Abstract:It is estimated that the majority (>99%) of microorganisms resist cultivation because they are in a viable but non-culturable (VBNC) state. Resuscitation-promoting factors (Rpfs) are one of the most important breakthroughs in the cultivation of VBNC bacteria. In the past decade, our group focused on resuscitation of VBNC bacteria by Rpf for exploring their potential environmental functions. Incorporated our previous studies in this mini-reiew, we provided the overview of formation and resuscitation of VBNC bacteria, and explored the relationship between "scout cells" and quorum-sensing based on the process of formation and reversion of VBNC cells. Moreover, we also summarized the VBNC bacteria possessed potential enivronmental functions which were previously isolated from various environments with Rpf addition. This review provides not only a new insight into revealing the the mechanisms of formation and resuscitation of VBNC bacteria, but also a theoretical basis for understanding and re-evaluating the role of Rpf method in the application for environmental bioremediation.

Abstract:LC3-associated phagocytosis (LAP) is an efficient process for the phagocytosis and degradation of invading pathogens. Recently, many studies show that LAP plays a very important role in the elimination of pathogenic microorganism, and its mechanism is different from the conventional phagocytosis and autophagy. Upon the extracellular stimulation, the activation of some autophagy related proteins in host cells induces the recruit of LC3 to the single membrane of phagosome, which promotes the efficiency of phagocytosis and killing of the pathogens. Different pathogens have different mechanisms to cope with the killing of LAP. The general mechanism of LAP, different responses to LAP to various pathogens and the research progress in recent years are reviewed in this paper.

Abstract:Corynebacterium pseudotuberculosis is an interesting bacterium due to its great zoonotic potential and because it causes considerable economic losses worldwide. The rapid development of omics technology has promoted the study of C. pseudotuberculosis pathogenicity. A total of 72 C. pseudotuberculosis genome sequences were obtained by genome sequencing analysis, and the structure, evolution and pathogenicity differences of C. pseudotuberculosis biovar equi or biovar ovis were identified. Based on the proteomics study, proteins such as phospholipase D (PLD), serine protease (CP40) and neuraminidase H (NanH) were found to be related to C. pseudotuberculosis physiology, toxicity and immunity, and other unknown functions. With the transcriptomics, it was found that defense against oxidative stress, adhesion and regulation were most typical in the gene expression difference in harsh environments such as high osmotic pressure, heat shock or acidic conditions. In this paper, we analyzed the C. pseudotuberculosis Xuanhan strain (XH02) we obtained with comparative genomic and SNP evolution, and the application of proteomics and transcriptomics in C. pseudotuberculosis pathogenesis was reviewed.

Abstract:Microbial electricigenic respiration is a microbial metabolism mode in which electroactive microorganisms (EAMs) degrade organics and then transfer the produced electrons to extracellular solid electrodes through the respiratory chain, while EAMs gain energy to support their own growth. Microbial electricigenic respiration has broad application prospects in renewable energy utilization and environmental remediation. How to improve the microbial electricigenic respiration performances of EAMs is at the heart of promotion of relevant techniques from lab-scale research to full-scale application, and the electron transfer between EAMs and extracellular solid electrodes plays a pivotal role in microbial electricigenic respiration. To date, there have been many reviews focusing on how to improve electrode materials, however, very few literatures summarize how to strengthen the electricigenic respiration capability of EAMs. In this review, we summarized the existing approaches for promoting the respiratory performances of EAMs from the following three aspects:the addition of chemical reagents for example surfactants and signal molecules, physical force application such as ultrasonic waves and magnetic field forces, and genetic modification for instance phenazine secretion-related gene overexpression and pilA gene heterogenous expression. Advantages and disadvantages of each method were introduced, mechanisms and effects were expatiated upon, and future research directions from the perspectives of the practical application and the mechanism research were proposed.

