identified candidate virulence-blocking amychelin siderophore compounds from actinomycetes. Subsequently, we developed unreported analogs of these virulence-blocking siderophores with improved potency by exploiting an Amycolatopsis methanolica strain 239T chorismate to salicylate a biosynthetic subpathway for mutasynthesis. This allowed us to generate the fluorinated amychelin, fluoroamychelin I, which rescued C. elegans from P. aeruginosa-mediated killing with an EC50 value of 1.4 µM, outperforming traditional antibiotics including ceftazidime and meropenem. In general, this paper describes an efficient platform for the identification and production of classes of anti-microbial compounds with potential unique modes of action.

Dual-function chromogenic screening-based CRISPR/Cas9 genome editing system for actinomycetes.

Actinobacteria are one of the most important sources of pharmaceutically valuable and industrially relevant secondary metabolites. Modern genome mining reveals that the potential for secondary metabolite production of actinomycetes has been underestimated. Recently, the establishment of CRISPR/Cas9-based genetic manipulation approaches in actinomycetes opened a new era for genome engineering of this type of organism. Compared with the traditional methods, the application of CRISPR/Cas9 shows several advantages in actinomycetes including higher efficiency and ease of operation. However, the screening process for the correctly edited mutants and the plasmid curing are still time- and labor-intensive. To address this problem, we developed an updated version of the CRISPR/Cas9 genome editing system for actinomycetes, based on two chromogenic reporter systems (GusA and IdgS). Our system facilitates both processes of positive clone screening and plasmid curing. Here, we demonstrate by three case studies in both model actinomycetes and non-model actinomycetes that this system is faster and more efficient. We performed the deletion of one single gene, actIORFI (SCO5087 of the actinorhodin gene cluster) in Streptomyces coelicolor M145, one small-size (5.5 kb) gene cluster (orange-pigmented carotenoid gene cluster), and one relatively large-size (61 kb) gene cluster (abyssomicin gene cluster) in Verrucosispora sp. MS100137. The results presented in this study indicate that this updated CRISPR/Cas9 system employing chromogenic reporters is versatile and broadly applicable in genome engineering of actinomycetes, not only for the largest genus Streptomyces.

Armeniaspirol Antibiotic Biosynthesis: Chlorination and Oxidative Dechlorination Steps Affording Spiro[4.4]non-8-ene.

Armeniaspirols are potent antibiotics containing an unusual spiro[4.4]non-8-ene moiety. Herein, we describe the cloning and functional analysis of the armeniaspirol biosynthetic gene cluster. Gene-inactivation studies and subsequent isolation of previously unknown biosynthetic intermediates shed light on intriguing biosynthetic details. Remarkably, deletion of ams15, which encodes a protein bearing a flavin-binding domain, led to the accumulation of several non-spiro intermediates with various numbers of chlorine substitutions on the pyrrole moiety. The di- and trichloropyrrole species were converted by Streptomyces albus expressing Ams15 into mono- and dichlorinated spiro derivatives, respectively. In addition, in vitro conversion of these non-spiro intermediates into des-N-methyl spiro intermediates by the cell lysate of the same recombinant strain proved Ams15 to be responsible for spiro formation through oxidative dehalogenation.

[Development of a method for cleaning outpatient data rapidly and generating statistical reports automatically to the analysis of time series on the air pollution and disease].

OBJECTIVE: To develop a method for cleaning the outpatient data rapidly and generating the statistical reports automatically. METHODS: Formulating the cleaning rules according to the data characters, writing programs to clean the individual cases and generate the statistical reports using the SQL language. RESULTS: It could clean the different individual cases rapidly, calculate the daily outpatient visits and generate the statistical reports automatically with high accuracy. CONCLUSION: The method can apply to the data processing of the hospital cases, first aid cases, cause-of-death data and health records. It not only can process large amounts of data flexibly, conveniently and quickly, but also has great practical value. So it is the necessary way to the health risk assessment of air pollution.

A new salicylate synthase AmS is identified for siderophores biosynthesis in Amycolatopsis methanolica 239(T).

Siderophores are important for the growth of bacteria or the applications in treatment of iron overload-associated diseases due to the iron-chelating property. Salicylate synthase played a key role in the biosynthesis of some NRPS-derived siderophores by the providing of an iron coordination moiety as the initial building block. A new salicylate synthase, namely AmS, was identified in the biosynthesis pathway of siderophore amychelin in Amycolatopsis methanolica 239(T), since it shunt chorismate, an integrant precursor, from primary to secondary metabolite flow. The amino acid sequence alignment and phylogenetic analysis showed that AmS grouped into a new cluster. In vitro assays of AmS revealed its wide temperature tolerance ranged from 0 to 40 °C and narrow pH tolerant ranged from 7.0 to 9.0. AmS was resistant to organic solvents and non-ionic detergents. Moreover, AmS converted chorismate to salicylate with K m of 129.05 µM, k cat of 2.20 min(-1) at optimal conditions, indicating its low substrate specificity and comparable velocity to reported counterparts (Irp9 and MbtI). These properties of AmS may improve the iron-seizing ability of A. methanolica to compete with its neighbors growing in natural environments. Most importantly, serine and cysteine residues were found to be important for the catalytic activity of AmS. This study presented AmS as a new cluster of salicylate synthase and the reaction mechanism and potential applications of salicylate synthase were highlighted as well.

[Appropriate dust control measures for jade carving operations].

OBJECTIVE: To provide the appropriate dust control measures for jade carving operations. METHODS: Dust concentrations in the workplace were measured according to GB/T 5748-85. Ventilation system of dust control were measured according to GB/T 16157-1996. Dust particle size distributions for different sources and particle size fraction collecting efficiencies of the dust collectors were measured with WY-1 in-stack 7 stage cascade impactors. RESULTS: On the basis of adopting wet process in the carving operations, local exhaust ventilation system for dust control was installed, which included: the special designed slot exhaust hoods with hood face velocity of 2.5 m/s and exhaust volume of 600 m3/h. The pipe sizes were determined according to the air volume passing through the pipe and the reasonable air velocities. Impinging scrubber or bag filter dust collector were selected to treat the dust laden air from the local exhaust ventilation system, which gave a total collecting efficiency of 97% for impinging scrubber and 98% for bag filter; The type of fan and its size were selected according to the total air volume of the ventilation system and maximum total pressure needed for the longest pipe line plus the pressure drop of the dust collector. CONCLUSIONS: Practical application showed that, after installation and use of the appropriate dust control measures, the dust concentrations in the workplaces could meet or nearly meet the national hygienic standard and the dust laden air at the local exhaust ventilation system could meet the national emission standard.