Kinetics of Bacterial Growth on Chlorinated Aliphatic Compounds

With the pure bacterial cultures Ancylobacter aquaticus AD20 and AD25, Xanthobacter autotrophicus GJ10, and Pseudomonas sp. strain AD1, Monod kinetics was observed during growth in chemostat cultures on 1,2-dichloroethane (AD20, AD25, and GJ10), 2-chloroethanol (AD20 and GJ10), and 1,3-dichloro-2-propanol (AD1). Both the Michaelis-Menten constants (K_m) of the first catabolic (dehalogenating) enzyme and the Monod half-saturation constants (K_s) followed the order 2-chloroethanol, 1,3-dichloro-2-propanol, epichlorohydrin, and 1,2-dichloroethane. The K_s values of strains GJ10, AD20, and AD25 for 1,2-dichloroethane were 260, 222, and 24 μM, respectively. The low K_s value of strain AD25 was correlated with a higher haloalkane dehalogenase content of this bacterium. The growth rates of strains AD20 and GJ10 in continuous cultures on 1,2-dichloroethane were higher than the rates predicted from the kinetics of the haloalkane dehalogenase and the concentration of the enzyme in the cells. The results indicate that the efficiency of chlorinated compound removal is indeed influenced by the kinetic properties and cellular content of the first catabolic enzyme. The cell envelope did not seem to act as a barrier for permeation of 1,2-dichloroethane.     

Stomatal Opening as a Practical Indicator of Moisture Stress in Cotton

The response of stomata to varying soil moisture content was studied in cotton by means of a new type of porometer suitable for field work. The results showed that maximum stomatal aperture was only attained when soil moisture was near the upper limit of the so called “available range”. These results indicate the possibility of making practical use of the new type of porometer for the purpose of determining soil moisture deficiency under field conditions, thus aiding in irrigation practices.     

Comparative Growth of Natural Bacterial Isolates on Various Lignin-Related Compounds

Bacterial strains were isolated on the basis of their ability to proliferate in a minimal medium containing one of a series of lignin-related compounds as the sole carbon and energy source. These included the aromatic monomers guaiacol, vanillic and coumaric acids, a dimer and a trimer possessing the arylglycerol-β-aryl ether linkage, anisoin, and both the ether-soluble and -insoluble fractions of kraft lignin. The growth of the strains on each of these compounds was measured. The results showed that the metabolic properties of the strains varied according to the structure of the carbon sources used for their selection. Spectrophotometric tracings of the culture medium during the log phase of growth of one of the strains on the β-O-4 dimer revealed decomposition with the release of guaiacol.     

Synergy of Fresh and Accumulated Organic Matter to Bacterial Growth

The main goal of this research was to evaluate whether the mixture of fresh labile dissolved organic matter (DOM) and accumulated refractory DOM influences bacterial production, respiration, and growth efficiency (BGE) in aquatic ecosystems. Bacterial batch cultures were set up using DOM leached from aquatic macrophytes as the fresh DOM pool and DOM accumulated from a tropical humic lagoon. Two sets of experiments were performed and bacterial growth was followed in cultures composed of each carbon substrate (first experiment) and by carbon substrates combined (second experiment), with and without the addition of nitrogen and phosphorus. In both experiments, bacterial production, respiration, and BGE were always higher in cultures with N and P additions, indicating a consistent inorganic nutrient limitation. Bacterial production, respiration, and BGE were higher in cultures set up with leachate DOM than in cultures set up with humic DOM, indicating that the quality of the organic matter pool influenced the bacterial growth. Bacterial production and respiration were higher in the mixture of substrates (second experiment) than expected by bacterial production and respiration in single substrate cultures (first experiment). We suggest that the differences in the concentration of some compounds between DOM sources, the co-metabolism on carbon compound decomposition, and the higher diversity of molecules possibly support a greater bacterial diversity which might explain the higher bacterial growth observed. Finally, our results indicate that the mixture of fresh labile and accumulated refractory DOM that naturally occurs in aquatic ecosystems could accelerate the bacterial growth and bacterial DOM removal.     

Construction of a New Bacterial Cloning Vector Using a Mutant Green Fluorescent Protein as an Indicator

A new bacterial cloning vector, pGreenLD, derived from the triple substitution mutated Aequorea v/ctor/a green fluorescent protein(GFP-S65A, V68L, S72A), when expressed in E. coli produced colonies which showed yellow-green colour under daylight and strong green fluorescence under long-wave ultraviolet light. It can be a useful vector for selecting foreign DNA fragment which was inserted into multiple cloning site based on the loss of the yellow-green color/green fluorescence of E. coli cells attributable to the insertional inactivation of GFP production.     