Abstract:Avian Tembusu virus (ATMUV) is a highly pathogenic virus infecting variety kinds of laying birds, resulting in appetite decreases and significant reduction in egg production. At the last stage, the affected animals may have neurological symptoms, such as muscle paralysis and dystaxia. In recent years, ATMUV spreads quickly in many provinces in China and has caused huge economic loss in poultry industry all over the world. Host innate immunity is the first line to defend against the infection of pathogens at early stages. Innate immune response is the nature ability of the organism to fight against the pathogenic microorganisms. On the other hand, adaptive immunity is the process by which the immune system produces specific antibodies by B cells and cellular immune response by T cells. Adaptive immunity is the key components of host immune system that contribute to the resistance to pathogens and are the basis of immunization. In this review, we will discuss host immune response to ATMUV infection and highlight the mechanisms by which ATMUV induces the host innate and adaptive immunity.

Abstract:[Objective] We studied the influence of three factors (region, gender and season) on intestinal bacterial community structure and diversity of Gansu zokor distributed in Liupan Mountains. [Methods] The V3-V4 regions of 16S rRNA genes from 36 Gansu zokors' cecum were sequenced using Illumina high-throughput sequencing technology. The sequences were used for analysis of species diversity, abundance and community structure of the intestinal bacteria. [Results] The intestinal bacteria of Gansu zokor mainly consisted of three phyla, among which Firmicutes was the most predominant phylum, and the others of Bacteroidetes and Proteobacteria. At the genus level, the predominant genera were Oscillospira, Ruminococcus, Coprococcus and Desulfovibrio. By comparing samples obtained from different regions, we found intestinal bacterial community structure presented a high similarity among the three counties (Pengyang county, Longde county, and Jingyuan county), whereas it showed a low similarity between Haiyuan county with other counties. Meanwhile, we concluded different sexes had different impacts on intestinal bacterial community structure of Gansu zokor, whereas female zokors are highly variable on the community structure of gut bacteria than male zokors. In addition, season might be an important modulator of the intestinal bacteria. The results showed that intestinal bacterial diversity of Gansu zokor was significantly higher in autumn than in spring, the similarity of bacterial community structure in autumn was also higher than in spring. [Conclusion] Region and gender merely had significant effects on intestinal bacterial community structure of Gansu zokor, whereas seasons might have significant impacts on both community structure and diversity of cecal bacteria significantly.

Abstract:[Objective] In order to study the function of amino acid sites A172 and S173 in the substrate entryway of alanine racemase TtAlr from Thermoanaerobacter tengcongensis MB4. [Methods] The mutant vectors were constructed by site-directed mutagenesis PCR using plasmid pET-TtAlr as the template and expressed in E. coli BL21(DE3). The enzymes were purified by affinity chromatography. D-amino acid oxidase coupling method was used to detect enzyme activity and stability of each mutant and wild type TtAlr proteins. [Results] Both TtAlr and mutant proteins were expressed and purified successfully. Results of enzymatic properties show that A172 site mutation could improve the catalytic activity of TtAlr, but the stability of enzyme proteins decreased significantly. Likewise, S173 site mutation could increase the optimal temperature of TtAlr, prolonged the half-life of the enzyme and improved its stability, but the catalytic activity of the enzyme decreased significantly. [Conclusion] A172 and S173 amino acids residues in the substrate entryway of alanine racemase TtAlr were the key sites that played a major role in the catalytic activity and stability of the enzyme protein.

Abstract:[Objective] In order to give a full overview including ecological distribution, community structure and phylogenic diversity.[Methods] All n-damo bacteria related sequences from NCBI databases (data collected until November 2016) and together with our experimental data from Dajiuhu surface peat samples through 16S rRNA illuminia sequencing were set to do the bioinformatics analysis.[Results]n-damo bacteria were mainly detected in the sediment, wetlands and paddy soil. The average detection rate of n-damo bacteria based on pmoA gene was 7 times more than that based on 16S rRNA gene, whereas Shannon-Wiener index (1.4-3.4) remained relatively stable. The abundance of NC10 was only 0.067% from Dajiuhu peatland, which was the lowest abundance in all published data, indicating nitrogen was a limiting factor for NC10. Group A including Methylomirabilis oxyfera which had been test to have the anaerobic methane oxidation accounts for only a little bit (less than 20%), suggesting that the real ecological potential of methane consumption by n-damo bacteria need more deep study. [Conclusion] The integrated analysis on n-damo bacteria deepen our knowledge of this functional group and may give some guideline for further study.

