Hong-Ying Hu

Analysis of Respiratory Quinones in Soil for Characterization of Microbiota

Abstract Method for the analysis of respiratory quinones in soil was developed to characterize soil microbiota. The respiratory quinones were extracted with a mixture of chloroform and methanol using a Wahling blender or a sonicator and cleaned-up by a silicagel column cartridge. The quinone species were determined by reverse-phase high performance liquid chromatography. Spectra of peaks were measured with a photodiode array detector to examine the purity. More than 90% of extractable quinones in soil were recovered by three extractions. The recovery of quinones, added to soil as freeze-dried powder of microorganisms, was higher than 96%. This procedure led to higher representative results as follows. Fluctuations within 95% of cumulative frequency were 13% for the extracted amount of quinones and 20% for the dissimilarity, respectively. The quinone profiles of four soils were determined as follows: a soil from the aerobic layer of a paddy field, an ando soil and two yellow upland soils which had received...

Development of a Novel Solid Phase Extraction Method for the Analysis of Bacterial Quinones in Activated Sludge with a Higher Reliability

A novel analytical method for analysis of microbial quinones in activated sludge sample was developed with improved reliability compared to the conventional method. The operating conditions for the extraction of quinones from activated sludge sample with a methanol-chloroform mixture and hexane were optimized. A solid phase extraction method using the Sep-Pak Plus Silica, a small column packed with silica gel, was employed for the purification and separation of quinones instead of column chromatography and thin layer column chromatography used in the conventional method. The analytical operation was simplified and the analytical time was shortened by a half or two-thirds of that required for the conventional method. The recoveries of quinones were markedly increased from 30-60% with the conventional method to as high as 90-101% with the improved method, demonstrating that the reliability of the analytical results of the improved method is significantly higher than that of the conventional method. The quinone content value obtained using the improved method was twice as high as that obtained using the conventional method for the same activated sludge sample, but the quinone profiles expressed as the mole fraction of each quinone type were the same for each method.

Quantitative analyses of the change in microbial diversity in a bioreactor for wastewater treatment based on respiratory quinones

Abstract The change in microbial diversity in a submerged aerobic biofilter, into which a persistent chemical of dimethylformamide was fed, were analyzed based on the quinone profiles using a new diversity index of the respiratory quinones (DQ) and an index of equitability of the distribution of quinone species (EQ) originally proposed in this study: DQ=(∑ f 1/2 k ) 2 , EQ=DQ/ n . Where, f k is the mole fraction of quinone species k and n is the number of quinone species, respectively. The DQ value increases with an increase in the number and the equitability of the distribution of quinone species. In the case where the mole fractions of all quinone species are equal, the value of DQ becomes equal to the number of quinone species and the EQ value becomes 1. A simulation analyses shows that DQ has a higher sensitivity to the change in number of quinone species than Shannons index, i.e. one of the most widely used diversity indices. The relative sensitivity of DQ to the change in mole fraction of quinone species is higher than that of Shannons diversity index for natural ecosystems in which many kinds of quinone species exist and occupy smaller fractions. The quinone profiles of the biofilm significantly changed and the values of DQ and EQ increased during the acclimation of the biofilm to dimethylformamide in the biofilter. The plot of the mole fraction of dominant quinone species to the DQ values showed higher sensitivity to the change in the microbial community. These results indicated that DQ and EQ are useful indices for the evaluation of the changes in the microbial community in response to environmental conditions.

Long-term changes in microbial community structure in soils subjected to different fertilizing practices revealed by quinone profile analysis

Abstract Quinone profile analysis of stored air-dried soils gave an approximation of the long-term changes in the microbial community structure in four soils subjected to different types of fertilizer application from 1987 to 1997: unfertilized soil (NF-soil), soil amended with chemical fertilizers (CF-soil), soil amended with chemical fertilizers and 40 t ha-1 y-1 of farmyard manure (CF+ FYM-soil), and soil amended with 400 t ha-1 y-1 of farmyard manure (FYM-soil). The carbon content increased, and the soil pH remained higher in the soils receiving farmyard manure. Principal component analysis of the quinone profiles of the soils indicated that the microbial community structure showed a high similarity among the four soils before the onset of cultivation and changed to a different community structure specific to the respective fertilizing practices except for the NF-soil. The specific quinone profile became stable after two cropping seasons in the FYM-soil, after 10 cropping seasons in the CF+ FYM-soil a...

Optimal operation of bioreactor system developed for the treatment of chromate wastewater using enterobacter cloacae HO-1

The characteristics of a novel bioreactor system developed for the simultaneous treatment of toxic hexavalent chromium (chromate) and high-strength organic pollution by introducing a chromate-resistant and chromate-reducing bacterium Enterobacter cloacae strain HO-1 (HO-1, hereafter) were studied to optimize its operating condition. Based upon the growth and chromate-reducing kinetics of HO-1, a mathematical model to simulate the change of chromate removal rate per unit volume of bioreactor and that per unit cell mass under the fed-batch operation was proposed. The effects of repeated cultivation of HO-1 with ORP-controlled aeration and the cumulative chromate reduction per unit cell mass on the microbial activity were investigated to represent the kinetic expression of chromate reduction. By using the mathematical model developed through the present research, the relationship between the operating conditions and the chromate reduction rate in the bioreactor was simulated and the model was verified by comparing it with the observed data. Thus the optimal operating conditions to give the higher reduction rate and the longer operating interval with and without aerated cultivation of HO-1 are extensively discussed. This bioreactor can be used for the treatment of chromate wastewater provided that some high concentration organic wastes are available. The rate of chromate reduction in the bioreactor is as high as 10 - 15 g-Cr 6+ /m 3 h. Chromium hydroxide, products of biological chromate reduction, can be recycled as a green pigment.