Pressate from Peat Dewatering as a Substrate for Bacterial Growth

This study considered the possibility of using water expressed during the drying of fuel-grade peat as a substrate for microbial growth. Highly humified peat pressed for 2.5 min at 1.96 MPa produced water with a chemical oxygen demand of 690 mg/liter. Several biological compounds could be produced by using the organic matter in expressed peat water as a substrate. These included polymers such as chitosan, contained in the cell wall of Rhizopus arrhizus, and two extracellular polysaccharides, xanthan gum and pullulan, produced by Xanthomonas campestris and Aureobasidium pullulans, respectively. A very effective surfactant was produced by Bacillus subtilis grown in the expressed water. Small additions of nutrients to the peat pressate were necessary to obtain substantial yields of products. The addition of peptone, yeast extract, and glucose improved production of the various compounds. Biological treatment improved the quality of the expressed water to the extent that in an industrial process it could be returned to the environment.     

厌氧污泥体系脱氢酶活性表征细菌数的研究

以硫化钠作还原剂,甲苯为提取溶剂,用氯化三苯基四氮唑(TTC)法测定污泥脱氢酶活性,探讨了脱氢酶活性与细菌数对数(lgABN)的相关性,并建立了脱氢酶活性表征细菌数的数学模型。结果表明:在lgABN为7.5～12范围内,脱氢酶表征的污泥活性与细菌数对数(lgABN)呈明显线性相关,从理论上分析了其惟一线性相关性;在细菌数的对数达到7.5～12的对数期、稳定期与衰亡期,脱氢酶活性换算成对应细菌数所绘制的曲线与细菌生长曲线有明显拟合;在有重金属离子存在的污泥体系中,脱氢酶活性可用来作为评价重金属离子的毒性指标;从而脱氢酶活性可取代细菌计数表征污泥活性。     

Biodegradability of Vegetation-Derived Dissolved Organic Carbon in a Cool Temperate Ombrotrophic Bog

Dissolved organic carbon (DOC) plays a key role in the peatland carbon balance and serves numerous ecological and chemical functions including acting as a microbial substrate. In this study, we quantify the concentration, biodegradability, and intrinsic properties of DOC obtained from peat, fresh material, and litter from nine species of ombrotrophic bog vegetation. Potential biodegradability was assessed by incubating vegetation extracts for 28 days in the dark and measuring percent DOC loss as the fraction of biodegradable DOC (%BDOC) while DOC properties were characterized using UV–Vis absorbance and fluorescence measurements. The mean initial DOC concentration extracted differed significantly among species (P < 0.05) and was significantly higher in fresh material, 217 ± 259 mg DOC l^?1, than either litter or peat extracts with mean concentrations of 82.1 ± 117 mg DOC l^?1 and 12.7 ± 1.0 mg DOC l^?1, respectively (P < 0.05). %BDOC also differed significantly among species (P < 0.05) and ranged from 52 to 73% in fresh cuttings with the greatest fraction observed in S. magellanicum; 22–46% in litter; and 24% in peat. The majority of variability (82.5%) in BDOC was explained by initial absorbance at 254 nm and total dissolved nitrogen concentration which was further resolved into significant non-linear relationships between %BDOC and both humic-like and protein-like DOC fractions (P < 0.05). Our results highlight the extremely heterogeneous nature of the surface vegetation-derived DOC input in peatlands and stress the importance of vegetation species in peatland ecosystem function.     

Effect of Organic Complex Compounds on Bacillus thermoamylovorans Growth and Glucose Fermentation

The effect of the concentration of a mixture (1/1 [wt/wt]) of yeast extract and bioTrypcase (YE+bT) on the growth and physiology of a new species, Bacillus thermoamylovorans, a moderately thermophilic, non-spore-forming, lactic acid-producing bacterium isolated from palm wine, was studied. At an initial glucose concentration of 100 mM, B. thermoamylovorans growth was limited when the concentration of YE+bT was lower than 5.0 g liter⁻¹; under these conditions, cellular yield reached a maximum value of 0.4 g of cells per g of YE+bT. Growth limitation due to deficiency in growth factors led to a significant shift in glucose metabolism towards lactate production. Lactate constituted 27.5 and 76% of the end products of glucose fermentation in media containing YE+bT at 20.0 and 1.0 g liter⁻¹, respectively. This result markedly differed from published data for lactic bacteria, which indicated that fermentative metabolism remained homolactic regardless of the concentration of YE. Our results showed that the ratio between cellular synthesis and energy production increased with the concentration of YE+bT in the culture medium. They indicate that the industrial production of lactic acid through glucose fermentation by B. thermoamylovorans can be optimized by using a medium where glucose is present in excess and the organic additives are limiting.     

