bluuebunny/crossref_metadata_2025 huggingface dataset:
Production and Further Characterization of an Alkaline Elastase Produced by Alkalophilic
Bacillus
Strain Ya-B
Authors: Tsai Ying-Chieh , Juang Ray-Yeng , Lin Sheun-Fuh , Chen Shiow-Wen , Yamasaki Makari , Tamura Gakuzo
Published: January 2020

The characteristics of the obligate alkalophilic
Bacillus
sp. strain Ya-B, which produces alkaline elastase extracellularly, were examined. This strain grew at pH 7.0 only in the presence of 1% or more NaCl. Its fatty acid distribution pattern was similar to that of other
Bacillus
species in which iso-C
15
and anteiso-C
15
were the most abundant fatty acids. About 120 mg of enzyme was recovered from 1 liter of culture broth in a medium (pH 10.1) containing mainly glucose, soymeal, and glycerol. The antiserum against this enzyme did not recognize microbial proteinases, such as subtilisins, but reacted with proteinase C, which was purified from commercial pronase. Chemical modification studies revealed that certain histidine and tyrosine residues might be involved in the enzyme activity. This enzyme underwent a partial unfolding at pHs higher than 12.0, as indicated by the circular dichroism study.
Transfer and Expression of Lithoautotrophy and Denitrification in a Host Lacking These Metabolic Activities
Authors: Schneider Beate , Nies Anke , Friedrich Bärbel
Published: January 2020

The conjugative 450-kilobase-pair megaplasmid pHG1 from
Alcaligenes eutrophus
H16 was transferred to the herbicide-degrading soil bacterium
A. eutrophus
JMP134. This transfer was achieved by means of RP4 mobilization and a Tn
5
-Mob insertion provided in
trans
on the megaplasmid replicon. Although kanamycin-resistant transconjugants also occurred with other gram-negative species such as
Rhizobium, Agrobacterium
, and thiobacteria,
A. eutrophus
JMP134 was the only recipient which stably maintained the megaplasmid. pHG1-containing transconjugants derived from JMP134 expressed all metabolic functions associated with the plasmid: the ability to oxidize hydrogen through catalysis of two hydrogenases, to assimilate carbon dioxide via the Calvin cycle pathway, and to grow with nitrate anaerobically. All of these metabolic activities were absent in the original strain JMP134.
Interaction of Ammonia Monooxygenase from
Nitrosomonas europaea
with Alkanes, Alkenes, and Alkynes
Authors: Hyman Michael R. , Murton Ian B. , Arp Daniel J.
Published: January 2020

Ammonia monooxygenase of
Nitrosomonas europaea
catalyzes the oxidation of alkanes (up to C
8
) to alcohols and alkenes (up to C
5
) to epoxides and alcohols in the presence of ammonium ions. Straight-chain, N-terminal alkynes (up to C
10
) all exhibited a time-dependent inhibition of ammonia oxidation without effects on hydrazine oxidation.
Ethanol Extraction Requirement for Purification of Protein Labeled with [
3
H]Leucine in Aquatic Bacterial Production Studies
Authors: Wicks Richard J. , Robarts Richard D.
Published: January 2020

The trichloroacetic acid (TCA)-insoluble fraction of water column bacteria labeled with [
3
H]leucine contained an ethanol-soluble fraction accounting for up to 44% of the label. A component of the ethanol-soluble fraction is [
3
H]leucine. Labeled-protein purification requires an ethanol wash step. Cold TCA can replace hot TCA for precipitation of labeled proteins.
Reductive dechlorination of hexachlorobenzene to tri- and dichlorobenzenes in anaerobic sewage sludge
Authors: Fathepure B Z , Tiedje J M , Boyd S A
Published: January 2020

Hexachlorobenzene was dechlorinated to tri- and dichlorobenzenes in anaerobic sewage sludge. The complete biotransformation of 190 microM hexachlorobenzene (approximately 50 ppm) occurred within 3 weeks. The calculated rate of hexachlorobenzene dechlorination was 13.6 mumol liter-1 day-1. Hexachlorobenzene was dechlorinated via two routes, both involving the sequential removal of chlorine from the aromatic ring. The major route was hexachlorobenzene----pentachlorobenzene----1,2,3,5-tetrachlorobenzene--- -1,3,5- trichlorobenzene. Greater than 90% of the added hexachlorobenzene was recovered as 1,3,5-trichlorobenzene, and there was no evidence for further dechlorination of 1,3,5-trichlorobenzene. The minor route was hexachlorobenzene----pentachlorobenzene----1,2,4,5-tetrachlorobenzene--- -1,2,4- trichlorobenzene----dichlorobenzenes. These results extend reductive dechlorination to poorly water soluble aromatic hydrocarbons which could potentially include other important environmental pollutants like polychlorinated biphenyls.
Effect of 5-Fluoro-2′-Deoxyuridine on [
3
H]Thymidine Incorporation by Bacterioplankton in the Waters of Southwest Florida
Authors: Jeffrey Wade H. , Paul John H.
Published: January 2020

