The value of basic research: discovery of Thermus aquaticus and other extreme thermophiles.
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Genetics↗
Biomedical subjects
T D Brock
Publications and source records attributed to T D Brock.
The road to Yellowstone--and beyond.
This memoir describes the professional life and times of Thomas D. Brock, with an emphasis on those aspects of his career relating to research in microbial ecology, and how this work led to field research in Yellowstone. The first discovery of extremely thermophilic bacteria is described, followed by a discussion of some of the consequences of this discovery for biotechnology and microbiology. Also covered briefly in this memoir are Brock's activities in textbook writing, publishing, computers, and the history of science.
Bacteria↗
The bacterial nucleus: a history.
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Bacteria↗
Significance of algal excretory products for growth of epilimnetic bacteria.
Light-stimulated uptake of CO(2) and differential filtration through Nucleopore filters were used to estimate the significance of phytoplankton excretion as a source of bacterial carbon in water samples collected at different seasons of the year in Lake Mendota, Wis. On an annual basis, about 14% of the estimated bacterial production was accounted for by algal excretion, although at certain times of year the fraction of bacterial carbon derived from algal excretion was considerably higher. About 20% of the annual primary production was estimated to pass through the bacterial component.
Journal Article↗
Assessing biomass and production of bacteria in eutrophic lake mendota, wisconsin.
Estimates were made of the biomass and production of heterotrophic bacteria in the epilimnion of Lake Mendota, Wis. Cell counts were done with epifluorescence microscopy and varied from 3 x 10 bacteria per ml in winter to 3 x 10 bacteria per ml in summer. Cell volumes were measured in scanning electron micrographs. The average cell volume was 0.159 mum. Annual variations and depth distribution were studied. Production was estimated from the frequency of dividing cells and from dark radioactive sulfate uptake. Annual productivity and daily average productivity were very close with both methods: 107 to 205 g of C per m per year for sulfate and 89 to 117 g of C per m per year for frequency of dividing cells. Zooplankton feeding removed 2 to 10% of the bacterial net production annually. When compared with biomass changes and losses due to zooplankton feeding, production values were very high. Therefore, it was suggested that other loss factors have to be more important than zooplankton feeding in controlling the bacterial population. Bacterial heterotrophic production was about 50% of gross primary production.
Journal Article↗
Dynamics of bacterial sulfate reduction in a eutrophic lake.
Bacterial sulfate reduction in the surface sediment and the water column of Lake Mendota, Madison, Wis., was studied by using radioactive sulfate (SO(4)). High rates of sulfate reduction were observed at the sediment surface, where the sulfate pool (0.2 mM SO(4)) had a turnover time of 10 to 24 h. Daily sulfate reduction rates in Lake Mendota sediment varied from 50 to 600 nmol of SO(4) cm, depending on temperature and sampling date. Rates of sulfate reduction in the water column were 10 times lower than that for the surface sediment and, on an areal basis, accounted for less than 18% of the total sulfate reduction in the hypolimnion during summer stratification. Rates of bacterial sulfate reduction in the sediment were not sulfate limited at sulfate concentrations greater than 0.1 mM in short-term experiments. Although sulfate reduction seemed to be sulfate limited below 0.1 mM, Michaelis-Menten kinetics were not observed. The optimum temperature (36 to 37 degrees C) for sulfate reduction in the sediment was considerably higher than in situ temperatures (1 to 13 degrees C). The response of sulfate reduction to the addition of various electron donors metabolized by sulfate-reducing bacteria in pure culture was investigated. The degree of stimulation was in this order: H(2) > n-butanol > n-propanol > ethanol > glucose. Acetate and lactate caused no stimulation.
Journal Article↗
Characterization of an acetate-decarboxylating, non-hydrogen-oxidizing methane bacterium.
A methanogenic bacterium, commonly seen in digested sludge and referred to as the "fat rod" or Methanobacterium soehngenii, has been enriched to a monoculture and is characterized. Cells are gramnegative, non-motile and appear as straight rods with flat ends. They form filaments which can grow to great lengths. The structure of the outer cell envelop is similar to Methanospirillum hungatii. The organism grows on a mineral salt medium with acetate as the only organic component. Acetate is the energy source, and methane is formed exclusively from the methyl group. Acetate and carbon dioxide act as sole carbon source and are assimilated in a molar ratio of about 1.9:1. The reducing equivalents necessary to build biomass from these two precursors are obtained from the total oxidation of some acetate. Hydrogen is not used for methane formation and is not needed for growth. Formate is cleaved into hydrogen and carbon dioxide. Coenzyme M was found to be present at levels of 0.35 nmol per mg of dry cells and F420 amounted to 0.55 microgram per mg protein. The mean generation time was 9 days at 33 degrees C.
Acetates↗
Anaerobic methane oxidation: occurrence and ecology.