Abstract:[Objective] Aimed to clarify the dynamic changes of airborne bacterial species and community diversity in sandstorm source areas in Xinjiang, we collected air samples from the dust source areas in the hinterland of Taklimakan and surroundings, and analyzed the community structure of airborne bacteria in the early, middle and late stages of sandstorm occurrence. [Methods] Based on the Illumina HiSeq sequencing platform, we constructed the 16S rRNA small fragments library by sequencing using double-end sequencing. [Results] The dust storm source bacteria belonged to 37 genera and affiliated to 4 phyla, and Proteobacteria accounted for 67.6%, Bacteroidetes accounted for 17.6%, Actinobacteria accounted for 11.7%, Firmicutes accounted for 2.9%. At genus level, there were altogether 8 different dominant genera in Xinjiang sandstorm source area, and the Non-metric Multidimensional Scaling result showed the significant differences in bacterial community composition at different sampling sites in different periods. The canonical correspondence analysis showed that the influence of airborne bacterial diversity in sandstorm area with environmental factors was as follows:altitude > latitude > longitude > humidity > air pressure > temperature with no significant difference. [Conclusion] The diversity and abundance of airborne bacterial community in the sandstorm source area in Xinjiang were very high. The differences of air bacterial community composition in the early, middle and late stages of sandstorm were significant in different areas. The influence of sandstorm on the indigenous bacterial communities in the five source areas was extremely significant.

Abstract:[Objective] A nitrous oxide reducing bacterium (DJ7) was isolated from paddy soil and its ability to reduce N₂O was detected under different incubation conditions. Aiming to understand the mechanisms of N₂O reduction and supply theoretical basis for regulation of N₂O emission. [Methods] The strain was isolated by microbial enrichment culture and identified by homogeneous analysis of its sequences of nosZ and 16S rRNA gene with the database of NCBI. The abilities of N₂O-reducing of DJ7 at different temperatures and O₂ concentrations were determined by measuring the amount of N₂O reduction through GC. [Results] The strain was characterized as Pseudomonas and contained nosZ gene, its N₂O reduction rate was more than 0.0219 μmol/min under anaerobic at 30℃. Although the increase of air O₂ concentration and shifts of incubation temperature at either direction resulted decreases of the capability of N₂O reduction, it maintained a certain ability in N₂O reduction. [Conclusion] The strain is an active facultative N₂O reducer belonging to Pseudomonas and possesses strong ability to reduce N₂O, especially under anaerobic condition. This strain may provide a new idea to alleviate the N₂O issue and play an active role in the ecological environment.

Abstract:[Objective] To understand the distribution of microorganisms and their effects on the waterlogged wooden relics collected from the "Nanhai No. 1" shipwreck. [Methods] Prokaryotic microorganisms in 10 wooden relics samples collected from the "Nanhai No. 1" shipwreck were identified by using high throughput sequencing technique. The composition and diversity of the bacterial communities in each sample were analyzed. [Results] Total 121 394 313 bacterial sequences with an average reading length of 458 bp were obtained. There were 3 780 different OTUs based on the similarity of the 97% sequences. According to the sequence in the GenBank database, 34 orders, 35 families and 187 genera of bacteria were involved in the wooded relics. Among these bacteria, most species were from Proteobacteria, and the species from Bacteroidetes ranked in the second. In the class level, the species from γ-Proteobacteria and α-Proteobacteria contributed more on community diversity. Hydrogenophaga, Lacibacter cauensis, Acidovorax delafieldii, Devosia, Sediminibacterium, Brevundimonas diminuta and Pseudomonas mendocina were found in all samples, and Muricauda, Thiobacillus, Rhodoplanes, Emticicia, Methylotenera mobilis and Flavobacterium were appeared with higher frequency of bacterial taxa. Principal component analysis showed that there were some differences in the composition of the bacterial community in different cultural relics. [Conclusion] There were rich biodiversity in prokaryotic microorganisms, both aerobic and anaerobic, in the archaeological wood from "Nanhai No. 1" shipwreck. The bacteria species, which participate in the decomposition of cellulose in the bacterial communities, resulted in a great challenge to the protection of cultural relics. At the same time, impact from iron-sulfur cycle bacteria should be taken into consideration due to they were existing in these relics.