The effect of 5-fluoro-2′-deoxyuridine (FdUrd) on [
methyl
-
3
H] thymidine incorporation by bacterioplankton populations in subtropical freshwater, estuarine, and oceanic environments was examined. In estuarine waters, intracellular isotope dilution was inhibited by FdUrd, which enabled us to estimate both intracellular and extracellular isotope dilution. In 2 of 10 cases, extracellular isotope dilution was significant. At low concentrations of [
methyl
-
3
H]thymidine or [6-
3
H]thymidine, FdUrd completely inhibited incorporation of radioactivity into protein and RNA. At high concentrations of [
3
H]thymidine, however, FdUrd had little effect on labeling patterns. The dihydrofolate reductase inhibitors amethopterin and trimethoprim had no effect on macromolecular labeling patterns. These results suggest that thymidylate synthase is not involved in nonspecific labeling and that FdUrd inhibits nonspecific labeling by blocking some other enzyme involved in thymidine catabolism. In oligotrophic oceanic and freshwater samples, FdUrd did not inhibit intracellular isotope dilution or [
3
H]thymidine labeling of protein and RNA, but caused some inhibition of [
3
H]thymidine incorporation into DNA. The ability of FdUrd to inhibit nonspecific macromolecular labeling during [
3
H]thymidine incorporation was significantly correlated (
r
= 0.84) with total thymidine incorporation (in picomoles per liter per hour). The results are discussed in terms of applications of FdUrd to routine bacterial production measurements and the general assumptions of [
3
H]thymidine incorporation.
Transposon Tn
5
-Generated
Bradyrhizobium japonicum
Mutants Unable To Grow Chemoautotrophically with H
2
Authors: Hom Sherman Siu Ming , Novak Patricia D. , Maier Robert J.
Published: January 2020

Twelve Tn
5
-induced mutants of
Bradyrhizobium japonicum
unable to grow chemoautotrophically with CO
2
and H
2
(Aut
−
) were isolated. Five Aut
−
mutants lacked hydrogen uptake activity (Hup
−
). The other seven Aut
−
mutants possessed wild-type levels of hydrogen uptake activity (Hup
+
), both in free-living culture and symbiotically. Three of the Hup
−
mutants lacked hydrogenase activity both in free-living culture and as nodule bacteroids. The other two mutants were Hup
−
only in free-living culture. The latter two mutants appeared to be hypersensitive to repression by oxygen, since Hup activity could be derepressed under 0.4% O
2
. All five Hup
−
mutants expressed both ex planta and symbiotic nitrogenase activities. Two of the seven Aut
−
Hup
+
mutants expressed no free-living nitrogenase activity, but they did express it symbiotically. These two strains, plus one other Aut
−
Hup
+
mutant, had CO
2
fixation activities 20 to 32% of the wild-type level. The cosmid pSH22, which was shown previously to contain hydrogenase-related genes of
B. japonicum
, was conjugated into each Aut
−
mutant. The Aut
−
Hup
−
mutants that were Hup
−
both in free-living culture and symbiotically were complemented by the cosmid. None of the other mutants was complemented by pSH22. Individual subcloned fragments of pSH22 were used to complement two of the Hup
−
mutants.
Production of Formaldehyde by Detergent-Treated Cells of a Methanol Yeast,
Candida boidinii
S2 Mutant Strain AOU-1
Authors: Sakai Yasuyoshi , Tani Yoshiki
Published: January 2020

Treatment of cells of a methanol yeast,
Candida boidinii
, with the cationic detergent cetyldimethylbenzyl-ammonium chloride (Cation M2) improved the production of formaldehyde. Formaldehyde production was improved twofold with respect to the initial amount of formaldehyde and 1.61-fold with respect to the final amount of formaldehyde after a 12-h reaction under optimized detergent treatment conditions. The treatment caused formaldehyde and formate dehydrogenases to leak out of the cells more rapidly than catalase, but there was no leakage of alcohol oxidase. The improvement in formaldehyde production was considered to be due to the increased permeability of yeast cell membranes and to lower activities of formaldehyde and formate dehydrogenases in Cation M2-treated cells than in intact cells. Changes in the ultrastructure of the cells were observed upon Cation M2 treatment. Several developed peroxisomes were observed in intact cells. After Cation M2 treatment, the cells were obviously damaged, and several peroxisomes seemed to have fused with each other.
Methylation of Halogenated Phenols and Thiophenols by Cell Extracts of Gram-Positive and Gram-Negative Bacteria
Authors: Neilson Alasdair H. , Lindgren Carin , Hynning Per-Åke , Remberger Mikael
Published: January 2020

O-methylation of 2,6-dibromophenol was studied in cell extracts prepared from
Rhodococcus
sp. strain 1395. O-methylation activity was enhanced by the addition of
S
-adenosyl-
l
-methionine but was not affected by the addition of 5-methyltetrahydrofolate nor by up to 10 mM MgCl
2
or EDTA. By using 2,6-dibromophenol, 4,5,6-trichloroguaiacol, and pentachlorothiophenol as the substrates, O-methylation activity was also demonstrated in extracts from two other
Rhodococcus
sp. strains, an
Acinetobacter
sp. strain, and a
Pseudomonas
sp. strain. A diverse range of chloro- and bromophenols, chlorothiophenols, chloro- and bromoguaiacols, and chloro- and bromocatechols were assayed as the substrates by using extracts prepared from strain 1395; all of the compounds were methylated to the corresponding anisoles, veratroles, or guaiacols, which have been identified previously from experiments using whole cells. The specific activity of the enzyme towards the thiophenols was significantly higher than it was towards all the other substrates—high activity was found with pentafluorothiophenol, although the activity with pentafluorophenol was undetectable with the incubation times used. For the chlorophenols, the position of the substituents was of cardinal importance. The enzyme had higher activity towards the halogenated catechols than towards the corresponding guaiacols, and selective O-methylation of the 3,4,5-trihalogenocatechols yielded predominantly the 3,4,5-trihalogenoguaiacols. As in experiments with whole cells, neither 2,4-dinitrophenol, hexachlorophene, nor 5-chloro- or 5-bromovanillin was O-methylated. The results showed conclusively that the methylation reactions were enzymatic and confirmed the conclusion from extensive studies using whole cells that methylation of halogenated phenols may be a significant alternative to biodegradation.
Inhibition of T-2 toxin production on high-moisture corn kernels by modified atmospheres
Authors: Paster N , Menasherov M
Published: January 2020

