Screening and Toxicity Determination of Intestinal Aerobic Bacteria in 2^(nd) instar Larvae of Dendrolimu Kikuchii

[ Objective ] The paper was to screen intestinal aerobic bacteria from 2nd instar larvae of Dendrolimu kikuchii and determine its toxicity. [ Method ] The intestinal samples of 2nd instar larvae of D. kikuchii were diluted in gredient and coated on plates to isolate strains, and a total of 5 strains of aerobic bacteria were isolated. With bacterial genomic DNA as template, the universal primers of bacterial 16S rDNA （27f and 1 492r） were used to amplify the template, 4 restriction enzymes Hae Ⅲ and Hind Ⅲ, Hinf Ⅰ and Taq Ⅰ were used to carry out ARDRA polymorphism analysis on PCR products. [ Result] Clustering map analysis showed that 5 strains of aerobic bacteria clustered into two different taxonomic operating units （OTU） in 95% similarity level, this indicated that the genetic diversity level of intestinal aerobic bacteria in 2nd instar larvae of D. kikuchii was relatively low. The indoor toxicity test showed that the death peaks of insects killed by intestinal bacteria were during 4 - 10 d, the insecticidal effect of strain 4 was the best with corrected mortality rate of 53.57% at 12 d. [ Conclusion] The paper provided reference for control of D. kikuchii.

Abundance of general aerobic heterotrophic bacteria in the Bering Sea and Chukchi Sea and their adaptation to temperature

The abundance of general aerobic heterotrophic bacteria(GAB) from the water and sediment in the Bering Sea and the Chukchi Sea was determined by using petri dish cultivation and counting method. The abundance of GAB among the different sea areas, sampling sites, layers of sediments surveyed and adaptability to differential temperatures was studied. The result obtained showed that: the occurrence percentage of GAB in the surface water was higher than that in sediment, but the abundance was only 0.17% of sediment. The occurrence percentage of GAB in surficial layer of sediment was higher than that in the other layers. The occurrence percentage of GAB in surficial layer of sediment was higher than that in the other layers. The occurrence percentage, abundance and its variation of GAB in the Bering Sea were higher than that in the Chukchi Sea respectively. The average value of the abundance in the all sediment surveyed was 3166.3×10 2 CFU·g -1(wet.). The abundance of GAB in sediment showed a trend: roughly higher in the lower latitudinal area than higher latitude. The results from temperature test mean that: the majority of bacteria tested were cold-adapted ones, minority might be mesophilic bacteria. The results indicated that, Arctic ocean bacteria had a stronger adaptability to environmental temperature.

Community structure of aerobic anoxygenic phototrophic bacteria in algae-and macrophyte-dominated areas in Taihu Lake,China

Aerobic anoxygenic phototrophic bacteria(AAPB)repre sent a major group of bacterioplankton assemblages in many water systems and some are assumed to be closely associated with phytoplankton.However,studies on relationships between AAPB and cyanobacterial blooms are in scarcity.The dynamics of the abundance and diversity of AAPB was compared based on pufM gene in Meiliang Bay(featured by cyanobacterial blooms)and East Bay(featured by macrophyte)of Taihu Lake,a shallow subtropical lake in the East China plain.AAPB abundance was not significantly different between the two sites,and they were positively correlated with dissolved organic carbon(DOC)concentration.The ratios of AAPB to total bacteria varied from 3.4%to 11.5%and peaked in winter in both site s.No significant differences of AAPB community compositions were detected between the two sites,but there was a separation between warm seasons(June,August,and October)and cold seasons(December,February,and April).Rhizobiales and Limnohabitans-like pufM sequences were significantly contributors for the difference between two seasons,and specially enriched in cold seasons.Chlorophyll a(Ch1 a)and DOC were the most significant variables influencing the AAPB community structure.Furthermore,Porphyrobacter and Rhodospirillales-like pufM sequences were positively correlated with Ch1 a,indicating potential influence of cyanobacterial blooms on these AAPB taxa.These results suggested that diverse AAPB ecotypes coexisted in Taihu Lake,and their ecological role in carbon cycling in the lake may not be ignored.