Abstract:[Objective] To study the effect of autotrophic and mixotrophic cultivation on the growth, division and biochemical components accumulation of Chlorella pyrenoidosa, then to explore the response mechanism on circadian rhythm under artificial cultivation and optimize the corresponding technology. [Methods] BG11 medium was used for autotrophic cultivation of Chlorella pyrenoidosa and BG11 medium supplemented with glucose of different concentrations (1, 5, 10, 20 g/L) for mixotrophic cultivation. The algae cell number was determined by a hemocytometer, biomass was measured by dry weight, and the size and division of algae cells were observed by a microscope. We used staining method for quantification the total lipids of Chlorella pyrenoidosa. Chlorophyll, protein and starch contents were determined by ultraviolet spectrophotometry. [Results] Glucose had a significant growth promoting effect on Chlorella pyrenoidosa, and the optimum concentration was 10 g/L. Under a circadian rhythm, about 76.00% of new daughter cells in the autotrophic cultivation group were proliferated under dark period on the second and fifth day, whereas 67.50% of the new daughter cells were generated under light period in the mixotrophic group. The silent period of the algae in the autotrophic group was 8 hours, but that in the mixotrophic condition was only 4 hours on the second day. On the eighth day, the contents of total lipids and chlorophyll of algae cells kept relatively stable under a light-dark period, the protein and starch content of that reached their respective peak after 8 and 12 hours in light. [Conclusion] Mixotrophic cultivation with glucose can increase the biomass of Chlorella pyrenoidosa notably. The growth, proliferation and the biochemical components accumulation of Chlorella pyrenoidosa were regulated by circadian rhythm, the algae in autotrophic culture mainly grow during day in light and undergo cell proliferation in dark. The machanism of mixotrophic cultivation increasing the biomass was shortening the growth silence period of algae cells, and increasing the proportion of larger cells for passing through commitment point of cell-cycle, it was especially obvious during light period. The protein and starch contents of Chlorella pyrenoidosa showed obvious circadian rhythm, their best harvest time were 8 and 12 hours after in light, respectively.

Abstract:[Objective] This study aims to investigate the effect of inositol, sialic acid, and L-fucose metabolic pathways on pathogenicity in the process of Aeromonas hydrophila infection. [Methods] By homologous recombination technology, the genes associated with myo-inositol, sialic acid and L-fucose metabolism, iolC, fucK and nanA, were inactivated. Then fifty percent lethal doses (LD₅₀) of the deletion mutants were determined in zebrafish and the bacterial loads in different tissues were assayed in crucian carp co-infected with the wild-type and its derivative mutant strains. [Results] The inactivation of all the three metabolic genes successfully blocked the ability of A. hydrophila to degrade the corresponding substrates. Deletion of iolC resulted in a nearly 12-fold increase in LD₅₀ in zebrafish, whereas deletion of nanA and fucK had no significant effect on LD₅₀. The bacterial loads of the wild strain were significantly higher than that of the iolC deletion strain in liver, spleen and kidney after co-infection, indicating an obvious competitive growth advantage of the wild strain. But for nanA and fucK mutant strains, after co-infection with the wild strain, there was no significant difference in bacterial loads of the wild-type and its derivative mutant strains in different tissues. [Conclusion] The inositol metabolic pathway plays an important role during A. hydrophila infection, while sialic acid and L-fucose metabolic pathways have no significant effect on the pathogenicity of this bacterium.