The fungus Fusarium sporotrichioides, capable of producing T-2 toxin (T-2), was grown on irradiated corn kernels remoistened to 22% and kept in atmospheres of different CO2-O2 combinations. The production of T-2 was totally inhibited under 60% CO2-20% O2, whereas only trace amounts were detected when the gas combination was 40% CO2-5% O2. Under all other combinations tested, the amount of T-2 produced was reduced by 25 to 50% as compared with the control. Fungal growth was not inhibited by any of the gas mixtures examined, and the growth rate (measured by direct plating, dilution method, and CO2 production) was almost identical to that in grains kept under air. It is concluded that although F. sporotrichioides is tolerant to high CO2 levels, T-2 formation on corn can be inhibited by CO2 concentrations less than that required to inhibit fungal growth.
Aerobic biomineralization of alpha-hexachlorocyclohexane in contaminated soil
Authors: Bachmann A , de Bruin W , Jumelet J C , Rijnaarts H H , Zehnder A J
Published: January 2020

The factors identified to be important for the aerobic biodegradation of alpha-hexachlorocyclohexane (alpha-HCH) in a soil slurry are temperature, auxiliary carbon source, substrate concentration, and soil inhomogeneities. Temperatures in the range of 20 to 30 degrees C were determined to be most favorable for biodegradation of alpha-HCH. No alpha-HCH biodegradation was detected at temperatures below 4 degrees C and above 40 degrees C. The addition of auxiliary organic carbon compounds showed repressive effects on alpha-HCH biomineralization. Increased oxygen partial pressures reduced the repressive effects of added auxiliary organic carbon compounds. A linear relationship between alpha-HCH concentration and its conversion rate was found in a Lineweaver-Burk plot. Inhomogeneities such as clumping of alpha-HCH significantly affected its biodegradation. Inhomogeneity as an influence on biodegradation has not drawn sufficient attention in the past, even though it certainly has affected both laboratory studies and the application of biotechnological methods to clean up contaminated sites. On the basis of metabolites detected during degradation experiments, the initial steps of aerobic alpha-HCH bioconversion in a soil slurry are proposed.
A quantitative study of enterotoxin production by sheep milk staphylococci
Authors: Bautista L , Gaya P , Medina M , Nuñez M
Published: January 2020

Of 124 staphylococcal strains isolated from sheep milk, 78 produced enterotoxin A, B, C, or D when evaluated by an enzyme-linked immunosorbent assay. Enterotoxins A and D, elaborated by 44 and 43 strains, respectively, showed the highest incidence. Enterotoxin production by coagulase-negative strains (one Staphylococcus cohnii, three S. epidermidis, five S. haemolyticus, and four S. xylosus) was detected. Linear and logarithmic-logarithmic regressions of optical density on enterotoxin concentration yielded the best-fitting equations for enterotoxin quantitation. A significantly higher incidence of enterotoxin producers and significantly higher levels of enterotoxins produced were recorded for coagulase-positive, thermostable nuclease-positive, hemolysis-positive, or mannitol-positive strains. Mannitol utilization was the best test for discriminating between enterotoxigenic and nonenterotoxigenic staphylococci.
A Positive Strain Identification Method for
Rhizobium meliloti
Authors: Wheatcroft Roger , Watson Robert J.
Published: January 2020

About 80% of
Rhizobium meliloti
strains contain 1 to 11 copies of insertion sequence ISR
m1
in their genomes (R. Wheatcroft and R. J. Watson, J. Gen. Microbiol. 134:113-121, 1988). Hybridization to separated genomic DNA fragments with an ISRm1-specific probe produces patterns of hybridization bands which are distinctive for each strain. These patterns can be compared between strains to prove or disprove common identity. In most cases relatedness can be inferred despite phenotypic differences or minor genomic alterations.
Preparation of a Cell-Free, Isobutene-Forming System from
Rhodotorula minuta
Authors: Fujii Takao , Ogawa Takahira , Fukuda Hideo
Published: January 2020

The cell-free isobutene-forming system of
Rhodotorula minuta
IFO 1102 was prepared. This system consisted of isovaleric acid, NADPH, and a cell extract. The maximal activity obtained by using the described cell-free system was 17 nl/mg of protein per h. It appears that this cell-free isobutene-forming system operates in living cells of
R. minuta.
Biosynthesis of Territrems by
Aspergillus terreus
Authors: Ling Kuo Huang , Peng Yun Wen
Published: January 2020

Different radioactive precursors were added to 8-day potato-dextrose liquid cultures of
Aspergillus terreus
23-1. Territrems were isolated from chloroform extracts of the cultures at day 14 and purified by thin-layer chromatography and high-pressure liquid chromatography. The territrem B obtained was treated with alkaline hydrogen peroxide, and 3, 4, 5-trimethoxy benzoic acid was isolated from an ethyl acetate extract of the reaction mixture and purified by thin-layer chromatography and high-pressure liquid chromatography. By comparison of the specific radioactivities of territrem B and its cleaved aromatic product (disintegrations per minute per micromole of compound), it was demonstrated that the radioactivity of territrem B was located mainly on its aromatic moiety when [U-
14
C]shikimate,
l
-[
methyl
-
14
C]methionine, and
l
-[
methyl
-
3
H]methionine were precursors; however, the radioactivity of territrem B was located mainly on its nonaromatic moiety when [2-
14
C]mevalonate was the precursor. Mevinolin, a specific inhibitor of β-hydroxyl β-methyl glutaryl coenzyme A reductase, was shown to inhibit production of territrems by
A. terreus
23-1. When [U-
14
C]acetate was used as a precursor, mevinolin inhibited the incorporation of radioactive carbon into territrem but mevinolin did not inhibit incorporation of radioactive carbon from [2-
14
C]mevalonate into territrem.
Novel biotransformations of 4-chlorobiphenyl by a Pseudomonas sp
Authors: Barton M R , Crawford R L
Published: January 2020