Linear relation between the amount of respiratory quinones and the microbial biomass in soil

Abstract A linear relationship was observed between the amount of total respiratory quinones and the microbial biomass measured by a fumigation-extraction method in 15 soil samples regardless of the significant differences in the composition of the quinone profiles, with one exception in a soil amended with a very high application rate of farmyard manure. It is suggested that the amount of total respiratory quinones can be used as an indicator of the microbial biomass in soil.

Respiratory quinone profile as a tool for refractory chemical biodegradation study

The respiratory quinone profile was successfully used as a tool for identifying the change of bacterial phase in the microbial film of aerobic submerged biofilter treating refractory chemicals such as dimethylformamide (DMF). It was ascertained that the coexistence of easily biodegradable substances such as glucose and peptone hindered the acclimation of bacteria inhabiting the microbial film in biofilter to DMF. The higher initial microbial concentration and the feeding of DMF as a sole organic substance accelerated the acclimation of microbial film to DMF. Appreciable changes in quinone profile were detected through the acclimation to DMF, and the results showed that DMF degrading bacteria such as Methylobacterium and Mycobacterium had grown in the biofilter.

Changes in the Microbial Community Structure in Soils Treated with a Mixture of Glucose and Peptone with Reference to the Respiratory Quinone Profile

Abstract Based on the respiratory quinone profile, changes in the structure of microbial communities in the soil samples from Nagoya University Farm were monitored after the treatment with 1% of a mixture of glucose and peptone. Samples of two soils differing in the fertilization history were examined: CF-soil with the application of only chemical fertilizers and FYM-soil with the application of only farmyard manure at a high rate. In the CF-soil, the amount of water-soluble organic carbon (WOC), indicator of the mixture of glucose and peptone, decreased to the original level after 14 d. After 7 d, the soil pH reached the maximum level, then decreased gradually. Changes in the inorganic nitrogen levels in the water extract also reflected the 14-d period of mineralization. The amount of respiratory quinones reached maximum levels after 7 d and gradually decreased, reflecting the changes in the microbial biomass. The quinone composition significantly changed during the 14-d period and returned to a profile similar to the original one after 28 d. Diversity of quinones significantly decreased during the 14-d period due to the predominance of ubiquinone with 9 isoprenoid units. In the FYM-soil, the amount of WOC decreased to the original level after 1 d, and the pH and inorganic nitrogen levels in the water extract reflected the one-day mineralization period, and nitrification started after 3 d. Although the amount of quinones indicated an increase in the microbial biomass for 14 d, the quinone composition did not change. These findings suggested that long-term application of farmyard manure resulted in stable microbial communities in response to the incorporation of organic matter in soil.

Quantitative detection of Enterobacter cloacae strain HO-1 in bioreactor for chromate wastewater treatment using polymerase chain reaction (PCR)

Abstract A novel method for the detection of specific microbes using the polymerase chain reaction (PCR) technique was developed and applied for the quantitative detection of Enterobacter cloacae strain HO-1 (HO-1, hereafter) in the experimental apparatus used for the treatment of wastewater containing hexavalent chromium (chromate). Note that HO-1 is capable of reducing toxic chromate to trivalent chromium. Based upon the preliminary experiments using pUC18 DNA, conditions giving a constant amplification rate of the targeted DNA with high reliability were determined for the quantitative detection of its initial concentration by the extrapolation. The cell density of HO-1 determined by the PCR technique was compared with the results counted on the selective medium agar plate conventionally used for the counting of HO-1. The procedure proposed here was successfully applied to estimate the cell density of Enterobacter cloacae HO-1 in the bioreactor for the treatment of chromate wastewater.

Effect of temperature on the reaction rate of bacteria inhabiting the aerobic microbial film for wastewater treatment

Abstract The effect of temperature on the reaction rate of bacteria inhabiting a microbial film was investigated based on the experimental observations of a submerged aerobic biofilter for wastewater treatment. BOD removal rate per unit surface area of packing material was not significantly affected by the decrease in temperature. The BOD removal rate was compensated with the increased number of bacterial cells eaven if cell activity was decreased. Activation energy for aerobic reaction for BOD removal and for carbon dioxide production in the microbial film was estimated to be in the range of 50–70 kJ·mol−1. This range of values coincided well with those previously reported in the literature for wastewater treatment and growth of pure bacterial culture. Moreover, it was ascertained that the production rate of carbon dioxide in the microbial film was in the range of those observed for E. coli and Salmonella sp. The production rate of carbon dioxide extrapolated from the present data using the activation energy mentioned above coincided well with the aerobic reaction rate observed in the compost of sewage sludge. Bacterial phase change in the film was studied with reference to the quinone profile as well.