Phylogenetic analysis of aerobic anoxygenic phototrophic bacteria and their relatives based on farnesyl pyrophosphate synthase gene

The study aims to reveal phylogenetic and evolutionary relationship between aerobic anoxygenic phototrophic bacteria（AAnPB） and their relatives,anaerobic anoxygenic phototrophic bacteria（AnAnPB） and nonphototrophic bacteria（NPB,which had high homology of 16S rDNA gene with AAnPB and fell into the same genus）,and validate reliability and usefulness of farnesyl pyrophosphate synthase（FPPS） gene for the phylogenetic determination.FPPS genes with our modified primers and 16S rDNA genes with general primers,were amplified and sequenced or retrieved from GenBank database.In contrast to 16S rDNA gene phylogenetic tree,AAnPB were grouped into two clusters and one branch alone with no intermingling with NPB and AnAnPB in the tree constructed on FPPS.One branch of AAnPB,in both trees,was located closer to outgroup species than AnAnPB,which implicated that some AAnPB would be diverged earlier in FPPS evolutionary history than AnAnPB and NPB.Some AAnPB and NPB were closer located in both trees and this suggested that they were the closer relatives than AnAnPB.Combination codon usage in FPPS with phylogenetic analysis,the results indicates that FPPS gene and 16S rRNA gene have similar evolutionary pattern but the former seems to be more reliable and useful in determining the phylogenic and evolutionary relationship between AAnPB and their relatives.This is the first attempt to use a molecular marker beside 16S rRNA gene for studying the phylogeny of AAnPB,and the study may also be helpful in understanding the evolutionary relationship among phototrophic microbes and the trends of photosynthetic genes transfer.

Geographical distribution of general aerobic heterotrophic bacteria in surficial sediments from the Chukchi Sea and Canadian Basin

This paper determined the abundance of General Aerobic heterotrophic Bacteria （GAB） in surficial sediments from the Chukchi Sea and the Canadian basin by using MPN and discussed their geographical distribution. The result shows that the determination percentages of the GAB were high, even till 100 percentage. The abundance range and averages of GAB for 4℃and 25℃ were from 4.00 × 10^2 to 2.40× 10^6 , 1.71×10^6 ind.·g^-1 （wet sample ） and from 2.40 ×10^5 to 2.40×10^7 , 1.10×10^7 ind. ·g^-1 （wet sample ） respectively. Not only the abundance range but also the averages of GAB in 25℃ were higher than that in 4℃. The abundance of GAB in sediments show a tendency that it is roughly greater in the lower latitudinal area than in the higher latitudinal area. The abundance of GAB increased from east to west as for the longitudinal distribution. With the water depth increasing, the abundance of GAB at 4℃ decreased, but GBA at 25℃ is not changed obviously with water depth. It seems that warmer circumstantial temperature is more suitable for some GAB.

Diversity of the aerobic anoxygenic phototrophy gene pufM in Arctic and Antarctic coastal seawaters

Aerobic anoxygenic phototrophic（AAP） bacteria serve important functions in marine carbon and energy cycling because of their capability to utilize dissolved organic substrates and harvest light energy.AAP bacteria are widely distributed in marine environments,and their diversity has been examined in marine habitats.However,information about AAP bacteria at high latitudes remains insufficient to date.Therefore,this study determined the summer AAP bacterial diversity in Arctic Kongsfjorden and in the Antarctic coastal seawater of King George Island on the basis of puf M,a gene that encodes a pigment-binding protein subunit of the reaction center complex.Four puf M clone libraries were constructed,and 674 positive clones were obtained from four investigated stations（two in Kongsfjorden and two in the Antarctic Maxwell Bay）.Arctic clones were clustered within the Alphaproteobacteria,whereas Antarctic clones were classified into the Alphaproteobacteria and Betaproteobacteria classes.Rhodobacteraceae-like puf M genes dominated in all samples.In addition,sequences closely related to puf M encoded on a plasmid in Sulfitobacter guttiformis were predominant in both Arctic and Antarctic samples.This result indicates the transpolar or even global distribution of puf M genes in marine environments.Meanwhile,differences between the Arctic and Antarctic sequences may prove polar endemism.These results indicate the important role of Rhodobacteraceae as AAP bacteria in bipolar coastal waters.

Fed-Batch Fermentation for Spinosad Production in an Improved Reactor

As a kind of aerobic bacteria, Saccharopolyspora spinosa exhibits a high demand for oxygen. In the fermentation process, the methods of increasing ventilation and improving agitation speed are usually adopted to achieve higher values of dissolved oxygen. These methods decrease the efficiency of spinosad biosynthesis. In this study, an improved reactor was designed to solve these problems. The exhaust gas reflux device, impellers, and baffles were improved. Furthermore, we established the kinetic models for the cell growth, substrate consumption and spinosad generation in batch fermentation process. The simulation results were in good agreement with the experimental data. Spinosad production reached 583.86 mg/L after employing the suitable feeding strategy by fed-batch fermentation in the improved reactor, whereas it was only 157.01 mg/L before optimization. The method described can provide insight to strengthen spinosad production and can be extended to the culturing process of filamentous aerobic bacteria. © 2017 Tianjin University and Springer-Verlag Berlin Heidelberg