A bacterium, tentatively identified as a representative of the genus Pseudomonas (strain MB86), was isolated from soil contaminated by wood-preservation chemicals by using 4-chlorobenzoate as an enrichment substrate. The pseudomonad was able to grow on 4-chlorobenzoic acid and 4-chlorobiphenyl as sole carbon and energy sources. Spent culture medium from 4-chlorobiphenyl-grown cells contained 4-chlorobenzoic acid, 4'-chloroacetophenone, 2-hydroxy,2-[4'-chlorophenyl] ethane, and 2-oxo,2-[4'-chlorophenyl] ethanol as metabolites. 4'-Chloroacetophenone was produced in large amounts, possibly as a dead-end metabolite.
Transfer and expression of mesophilic plasmid-mediated degradative capacity in a psychrotrophic bacterium
Authors: Kolenc R J , Inniss W E , Glick B R , Robinson C W , Mayfield C I
Published: January 2020

A psychrotrophic bacterium, originally isolated from a natural aquatic environment, was characterized and identified as Pseudomonas putida Q5 for use as a representative recipient for biodegradative genes from a mesophilic microorganism. The TOL plasmid pWWO of the mesophile P. putida PaW1 was successfully transferred by conjugation to the naturally isolated psychrotroph P. putida Q5, as shown by plasmid analysis by agarose gel electrophoresis. Expression of the genes encoded by the mesophilic TOL plasmid in the psychrotroph was shown by the fact that the transconjugant (designated P. putida Q5T) had the capacity to degrade and utilize toluate (1,000 mg/liter) as a sole source of carbon at temperatures as low as 0 degrees C. Comparison of growth rates over a wide temperature range (0 to 30 degrees C) indicated that the physiological activity of the transconjugant was not reduced and that the plasmid DNA from the mesophile and its encoded enzymes functioned effectively in the psychrotroph at temperatures well below those at which the mesophile could grow. The production and demonstrated functioning of P. putida Q5T illustrates the possibility of developing specific degradative capacities in bacteria which can readily function at low temperatures in chemically contaminated environments or in industrial wastewater treatment systems.
Direct Characterization of Methanogens in Two High-Rate Anaerobic Biological Reactors
Authors: Kobayashi Hester A. , de Macario Everly Conway , Williams Regan S. , Macario Alberto J. L.
Published: January 2020

The methanogenic flora from two types of turbulent, high-rate reactors was studied by immunologic methods as well as by phase-contrast, fluorescence, and scanning electron microscopy. The reactors were a fluidized sand-bed biofilm ANITRON reactor and an ultrafiltration membrane-associated suspended growth MARS reactor (both trademarks of Air Products and Chemicals, Inc., Allentown, Pa.). Conventional microscopic methods revealed complex mixtures of microbes of a range of sizes and shapes, among which morphotypes resembling
Methanothrix
spp. and
Methanosarcina
spp. were noticed. Precise identification of these and other methanogens was accomplished by antigenic fingerprinting with a comprehensive panel of calibrated antibody probes of predefined specificity spectra. The methanogens identified showed morphotypes and antigenic fingerprints indicating their close similarity with the following reference organisms:
Methanobacterium formicicum
MF and
Methanosarcina barkeri
W in the ANITRON reactor only;
Methanosarcina barkeri
R1M3,
M. mazei
S6,
Methanogenium cariaci
JR1, and
Methanobrevibacter arboriphilus
AZ in the MARS reactor only; and
Methanobrevibacter smithii
ALI and
Methanothrix soehngenii
Opfikon in both reactors. Species diversity and distribution appeared to be, at least in part, dependent on the degree of turbulence inside the reactor.
Incidence and characterization of Bacillus cereus isolates contaminating dairy products
Authors: Wong H C , Chang M H , Fan J Y
Published: January 2020

A total of 293 dairy products purchased from local markets were examined to determine the incidence of and characterize Bacillus cereus. Isolations were made on mannitol-egg yolk-polymyxin B agar medium and confirmed by several staining and biochemical tests. B. cereus occurred in 17% of fermented milks, 52% of ice creams, 35% of soft ice creams, 2% of pasteurized milks and pasteurized fruit- or nut-flavored reconstituted milks, and 29% of milk powders, mostly in fruit- or nut-flavored milk mixes. The average population of B. cereus in these dairy products was 15 to 280 CFU/ml or CFU/g (range, 5 to 800). The characteristics of these B. cereus isolates in terms of heat resistance, biochemical reactions, and antibiotic susceptibility were similar to previously reported data except for a higher utilization of sucrose. Some isolates were especially resistant to carbenicillin, nalidixic acid, streptomycin, and tetracycline. The MICs for the isolates were also determined. All of the tested isolates lysed rabbit erythrocytes; 98% showed verotoxicity, 68% showed cytotonic toxicity for CHO cells, and 3 of 11 selected isolates that showed strong hemolysin activity killed adult mice.
Analysis of drug resistance in the archaebacterium Methanococcus voltae with respect to potential use in genetic engineering
Authors: Possot O , Gernhardt P , Klein A , Sibold L
Published: January 2020

The sensitivity of the methanogenic archaebacterium Methanococcus voltae to 12 inhibitors was tested in liquid medium. Four compounds appeared to be inhibitors of growth. Their MICs were as follows: pseudomonic acid, 0.1 micrograms/ml (0.19 microM); puromycin, 2 micrograms/ml (3.6 microM); methionine sulfoximine, 30 micrograms/ml (170 microM); and fusidic acid, 100 micrograms/ml (170 microM). On solid medium, the MICs were similar and the frequency of spontaneous resistance was found to be 5 X 10(-5) (methionine sulfoximine), 10(-7) (pseudomonic acid), and less than 10(-7) (puromycin and fusidic acid). Pseudomonic acid was found to inhibit isoleucyl-tRNA synthetase activity as measured by the in vitro aminoacylation of M. voltae tRNA with L-[U-14C]isoleucine. Fusidic acid and puromycin were shown to inhibit poly(U)-dependent polyphenylalanine synthesis in S30 extracts. Acetylpuromycin was inhibitory at much higher concentrations both in vivo and in vitro for M. voltae. Thus, the pac gene of Streptomyces alboniger, which is responsible for acetylation of puromycin and which conferred resistance to puromycin when introduced in eubacteria and eucaryotes, is a potential selective marker in gene transfer experiments with M. voltae. The latter was recently shown to be transformable. The same would be true for the cat gene of Tn9, which encodes resistance to fusidic acid in eubacteria in addition to resistance to chloramphenicol.
Development of improved defined media for Clostridium botulinum serotypes A, B, and E
Authors: Whitmer M E , Johnson E A
Published: January 2020