Abstract:[Objective] To study the role of Peste des petits ruminants virus envelope glycoproteins hemagglutinin and fusion during the fusion process. [Methods] The eukaryotic expression recombinant plasmids pCMV-HA-H, pCAGGS-Flag-F, pCMV-Myc-SLAM and pCMV-Myc-Nectin 4 sequenced correctly were transfected or co-transfected into CHO-K1 cells, and then the fusion role of PPRV H and F proteins was analyzed by microscopy and indirect immunofluorescence technique. [Results] The formation of syncytium was observed in the co-transfected cells while it was not found in the single plasmid transfected cells and the negative control. The number of syncytia in cells co-expressing H and F proteins was significantly more than that of the other groups. The H and F proteins were co-redistributed to form the polar caps (co-capping) in the same position. [Conclusion] PPRV F protein is an essential protein for fusion of the viral membrane with host cell membrane, but it must interact with PPRV H to make virus invasion successfully.

Abstract:[Objective] Through complex of whitening ingredients from different Chinese herbs, we studied the active ingredients and effectiveness of compound Chinese herbs fermentation broth. [Methods] The tyrosinase inhibition rate was used as the index, the best compound composition was obtained. The contents of total polyphenols, polysaccharides, flavonoids, and protein in the fermentation broth were measured by UV spectrophotometry, and amino acids compositions were analyzed by HPLC. We evaluated their antioxidant activity in fermentation broth by total reducing power and radical scavenging tests. The whitening efficacy was evaluated by tyrosinase inhibition test. [Results] Rubifructus had the best inhibition of tyrosinase activity by orthogonal designed experiment. The fermentation broth of compound Chinese herbs contained 147.8 g/kg of total polyphenols, 4.360 g/kg of polysaccharides, 1.170 g/kg of flavonoids and 1.220 g/kg of total protein. The fermentation broth of compound Chinese herbs had strong total reducing power, hydroxyl radical scavenging ability and 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activity. Tyrosinase inhibition rate of 5% (W/V) compound Chinese herbs fermentation broth reached 82.41%. [Conclusion] The fermentation broth of compound Chinese herbs contained multiple active ingredients, with antioxidant activity and whitening efficacy.

Abstract:[Objective] Candida tropicalis has become an important industrial strain to produce dicarboxylic acids due to its high ω-oxidation activity. Fatty aldehyde dehydrogenases (FALDHs) play important roles in the ω-oxidation pathway, converting fatty aldehydes to fatty acids. However, FALDHs characterization and their roles in the synthesis of dibasic acids have not been studied in depth. Therefore, we cloned two genes CtAld1 and CtAld2 encoding FALDHs and evaluated their function in cell phenotype, enzyme activity and dicarboxylic acids accumulation. [Methods] We screened two genes of CtAld1 and CtAld2 through genome mining and sequence alignment. Based on sequence analysis, we deleted CtAld1 and CtAld2 either separately or accumulatively by homologous recombination method, and generated various mutants. The effect of deletion of CtAld1 and CtAld2 on cell growth, FALDH activity and dodecanedioic acid (DCA12) production were evaluated and compared. [Results] XZX-1 (ΔCtAld1/ΔCtAld1), XZX-2 (ΔCtAld2/ΔCtAld2) and XZX-12 (ΔCtAld1/ΔCtAld1, ΔCtAld2/ΔCtAld2) were obtained. When using dodecane as sole carbon source, deletion of CtAld2 gene significantly inhibited cell growth, and the intracellular FALDH activity was only 30% of the parental strain. Deleting CtAld1 had a slight promotion of cell growth, however intracellular FALDH activity was decreased to some extent. Furthermore, simultaneous deletion of CtAld1 and CtAld2 significantly impaired the cell growth performance and decreased FALDH activity, thus causing a distinct lower DCA12 yield compared to the wild type strain. [Conclusion] The deletion of CtAld1 in C. tropicalis could reduce the yield of DCA12. And CtAld2 plays an important role in the growth on dodecane and production of DCA12. To our knowledge, they could be recruited as target genes for metabolic engineering of ω-oxidation pathway in C. tropicalis.