A novel sampling method for the investigation of gut microbiota

AIM： In order to characterize the qualitative and quantitative microorganisms in different sites of the lower digestive tract （LDT） in healthy volunteers, a specific technique was developed for collecting mucous of the distal ileum, colon and rectum. METHODS： A polyethylene tube was designed to go through the colonoscope channel with a No. 8 French tube. In order to avoid internal contamination, the distal extremity was protected with a membrane of microfilm after being sterilized in ethilene oxid. To facilitate the aspiration of a precise volume, its interior was coated with silicone. One hundred rnicrolliter （0.1 mL） sample of mucous was collected and transferred into an Eppenddorff tube containing nine hundred rnicrolliter （0.9mL） of VMGA-3 （viable medium of Goteborg）. This procedure was repeated at each site of the LDT with a new sterilized catheter. RESULTS： All sites revealed the ＂non pathogenic＂ anaerobic bacteria Veillonella sp （average 10s colony forming units/mL-CFU/mL）, allowing to conclude an environment of low oxidation-reduction potential （redox） in the LDT. It was also characterized the presence of Klebisiella sp with significant statistical predominance （SSP） in the ileum. Enterobacter sp was found with SSP in the sigrnoid colon, Bacteroides sp non-pigmented （npg） and Ecoli with SSP in the sigrnoid colon and rectum, Enterococcus sp and Lactobacillus sp with SSP in the rectum, all in a mean concentration of 10s CFU/mL CONCLUSION： This procedure is feasible and efficient and can point out a similar distribution of the aerobic and anaerobic bacteria with the presence of biological markers of normal microbiota in the LDT.

Environmental control of growth and BChl a expression in an aerobic anoxygenic phototrophic bacterium, Erythrobacter longus (DSMZ6997)

anoxygenic phototrophic bacteria （AAPB）, which form a unique functional group of heterotrophic bacteria, have the abilit,to utilize light energy. The impact of carbon source and light intensity on the growth and bacteriochlorophyn a （ BChl a） expression of a typical strain of AAPB, Erythrobacter longus strain DSMZ6997 was examined during batch culture and continuous culture. The results showed that the expression of BChl a in DSMZ6997 was regulated by both carbon-source and light conditions, and was stimulated by low availability of carbon but inhibited by light to a certain extent at 300 lx and completely at 1 500 lx. In contrast, cell abundance, and even cell size of this strain, was substantially enhanced under light/dark cycle cultivation conditions over dark conditions, indicating the promotion of growth by light. These results led to the conclusion that utilization of light through BChl a helps AAPB to survive under carbon stress, while light at high intensity is harmful to the synthesis of BChl a in AAPB.

Microbial Limit Test of Zhideke Granules

[Objectives]To test the microbial limit of Zhideke granules.[Methods]According to the relevant regulations on the testing of granules in Chinese Pharmacopoeia in 2015,the applicability of the methods for counting the total number of aerobic bacteria,molds and yeasts and objectionable organisms in Zhideke granules was studied.[Results]The total number of aerobic bacteria in Zhideke granules was determined by dilution plate method(1∶20,1 mL per dish),the total number of molds and yeasts was determined by conventional plate method(1∶10,1 mL per dish),and E.coli was detected by conventional method of TTB(1∶10 test solution,10-100 mL TSB).[Conclusions]The method was suitable for the microbial limit test of Zhideke granules.

Biological Degradation of Some Organic Compounds Enrolled in Paper Industry—A Pollution Prevention Approach

Evaluation of the elimination and the “ultimate” biodegradation by aerobic microorganisms of some organic compounds commonly used in paper manufacturing technology was investigated. Biodegradation lines of nine organic compounds were determined as percentage removal of chemical oxygen demand (COD) over 7 days incubation. The results of the biodegradability test clearly revealed that some of the organic compounds under investigation are highly biodegradable, while others rank from fairly to even non biodegradable. Significant biodegradation results were recorded for anti coating ester (95.0%), basoplast 200D (85.3%) and basoplast PR 8050(87.6%). A bleaching agent (formamidin sulfinic acid), ukanol BSA and solidurit KM demonstrate moderate biodegradation results of 62.1%, 76.2% and 69.8%, respectively. Poor biodegradation results for Hedifix M/35 (12.7%), basazol orange (34.9%) and basazol brown (29.0%) were recorded. Accordingly, appropriate precaution should be taken into consideration when using these compounds for industrial applications.