The minimal nutritional growth requirements were determined for strains Okra B and Iwanai E, which are representatives of groups I and II, respectively, of Clostridium botulinum. These type B and E strains differed considerably in their nutrient requirements. The organic growth factors required in high concentrations by the Okra B strain (group I) were arginine and phenylalanine. Low concentrations (less than or equal to 0.1 g/liter) of eight amino acids (methionine, leucine, valine, isoleucine, glycine, histidine, tryptophan, and tyrosine) and of five vitamins (pyridoxamine, p-aminobenzoic acid, biotin, nicotinic acid, and thiamine) were also essential for biosynthesis. The 10 required amino acids could be replaced by intact protein of known composition by virtue of the bacterium's ability to synthesize proteases. Glucose or other carbohydrates were not essential for Okra B, although they did stimulate growth. Quantitatively, the most essential nutrients for Okra B were arginine and phenylalanine. In contrast, the nonproteolytic strain, Iwanai E (group II), did not require either arginine or phenylalanine. It required glucose or another carbohydrate energy source for growth and did not utilize arginine or intact protein as a substitute source of energy. Iwanai E utilized ammonia as a nitrogen source, although growth was stimulated significantly by organic nitrogenous nutrients, especially glutamate and asparagine. Iwanai E also required biosynthesis levels of seven amino acids (histidine, isoleucine, leucine, tryptophan, tyrosine, valine, and serine), adenine, and six vitamins (biotin, thiamine, pyridoxamine, folic acid, choline, and nicotinamide). Calcium pantothenate also stimulated growth. On the basis of the nutritional requirements, chemically defined minimal media have been constructed for C. botulinum serotypes A, B, E, and F (proteolytic).(ABSTRACT TRUNCATED AT 250 WORDS)
Conjugal Transfer in Lactic Streptococci of Plasmid-Encoded Insensitivity to Prolate- and Small Isometric-Headed Bacteriophages
Authors: Jarvis Audrey W.
Published: January 2020

Eight of 40 strains of
Streptococcus lactis
and
S. lactis
subsp.
diacetylactis
were able to conjugally transfer a degree of phage insensitivity to
Streptococcus lactis
LM0230. Transconjugants from one donor strain,
S. lactis
subsp.
diacetylactis
4942, contained a 106-kilobase (kb) cointegrate plasmid, pAJ1106. The plasmid was conjugative (Tra
+
) and conferred phage insensitivity (Hsp) and lactose-fermenting ability (Lac) in
S. lactis
and
Streptococcus cremoris
transconjugants. The phage resistance mechanism was effective against prolate- and small isometric-headed phages at 30°C. In
S. lactis
transconjugants, the phage resistance mechanism was considerably weakened at elevated temperatures. A series of deletion plasmids was isolated from transconjugants in
S. cremoris
4854. Deletion plasmids were pAJ2074 (74 kb), Lac
+
, Hsp
+
, Tra
+
; pAJ3060 (60 kb), Lac
+
, Hsp
+
; and pAJ4013 (13 kb), Lac
+
. These plasmids should facilitate mapping Hsp and
tra
genes, with the aim of constructing phage-insensitive strains useful to the dairy industry.
Dual-Label Radioisotope Method for Simultaneously Measuring Bacterial Production and Metabolism in Natural Waters
Authors: Jonas Robert B. , Tuttle Jon H. , Stoner Daphne L. , Ducklow Hugh W.
Published: January 2020

Bacterial production and amino acid metabolism in aquatic systems can be estimated by simultaneous incubation of water samples with both tritiated
methyl
-thymidine and
14
C-labeled amino acids. This dual-label method not only saves time, labor, and materials, but also allows determination of these two parameters in the same microbial subcommunity. Both organic carbon incorporation and respiration can be estimated. The results obtained with the dual-label technique are not significantly different from single-radiolabel methods over a wide range of bacterial activity. The method is particularly suitable for large-scale field programs and has been used successfully with eutrophic estuarine samples as well as with oligotrophic oceanic water. In the mesohaline portion of Chesapeake Bay, thymidine incorporation ranged seasonally from 2 to 635 pmol liter
−1
h
−1
and amino acid turnover rates ranged from 0.01 to 28.4% h
−1
. Comparison of thymidine incorporation with amino acid turnover measurements made at a deep, midbay station in 1985 suggested a close coupling between bacterial production and amino acid metabolism during most of the year. However, production-specific amino acid turnover rates increased dramatically in deep bay waters during the spring phytoplankton bloom, indicating transient decoupling of bacterial production from metabolism. Ecological features such as this are readily detectable with the dual-label method.
Lactic acid concentration as an indicator of acceptability in refrigerated or freeze-thawed ground beef
Authors: Nassos P S , King A D , Stafford A E
Published: January 2020

Lactic acid concentrations increased in refrigerated and freeze-thawed anaerobically stored ground beef. Bacterial counts were higher in refrigerated samples, but the ratios of gram-positive bacteria in refrigerated and freeze-thawed samples were the same. No differences in appearance or odor between refrigerated and freeze-thawed samples were noted after 2 days of aerobic storage. Initial lactic acid concentration can be used to predict the shelf life of frozen beef.
Effect of Iron Limitation on “
Pseudomonas plantarii
” Growth and Tropolone and Protein Production
Authors: Azegami Koji , Nishiyama Koushi , Kato Hajime
Published: January 2020