Anoxic sediments and digested sewage sludge anaerobically oxidized methane to carbon dioxide while producing methane. This strictly anaerobic process showed a temperature optimum between 25 and 37 degrees C, indicating an active microbial participation in this reaction. Methane oxidation in these anaerobic habitats was inhibited by oxygen. The rate of the oxidation followed the rate of methane production. The observed anoxic methane oxidation in Lake Mendota and digested sewage sludge was more sensitive to 2-bromoethanesulfonic acid than the simultaneous methane formation. Sulfate diminished methane formation as well as methane oxidation. However, in the presence of iron and sulfate the ratio of methane oxidized to methane formed increased markedly. Manganese dioxide and higher partial pressures of methane also stimulated the oxidation. The rate of methane oxidation in untreated samples was approximately 2% of the CH(4) production rate in Lake Mendota sediments and 8% of that in digested sludge. This percentage could be increased up to 90% in sludge in the presence of 10 mM ferrous sulfate and at a partial pressure of methane of 20 atm (2,027 kPa).
Journal Article↗
Biological energy production in the apparent absence of electron transport and substrate level phosphorylation.
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Acetates↗
Decomposition of blue-green algal (cyanobacterial) blooms in lake mendota, wisconsin.
Decomposition of natural populations of Lake Mendota phytoplankton dominated by blue-green algae (cyanobacteria) was monitored by using oxygen uptake and disappearance of chlorophyll, algal volume (fluorescence microscopy), particulate protein, particulate organic carbon, and photosynthetic ability (CO(2) up-take). In some experiments, decomposition of C-labeled axenic cultures of Anabaena sp. was also measured. In addition to decomposition, mineralization of inorganic nitrogen and phosphorus were followed in some experiments. Decomposition could be described as a first-order process, and the rate of decomposition was similar to that found by others using pure cultures of eucaryotic algae. Nitrogen and phosphorus never limited the decomposition process, even when the lake water was severely limited in soluble forms of these nutrients. This suggests that the bacteria responsible for decomposition can obtain all of their key nutrients for growth from the blue-green algal cells. Filtration of lake water through plankton netting that removed up to 90% of the algal biomass usually did not cause a similar decrease in oxygen demand, suggesting that most of the particulate organic matter used for respiration of the decomposing bacteria was in a small-particle fraction. Short-term oxygen demand correlated well with the particulate chlorophyll concentration of the sample, and a relationship was derived that could be used to predict community respiration of the lake from chlorophyll concentration. Kinetic analysis showed that not all analyzed components disappeared at the same rate during the decomposition process. The relative rates of decrease of the measured parameters were as follows: photosynthetic ability > algal volume > particulate chlorophyll > particulate protein. Decomposition of C-labeled Anabaena occurred at similar rates with aerobic epilimnetic water and with anaerobic sediment, but was considerably slower with anaerobic hypolimnetic water. Of the various genera present in the lake, Aphanizomenon and Anabaena were more sensitive to decomposition than was Microcystis. In addition to providing a general picture of the decomposition process, the present work relates to other work on sedimentation to provide a detailed picture of the fate of blue-green algal biomass in a eutrophic lake ecosystem.
Journal Article↗
Measuring radioactive methane with the liquid scintillation counter.
Although a gas proportional counter is the most convenient method of measuring the radioactivity of fixed gases such as methane, it cannot be used when high nonradioactive concentrations of methane are present in the gas phase, due to quenching. If only methane and carbon dioxide are present in radioactive form in the gas phase, a liquid scintillation method for measuring these substances can be used. The procedure is described in detail, and the solubility of methane in liquid scintillation cocktails is determined.
Journal Article↗
Lytic organisms and photooxidative effects: influence on blue-green algae (cyanobacteria) in lake mendota, wisconsin.
The effects of exposure to high light intensities on blue-green algal (cyanobacterial) populations were examined in Lake Mendota, Wis. The algal populations were shown to be susceptible to inhibition of photosynthetic activity and pigment bleaching as a result of exposure. These effects generally influence only a small percentage of the lake population and thus are probably not important in causing major declines in chlorophyll a. Lytic organisms were shown to increase in numbers in the lake in response to the seasonal development of blue-green algae, reaching values of greater than 1,000 plaque-forming units per ml in midsummer. Both bacteria and protozoa were observed in plaque zones, but it could not be determined whether these lytic organisms had a major role in algal biomass declines.
Journal Article↗
Methane formation and methane oxidation by methanogenic bacteria.
Methanogenic bacteria were found to form and oxidize methane at the same time. As compared to the quantity of methane formed, the amount of methane simultaneously oxidized varied between 0.3 and 0.001%, depending on the strain used. All the nine tested strains of methane producers (Methanobacterium ruminantium, Methanobacterium strain M.o.H., M. formicicum, M. thermoautotrophicum, M. arbophilicum, Methanobacterium strain AZ, Methanosarcina barkeri, Methanospirillum hungatii, and the "acetate organism") reoxidized methane to carbon dioxide. In addition, they assimilated a small part of the methane supplied into cell material. Methanol and acetate also occurred as oxidation products in M. barkeri cultures. Acetate was also formed by the "acetate organism," a methane bacterium unable to use methanogenic substrates other than acetate. Methane was the precursor of the methyl group of the acetate synthesized in the course of methane oxidation. Methane formation and its oxidation were inhibited equally by 2-bromoethanesulfonic acid. Short-term labeling experiments with M. thermoautotrophicum and M. hungatii clearly suggest that the pathway of methane oxidation is not identical with a simple back reaction of the methane formation process.