The response of structure and nitrogen removal function of the biofilm on submerged macrophytes to high ammonium in constructed wetlands

The ammonium exceedance discharge from sewage treatment plants has a great risk to the stable operation of subsequent constructed wetlands(CWs).The effects of high ammonium shocks on submerged macrophytes and epiphytic biofilms on the leaves of submerged macrophytes in CWs were rarely mentioned in previous studies.In this paper,the 16S rRNA sequencing method was used to investigate the variation of the microbial communities in biofilms on the leaves of Vallisneria natans plants while the growth characteristics of V.natans plants were measured at different initial ammonium concentrations.The results demonstrated that the total chlorophyll and soluble sugar synthesis of V.natans plants decreased by 51.45%and 57.16%,respectively,and malondialdehyde content increased threefold after8 days if the initial NH_(4)^(+)-N concentration was more than 5 mg/L.Algal density,bacterial quantity,dissolved oxygen,and pH increased with high ammonium shocks.The average removal efficiencies of total nitrogen and NH_(4)^(+)-N reached 73.26%and 83.94%,respectively.The heat map and relative abundance analysis represented that the relative abundances of phyla Proteobacteria,Cyanobacteria,and Bacteroidetes increased.The numbers of autotrophic nitrifiers and heterotrophic nitrification aerobic denitrification(HNAD)bacteria expanded in biofilms.In particular,HNAD bacteria of Flavobacterium,Hydrogenophaga,Acidovorax,Acinetobacter,Pseudomonas,Aeromonas,and Azospira had higher abundances than autotrophic nitrifiers because there were organic matters secreted from declining leaves of V.natans plants.The analysis of the nitrogen metabolic pathway showed aerobic denitrification was the main nitrogen removal pathway.Thus,the nitrification and denitrification bacterial communities increased in epiphytic biofilms on submerged macrophytes in constructed wetlands while submerged macrophytes declined under ammonium shock loading.

Microprofiles of activated sludge aggregates using microelectrodes in completely autotrophic nitrogen removal over nitrite （CANON） reactor

Microsensor measurements and fluorescence in situ hybridization （FISH） analysis were combined to investigate the microbial populations and activities in a laboratory-scale sequencing batch reactor （SBR） for completely autotrophic nitrogen removal over nitrite （CANON）. Fed with synthetic wastewater rich in ammo- nia, the SBR removed 82.54-5.4% ofinfluent nitrogen and a maximum nitrogen-removal rate of 0.52kgN.m 3.d-l was achieved. The FISH analysis revealed that aerobic ammonium-oxidizing bacteria （AerAOB） Nitrosomonas and anaerobic ammonium-oxidizing bacteria （AnAOB） dominated the community. To quantify the microbial activities inside the sludge aggregates, microprofiles were measured using pH, dissolved oxygen （DO）, NH＋, NO＋ and NO3 microelectrodes. In the outer layer of sludge aggregates （0-700 μm）, nitrite-oxidizing bacteria （NOB） showed high activity with 4.1 gmol-cm3＋ h-1 of maximum nitrate production rate under the condition of DO concentration higher than 3.3 mg. L 1. Maximum AerAOB activity was detected in the middle layer （depths around 1700 μm） where DO concentration was 1.1 μg. L-1. In the inner layer （2200-3500μm）, where DO concentration was below 0.9 mg. L-1, AnAOB activity was detected. We thus showed that information obtained from microscopic views can be helpful in optimizing the SBR performance.

The research of typical microbial functional group reveals a new oceanic carbon sequestration mechanism——A case of innovative method promoting scientific discovery

Marine microbes are major drivers of marine biogeochemical cycles and play critical roles in the ecosystems. Aerobic anoxygenic phototrophic bacteria(AAPB) are an important bacterial functional group with capability of harvesting light energy and wide distribution, and appear to have a particular role in the ocean's carbon cycling. Yet the global pattern of AAPB distribution was controversial at the beginning of the 21 st century due to the defects of the AAPB enumeration methods. An advanced time-series observation-based infrared epifluorescence microscopy(TIREM) approach was established to amend the existing AAPB quantitative deviation and led to the accurate enumeration of AAPB in marine environments. The abundance of AAPB and AAPB% were higher in coastal and continental shelf waters than in oceanic waters, which does not support the idea that AAPB are specifically adapted to oligotrophic conditions due to photosynthesis in AAPB acting a supplement to their organic carbon respiration. Further investigation revealed that dependence of AAPB on dissolved organic carbon produced by phytoplankton(PDOC) may limit their competition and control AAPB distribution. So, the selection of carbon sources by AAPB indicated that they can effectively fractionate the carbon flow in the sea. Enlightened by these findings, the following studies on the interactions between marine microbes and DOC led to the discovery of a new mechanism of marine carbon sequestration—the Microbial Carbon Pump(MCP). The conceptual framework of MCP addresses the sources and mechanism of the vast DOC reservoir in the ocean and represents a breakthrough in the theory of ocean carbon sequestration.