The addition of iron to an iron-deficient medium markedly enhanced the growth of “
Pseudomonas plantarii
” and the production of red crystals. However, it markedly reduced the amount of dissolved tropolone, a product of the bacterium and an iron chelator, and the production of an iron-regulated protein (78 kilodaltons). The red crystal was complex, composed of tropolone and iron, the ratio being 3:1.
Anaerobic Microbial Dissolution of Transition and Heavy Metal Oxides
Authors: Francis A. J. , Dodge C. J.
Published: January 2020

Anaerobic microbial dissolution of several crystalline, water-insoluble forms of metal oxides commonly associated with the waste from energy production was investigated. An anaerobic N-fixing
Clostridium
sp. with an acetic, butyric, and lactic acid fermentation pattern, isolated from coal-cleaning waste, solubilized Fe
2
O
3
and MnO
2
by direct enzymatic reduction; CdO, CuO, PbO, and ZnO were solubilized by indirect action due to the production of metabolites and the lowering of the pH of the growth medium. Extracellular heat-labile components of the cell-free spent medium obtained from cultures without oxide solubilized a significant amount of Fe
2
O
3
(1.7 μmol); however, direct contact with the bacterial cells resulted in the complete dissolution (4.8 μmol) of the oxide. Under identical conditions, the cell-free spent medium solubilized only a small amount of MnO
2
(0.07 μmol), whereas 2.3 μmol of the oxide was solubilized by direct bacterial contact. Reduction of Fe
2
O
3
and MnO
2
by
Clostridium
sp. proceeds at different rates and, possibly, by different enzymatic systems. Fe(III) and Mn(IV) oxides appear to be used as sinks for excess electrons generated from glucose fermentation, since there is no apparent increase in growth of the bacterium concomitant with the reduction of the oxides. Dialysis bag experiments with Co
2
O
3
indicate that there is a slight dissolution of Co (0.16 μmol) followed by precipitation or biosorption. Although Mn
2
O
3
, Ni
2
O
3
, and PbO
2
may undergo reductive dissolution from a higher to a lower oxidation state, dissolution by direct or indirect action was not observed. Also, Cr
2
O
3
and NiO were not solubilized by direct or indirect action. Significant amounts of solubilized Cd, Cu, and Pb were immobilized by the bacterial biomass, and the addition of Cu
2+
inhibited the growth of the bacterium.
Biotransformation of Dehydroabietic, Abietic, and Isopimaric Acids by
Mortierella isabellina
Immobilized in Polyurethane Foam
Authors: Kutney James P. , Berset Jean-Daniel , Hewitt Gary M. , Singh Mahatam
Published: January 2020

Polyurethane foam supports immobilization of
Mortierella isabellina
ATCC 38063, a zygomycete that hydroxylates and thereby detoxifies resin acids found in certain pulp mill effluents. The fungus becomes entrapped as it threads growing mycelia through the open foam matrix. The tenacity of binding and the amount of biomass bound depend on initial spore numbers and nutrient concentration. Optimal dehydroabietic acid transformation occurs with early-stationary-phase foam-bound mycelia suspended in buffer at pH 6.5 to 8.5 with aeration >0.1 liter liter
−1
min
−1
and near a temperature maximum of 33°C. Neither a greater number of similarly loaded foam cubes nor a greater surface area for similar amounts of biomass increased transformation rates. Compared to free mycelial cultures, foam presence retards growth and decreases the rate of precursor transformation but does not alter the nature of the metabolites formed from dehydroabietic acid, abietic acid, or isopimaric acid. Diffusional barriers or binding of substrates in an inaccessible manner may cause the observed inhibitions. Overall, foam immobilization stabilizes enzymic activity and is a facile laboratory process with scaleup potential.
Molecular Cloning and Expression of a Xylanase Gene from
Bacillus polymyxa
in
Escherichia coli
Authors: Yang Robert C. A. , MacKenzie C. Roger , Bilous Doris , Seligy Verner L. , Narang Saran A.
Published: January 2020

Genomic fragments of
Bacillus polymyxa
derived from separate and complete digestion by
Eco
RI,
Hin
dIII, and
Bam
HI were ligated into the corresponding sites of pBR322, and the resulting chimeric plasmids were transformed into
Escherichia coli.
Of 6,000 transformants screened, 1 (pBPX-277) produced a clear halo on Remazol brilliant blue xylan plates. The insert in the pBPX-277 recombinant, identified as an 8.0-kilobase
Bam
HI fragment of
B. polymyxa
, was subsequently subjected to extensive mapping and a series of subclonings into pUC19. A 2.9-kilobase
Bam
HI-
Eco
RI subfragment was found to code for xylanase activity. Xylanase activity expressed by
E. coli
harboring the cloned gene was located primarily in the periplasm and corresponded to one of two distinct xylanases produced by
B. polymyxa.
Xylanase expression by the cloned gene occurred in the absence of xylan and was reduced by glucose and xylose. Southern blot hybridization with the cloned fragment as a probe against complete genomic digests of the bacilli
B. polymyxa, B. circulans
, and
B. subtilis
revealed that the cloned xylanase gene was unique to
B. polymyxa.
The xylanase expressed by the cloned gene had a molecular weight of approximately 48,000 and an isoelectric point of 4.9.
Circular Permutation of the Genome of a Temperate Bacteriophage from
Streptococcus cremoris
BK5
Authors: Lakshmidevi G. , Davidson Barrie E. , Hillier Alan J.
Published: January 2020