Bacterial Accumulation of Metallic Compounds

Xanthomonas maltophilia accumulated metals or metallic compounds within the cell in the form of one or a few particles, in a peculiar condition similar to that of an electrolyte solution in an electric cell. The particles consisted of many little granules, most of which were assumed to be composed mainly of a compound of zinc and phosphorus in an amorphous state.     

Bacterial Degradation of Detergent Compounds

A survey for surfactant degradation among aerobic bacteria has been undertaken. Tests have been made in peptone medium where such a degradation, if it occurs, will be gratuitous. Tallow-alkyl-sulfate, alkyl-ethoxylate-sulfate, and linear-alkyl-benzene-sulfonate were used. Forty-five strains of 34 species in 19 genera degrade one or more of these detergent compounds. With some species, the surfactant inhibits degradation without inhibiting growth, whereas with one species slight degradation took place even at a toxic concentration of surfactant.     

Extensive Identification of Bacterial Riboflavin Transporters and Their Distribution across Bacterial Species

Riboflavin, the precursor for the cofactors flavin mononucleotide (FMN) and flavin adenine dinucleotide, is an essential metabolite in all organisms. While the functions for de novo riboflavin biosynthesis and riboflavin import may coexist in bacteria, the extent of this co-occurrence is undetermined. The RibM, RibN, RfuABCD and the energy-coupling factor-RibU bacterial riboflavin transporters have been experimentally characterized. In addition, ImpX, RfnT and RibXY are proposed as riboflavin transporters based on positional clustering with riboflavin biosynthetic pathway (RBP) genes or conservation of the FMN riboswitch regulatory element. Here, we searched for the FMN riboswitch in bacterial genomes to identify genes encoding riboflavin transporters and assessed their distribution among bacteria. Two new putative riboflavin transporters were identified: RibZ in Clostridium and RibV in Mesoplasma florum. Trans-complementation of an Escherichia coli riboflavin auxotroph strain confirmed the riboflavin transport activity of RibZ from Clostridium difficile, RibXY from Chloroflexus aurantiacus, ImpX from Fusobacterium nucleatum and RfnT from Ochrobactrum anthropi. The analysis of the genomic distribution of all known bacterial riboflavin transporters revealed that most occur in species possessing the RBP and that some bacteria may even encode functional riboflavin transporters from two different families. Our results indicate that some species possess ancestral riboflavin transporters, while others possess transporters that appear to have evolved recently. Moreover, our data suggest that unidentified riboflavin transporters also exist. The present study doubles the number of experimentally characterized riboflavin transporters and suggests a specific, non-accessory role for these proteins in riboflavin-prototrophic bacteria.     

Kinetics of Utilization of Organic Compounds in the Growth of Mycobacterium tuberculosis.

Bowles, Jean A. (University of Colorado School of Medicine, Denver), and William Segal. Kinetics of utilization of organic compounds in the growth of Mycobacterium tuberculosis. J. Bacteriol. 90:157-163. 1965.-To obtain a workable system for a study of the kinetics of nutrient utilization (based on specific quantitative assay) by Mycobacterium tuberculosis, several cultural refinements were introduced: the use of shake culture, a 40-fold increase in the size of inoculum, substitution of glutamate for asparagine as nitrogen source, and elimination of glucose from the medium with glycerol remaining as carbon source. These modifications resulted in reduction to a tenth of the lag phase of glycerol utilization (from 40 to 4 days), and in a greatly increased rate of growth. Both coordinate and sequential patterns of nutrient utilization were in evidence, except in the case of citrate, which was never utilized under a variety of conditions of culture. The coordinate pattern of glucose-glutamate and glucose-glycerol utilization would appear to rule out catabolite repression by glucose. However, elimination of glucose from the medium resulted in elimination of the 4-day lag period before glutamate utilization was initiated, leaving open to question the role of glucose in this system. Evidence is presented for the hypothesis that the sequential pattern of glutamate-glycerol utilization is a function of glutamate repression of glycerol oxidation in the growth of M. tuberculosis, although no diauxie effect is apparent. In a determination of which nutrient-utilization systems were regulated by induction, only in the case of glycerol was evidence obtained for an inducible system. The enzymatic mechanisms underlying these patterns of nutrient utilization are presently being investigated.     