Acetates↗
Dimethyl sulfoxide as an electron acceptor for anaerobic growth.
The isolation from lake mud of a bacterium which can use dimethyl sulfoxide (DMSO) as an electron acceptor for growth is described. The isolate, called strain DL-1, was a small, gram negative, non-motile spiral. The sole product of DMSO reduction was dimethyl sulfide (DMS). Other electron acceptors used by the isolate included sulfite, thiosulfate, elemental sulfur, methionine sulfoxide, tetramethylene sulfoxide, nitrate, and oxygen (microaerophilically). Sulfate was not reduced and could not even be assimilated. Lactate or succinate could serve as electron donors, with acetate as the main product. Hydrogen could be used as an electron donor if acetate was present in the medium as a carbon source. The organism has a c-type cytochrome, and most likely uses electron transport phosphorylation during DMSO reduction. Cultures of Desulfovibrio sp., Escherichia coli, Pseudomonas aeruginosa, and Proteus vulgaris were tested for growth using DMSO as an electron acceptor, and only the Proteus strain grew. Both Proteus and strain DL-1 are versatile at coupling reductions with energy generation. There is a marked resemblance between strain DL-1 and the recently described sulfur-reducing spirillum of Wolfe and Pfennig.
Bacteria↗
Dimethyl sulphoxide reduction by micro-organisms.
Dimethyl sulphoxide (DMSO) was reduced to dimethyl sulphide by a wide variety of micro-organism, including prokaryotes and eukaryotes, aerobes and anaerobes. Dimethyl sulphone was not reduced by any of the organisms tested. Cell-free extracts of Escherichia coli reduced DMSO using reduced pyridine nucleotides as electron donors. Activity was greater in anaerobically grown cells than in those grown aerobically. Two other sulphoxides, methionine sulphoxide and tetramethylene sulphoxide, substantially inhibited DMSO reduction by extracts. Mutants of E. coli, which were unable to reduce biotin sulphoxide to biotin, were tested for their ability to reduce DMSO in whole cells and extracts. These mutants were in four different gene loci, bisA to bisD. DMSO reductase activity of the mutants was generally less than that of the wild-type strain, and activity depended upon the gene locus involved, the growth medium and the growth conditions. Only the bisA mutant had very low activity under all conditions. All of the bis mutants were able to grow using methionine sulphoxide as a sulphur source, indicating that biotin sulphoxide and methionine sulphoxide are reduced by different enzyme systems. DMSO may be reduced by both of these enzyme systems.
Anaerobiosis↗
Methane, carbon dioxide, and hydrogen sulfide production from the terminal methiol group of methionine by anaerobic lake sediments.
A significant portion of the sulfide in lake sediments may be derived from sulfur-containing amino acids. Methionine degradation in Lake Mendota (Wisconsin) sediments was studied with gas chromatographic and radiotracer techniques. Temperature optimum and inhibitor studies showed that this process was biological. Methane thiol and dimethyl sulfide were produced in sediments when 1-mumol/ml unlabeled methionine was added. When chloroform (an inhibitor of one-carbon metabolism) was added to the sediments, methane thiol, carbon disulfide, and n-propane thiol were produced, even when no methionine was added. When S-labeled methionine was added to the sediments in tracer quantities (1.75 nmol/ml), labeled hydrogen sulfide was produced, and a roughly equal amount of label was incorporated into insoluble material. Methane and carbon dioxide were produced from [methyl-C]methionine. Evidence is given favoring methane thiol as an intermediate in the formation of methane, carbon dioxide, and hydrogen sulfide from the terminal methiol group of methionine. Methionine may be an important source of sulfide in lake sediments.
Journal Article↗
Hydrocarbon biodegradation in hypersaline environments.
When mineral oil, hexadecane, and glutamate were added to natural samples of varying salinity (3.3 to 28.4%) from salt evaporation ponds and Great Salt Lake, Utah, rates of metabolism of these compounds decreased as salinity increased. Rate limitations did not appear to relate to low oxygen levels or to the availability of organic nutrients. Some oxidation of l-[U-C]glutamic acid occurred even at extreme salinities, whereas oxidation of [1-C]hexadecane was too low to be detected. Gas chromatographic examination of hexane-soluble components of tar samples from natural seeps at Rozel Point in Great Salt Lake demonstrated no evidence of biological oxidation of isoprenoid alkanes subject to degradation in normal environments. Some hexane-soluble components of the same tar were altered by incubation in a low-salinity enrichment culture inoculated with garden soil. Attempts to enrich for microorganisms in saline waters able to use mineral oil as a sole source of carbon and energy were successful below, but not above, about 20% salinity. This study strongly suggests a general reduction of metabolic rate at extreme salinities and raises doubt about the biodegradation of hydrocarbons in hypersaline environments.
Journal Article↗