The temperate bacteriophage BK5-T was isolated from
Streptococcus cremoris
BK5 by induction with mitomycin C. Electron microscopy revealed that BK5-T DNA consists of linear molecules, ranging in size from 39.7 to 46 kilobase pairs. Restriction analysis of self-ligated BK5-T DNA showed that the ends of the DNA were not cohesive. The
Eco
RI restriction fragments of the phage genome were cloned into pACYC184. Restriction enzyme analysis of both the phage DNA and the cloned
Eco
RI fragments with
Eco
RI,
Bst
EII,
Pst
I,
Cla
I, and
Xba
I yielded a 37.6-kilobase-pair-long circular restriction map for the phage genome. It was concluded that the BK5-T DNA molecules in the population differ in their sequence by a circular permutation and that individual DNA molecules are terminally redundant. The map location of the sites at which packaging of BK5-T DNA into phage heads is initiated (
pac
) and at which the phage integrates into the bacterial chromosome (
att
) were established.
Isolation of Bacteria Which Can Grow at Both High pH and Low Temperature
Authors: Kimura Takuhei , Horikoshi Koki
Published: January 2020

Fifty bacterial strains able to grow at pH 10 and 0°C were isolated from soils, and growth characteristics of three selected strains were investigated. Strain 207, which showed the best growth rate of all the isolates at the conditions described above, could grow at a temperature of −5 to 39°C at pH 8.5. The optimum pH for this strain changed from 9.5 at 10°C to 9.0 at 20°C.
Effect of a Killer Toxin of Yeast on Eucaryotic Systems
Authors: Pfeiffer Peter , Radler Ferdinand , Caspritz Gert , Hänel Heinz
Published: January 2020

The
Saccharomyces cerevisiae
killer toxin KT 28, which inhibits sensitive yeasts, was shown to have no effect on several pathogenic fungi or on the protozoan
Trichomonas vaginalis.
At concentrations of about 0.1 mg/ml, a partial inhibition of the skin pathogenic fungi
Trichophyton rubrum
and
Microsporum canis
was observed at pH 6.5. No pharmacological activity was detected in various tests with several animal organs.
Influence of Storage at Freezing and Subsequent Refrigeration Temperatures on β-Galactosidase Activity of
Lactobacillus acidophilus
Authors: Gilliland S. E. , Lara R. C.
Published: January 2020

The ability of three strains of
Lactobacillus acidophilus
to survive and retain β-galactosidase activity during storage in liquid nitrogen at −196°C and during subsequent storage in milk at 5°C was tested. The level of β-galactosidase activity varied among the three strains (0.048 to 0.177 U/10
7
organisms). Freezing and storage at −196°C had much less adverse influence on viability and activity of the enzyme than did storage in milk at 5°C. The strains varied in the extent of the losses of viability and β-galactosidase activity during both types of storage. There was not a significant interaction between storage at −196°C and subsequent storage at 5°C. The strains that exhibited the greatest losses of β-galactosidase activity during storage in milk at 5°C also exhibited the greatest losses in viability at 5°C. However, the losses in viability were of much greater magnitude than were the losses of enzymatic activity. This indicates that some cells of
L. acidophilus
which failed to form colonies on the enumeration medium still possessed β-galactosidase activity. Cultures of
L. acidophilus
to be used as dietary adjuncts to improve lactose utilization in humans should be carefully selected to ensure that adequate β-galactosidase activity is provided.
A model for analyzing growth kinetics of a slowly growing Mycobacterium sp
Authors: Lambrecht R S , Carriere J F , Collins M T
Published: January 2020

This report describes a simple method for quantifying viable mycobacteria and for determining generation time. We used statistical models and computer analysis of growth curves generated for the slowly growing mycobacterium Mycobacterium paratuberculosis under controlled conditions to derive a mathematical formula relating the dependent variable, growth, to the independent variables, log10 number of organisms in the inoculum (inoculum size) and incubation time. Growth was measured by a radiometric method which detects 14CO2 release during metabolism of a 14C-labeled substrate. The radiometric method allowed for early detection of growth and detected as few as three viable bacteria. The coefficient of variation between culture vials inoculated with the same number of M. paratuberculosis was 0.083. Radiometric measurements were highly correlated to spectrophotometric and plate count methods for measuring growth (r = 0.962 and 0.992, respectively). The proportion of the total variability explained by the model in a goodness of fit test was 0.9994. Application of the model to broth cultures provided accurate estimates of the number of M. paratuberculosis (standard error = 0.21, log10 scale) and the growth rate (coefficient of variation, 0.03). Generation time was observed to be dependent upon the number of organisms in the inoculum. The model accurately described all phases of growth of M. paratuberculosis and can likely be applied to other slowly growing microorganisms.
Two-Stage Mineralization of Phenanthrene by Estuarine Enrichment Cultures
Authors: Guerin William F. , Jones Galen E.
Published: January 2020

The polycyclic aromatic hydrocarbon phenanthrene was mineralized in two stages by soil, estuarine water, and sediment microbial populations. At high concentrations, phenanthrene was degraded, with the concomitant production of biomass and accumulation of Folin-Ciocalteau-reactive aromatic intermediates. Subsequent consumption of these intermediates resulted in a secondary increase in biomass. Analysis of intermediates by high-performance liquid chromatography, thin-layer chromatography, and UV absorption spectrometry showed 1-hydroxy-2-naphthoic acid (1H2NA) to be the predominant product. A less pronounced two-stage mineralization pattern was also observed by monitoring
14
CO
2
production from low concentrations (0.5 mg liter
−1
) of radiolabeled phenanthrene. Here, mineralization of
14
C-labeled 1H2NA could explain the incremental
14
CO
2
produced during the later part of the incubations. Accumulation of 1H2NA by isolates obtained from enrichments was dependent on the initial phenanthrene concentration. The production of metabolites during polycyclic aromatic hydrocarbon biodegradation is discussed with regard to its possible adaptive significance and its methodological implications.
Trichloroethylene Biodegradation by a Methane-Oxidizing Bacterium
Authors: Little C. Deane , Palumbo Anthony V. , Herbes Stephen E. , Lidstrom Mary E. , Tyndall Richard L. , Gilmer Penny J.
Published: January 2020