The Effects of Cyanobacterial Exudates on Bacterial Growth and Biodegradation of Organic Contaminants

The pulp and paper industry largely depends on the biodegradation activities of heterotrophic bacteria to remove organic contaminants in wastewater prior to discharge. Our recent discovery of extensive cyanobacterial communities in pulp and paper waste treatment systems led us to investigate the potential impacts of cyanobacterial exudates on growth and biodegradation efficiency of three bacterial heterotrophs. Each of the three assessed bacteria represented different taxa commonly found in pulp and paper waste treatment systems: a fluorescent Pseudomonad, an Ancylobacter aquaticus strain, and a Ralstonia eutropha strain. They were capable of utilizing phenol, dichloroacetate (DCA), or 2,4-dichlorophenoxyacetic acid (2,4-D), respectively. Exudates from all 12 cyanobacterial strains studied supported the growth of each bacterial strain to varying degrees. Maximum biomass of two bacterial strains positively correlated with the total organic carbon content of exudate treatments. The combined availability of exudate and a known growth substrate (i.e., phenol, DCA, or 2,4-D) generally had a synergistic affect on the growth of the Ancylobacter strain, whereas mixed effects were seen on the other two strains. Exudates from four representative cyanobacterial strains were assessed for their impacts on phenol and DCA biodegradation by the Pseudomonas and Ancylobacter strains, respectively. Exudates from three of the four cyanobacterial taxa repressed phenol biodegradation, but enhanced DCA biodegradation. These dissimilar impacts of cyanobacterial exudates on bacterial degradation of contaminants suggest a species-specific association, as well as a significant role for cyanobacteria during the biological treatment of wastewaters.     

Volatile Organic Compounds Associated with Microbial Growth in Automobile Air Conditioning Systems

Volatile organic compounds from Penicillium viridicatum and Methylobacterium mesophilicum growing on laboratory media and on component materials of automobile air conditioners were analyzed with gas chromatography and mass spectrometry. P. viridicatum produced compounds such as 4-methyl thiazole, terpenes and alcohols, whereas M. mesophilicum produced dimethyl disulfide, dimethyl trisulfide, and chlorophenol with growth on laboratory media. In comparison with laboratory media, fewer volatiles were detected from colonized foam insulation materials. Biofilms of M. mesophilicum on aluminum evaporator components produced mainly dimethyl disulfide. These biofilms, after inoculation with P. viridicatum, produced offensive smelling alcohols and esters such as 2-methyl propanol, 3-penten-2-ol, and the ethyl ester of butanoic acid. The moisture and substrates innate to the automobile air conditioning systems provided an environment suitable for microbial biofilm development and odor production. Reduction of retained moisture in the air conditioning system coupled with use of less susceptible or antimicrobial substrates are advised for remediation of the noxious odors. Received: 26 February 2000 / Accepted: 2 May 2000     

Modelling Bacterial Growth of Lactobacillus curvatus as a Function of Acidity and Temperature

Models that describe the effect of acidity, temperature, and the combined effect of these variables on the growth parameters of Lactobacillus curvatus are developed and validated. Growth parameters (lag time, specific growth rate, and maximum population density) were calculated from growth data at different temperature-acidity combinations. Experiments were set up to assess the quantitative effects of temperature and acidity on the growth parameters rather than for parameter estimation solely. The effect of acidity is monitored at several constant temperature values. Models are set up and fitted to the data. The same procedure is used at constant acidity values to model the effect of temperature. For lag time, specific growth rate, and maximum population density, the effect of temperature could be multiplied with the effect of acidity to obtain combinatory models that describe the effect of both controlling factors on the growth parameters. Lag time measurements showed large deviations, and therefore the lag time models developed can only be used to estimate the order of magnitude of lag time.     

Identifying Bioaccumulative Halogenated Organic Compounds Using a Nontargeted Analytical Approach: Seabirds as Sentinels

Persistent organic pollutants (POPs) are typically monitored via targeted mass spectrometry, which potentially identifies only a fraction of the contaminants actually present in environmental samples. With new anthropogenic compounds continuously introduced to the environment, novel and proactive approaches that provide a comprehensive alternative to targeted methods are needed in order to more completely characterize the diversity of known and unknown compounds likely to cause adverse effects. Nontargeted mass spectrometry attempts to extensively screen for compounds, providing a feasible approach for identifying contaminants that warrant future monitoring. We employed a nontargeted analytical method using comprehensive two-dimensional gas chromatography coupled to time-of-flight mass spectrometry (GC×GC/TOF-MS) to characterize halogenated organic compounds (HOCs) in California Black skimmer (Rynchops niger) eggs. Our study identified 111 HOCs; 84 of these compounds were regularly detected via targeted approaches, while 27 were classified as typically unmonitored or unknown. Typically unmonitored compounds of note in bird eggs included tris(4-chlorophenyl)methane (TCPM), tris(4-chlorophenyl)methanol (TCPMOH), triclosan, permethrin, heptachloro-1''-methyl-1,2''-bipyrrole (MBP), as well as four halogenated unknown compounds that could not be identified through database searching or the literature. The presence of these compounds in Black skimmer eggs suggests they are persistent, bioaccumulative, potentially biomagnifying, and maternally transferring. Our results highlight the utility and importance of employing nontargeted analytical tools to assess true contaminant burdens in organisms, as well as to demonstrate the value in using environmental sentinels to proactively identify novel contaminants.