Trichloroethylene (TCE), a common groundwater contaminant, is a suspected carcinogen that is highly resistant to aerobic biodegradation. An aerobic, methane-oxidizing bacterium was isolated that degrades TCE in pure culture at concentrations commonly observed in contaminated groundwater. Strain 46-1, a type I methanotrophic bacterium, degraded TCE if grown on methane or methanol, producing CO
2
and water-soluble products. Gas chromatography and
14
C radiotracer techniques were used to determine the rate, methane dependence, and mechanism of TCE biodegradation. TCE biodegradation by strain 46-1 appears to be a cometabolic process that occurs when the organism is actively metabolizing a suitable growth substrate such as methane or methanol. It is proposed that TCE biodegradation by methanotrophs occurs by formation of TCE epoxide, which breaks down spontaneously in water to form dichloroacetic and glyoxylic acids and one-carbon products.
High-efficiency, one-step starch utilization by transformed Saccharomyces cells which secrete both yeast glucoamylase and mouse alpha-amylase
Authors: Kim K , Park C S , Mattoon J R
Published: January 2020

Transformed, hybrid Saccharomyces strains capable of simultaneous secretion of glucoamylase and alpha-amylase have been produced. These strains could carry out direct, one-step assimilation of starch, with conversion efficiency greater than 93% during a 5-day growth period. One of the transformants converted 92.8% of available starch into reducing sugars in only 2 days. Glucoamylase secretion by these strains resulted from expression of one or more chromosomal STA genes derived from Saccharomyces diastaticus. The strains were transformed by a plasmid (pMS12) containing mouse salivary alpha-amylase cDNA in an expression vector containing yeast alcohol dehydrogenase promoter and a segment of yeast 2 micron plasmid. The major starch hydrolysis product produced by crude amylases found in culture broths was glucose, indicating that alpha-amylase and glucoamylase acted cooperatively.
Phenotypic and genotypic comparison of Escherichia coli from pristine tropical waters
Authors: Bermúdez M , Hazen T C
Published: January 2020

Nine fecal-coliform-positive strains were isolated from pristine sites in a tropical rain forest. These sites included nonpolluted rivers and water from bromeliads (epiphytes) which were 30 ft (ca. 910 cm) above the ground. Phenotypically, all of these isolates were identified as Escherichia coli. Their DNA was isolated and purified, and the base composition (G + C content) was determined and compared with that of E. coli B (ATCC 11303). The DNA from the environmental isolates was also hybridized to radiolabeled DNA from E. coli B. Eight strains had a DNA base composition similar to that of E. coli B and gave more than 75% homology with E. coli B. One strain had a different DNA base composition and a relatively low percentage of homology with the reference strain. The finding of E. coli in pristine tropical waters suggests that this bacterium could be a natural inhabitant in these environments and is not a reliable indicator of recent human fecal contamination in tropical waters. The indicators that are currently used in the tropics to test the biological quality of water should be reevaluated.
Strain Improvement of
Rhodotorula graminis
for Production of a Novel
l
-Phenylalanine Ammonia-Lyase
Authors: Orndorff Steve A. , Costantino Nina , Stewart David , Durham Don R.
Published: January 2020

l
-Phenylalanine ammonia-lyase (PAL; EC 4.3.1.5) from
Rhodotorula rubra
has been used in the commercial manufacture of
l
-phenylalanine from
trans
-cinnamic acid. In this study,
R. graminis
PAL was investigated. Mutant strain GX6000 was isolated after ethyl methanesulfonate mutagenesis of wild-type
R. graminis
GX5007 by selecting for resistance to phenylpropiolic acid, an analog of
trans
-cinnamic acid. Mutant strain GX6000 produced inducible PAL at levels four- to fivefold higher than had wild-type
R. graminis.
Furthermore, this strain had several other physiological traits that make it more commercially useful than
R. rubra.
For example, during fermentation, the PAL half-life was three- to fivefold longer, PAL specific activity was six to seven times higher, and PAL synthesis was significantly less inhibited by temperatures above 30°C. Induction of PAL in strain GX6000 appeared to be less tightly regulated;
l
-leucine acted synergistically with
l
-phenylalanine, the physiological inducer, to increase the PAL specific activity and titer to 165 U/g (dry weight) and 3,000 U/liter, respectively, a 40% increase over the effect of
l
-phenylalanine alone. Strain GX6000 PAL showed significantly greater stability in bioreactors for the synthesis of
l
-phenylalanine, a finding that is consistent with the stability properties observed during fermentation.
Denitrification in a Sand and Gravel Aquifer
Authors: Smith Richard L. , Duff John H.
Published: January 2020

Denitrification was assayed by the acetylene blockage technique in slurried core material obtained from a freshwater sand and gravel aquifer. The aquifer, which has been contaminated with treated sewage for more than 50 years, had a contaminant plume greater than 3.5-km long. Near the contaminant source, groundwater nitrate concentrations were greater than 1 mM, whereas 0.25 km downgradient the central portion of the contaminant plume was anoxic and contained no detectable nitrate. Samples were obtained along the longitudinal axis of the plume (0 to 0.25 km) at several depths from four sites. Denitrification was evident at in situ nitrate concentrations at all sites tested; rates ranged from 2.3 to 260 pmol of N
2
O produced (g of wet sediment)
−1
h
−1
. Rates were highest nearest the contaminant source and decreased with increasing distance downgradient. Denitrification was the predominant nitrate-reducing activity; no evidence was found for nitrate reduction to ammonium at any site. Denitrifying activity was carbon limited and not nitrate limited, except when the ambient nitrate level was less than the detection limit, in which case, even when amended with high concentrations of glucose and nitrate, the capacity to denitrify on a short-term basis was lacking. These results demonstrate that denitrification can occur in groundwater systems and, thereby, serve as a mechanism for nitrate removal from groundwater.