bluuebunny/crossref_metadata_2025 huggingface dataset:
Fate of 2,2,2-trichloroacetaldehyde (chloral hydrate) produced during trichloroethylene oxidation by methanotrophs
Authors: Newman L M , Wackett L P
Published: January 2020

Four different methanotrophs expressing soluble methane monooxygenase produced 2,2,2-trichloroacetaldehyde, or chloral hydrate, a controlled substance, during the oxidation of trichloroethylene. Chloral hydrate concentrations decreased in these cultures between 1 h and 24 h of incubation. Chloral hydrate was shown to be biologically transformed to trichloroethanol and trichloroacetic acid by Methylosinus trichosporium OB3b. At elevated pH and temperature, chloral hydrate readily decomposed and chloroform and formic acid were detected as products.
Amine Oxidases of Marine Phytoplankton
Authors: Palenik Brian , Morel François M. M.
Published: January 2020

Some phytoplankton utilized a novel mechanism for obtaining nitrogen from primary amines. They oxidized the primary amines to produce extracellular hydrogen peroxide and aldehydes and used the third reaction product, ammonium, as a nitrogen source. The specificity, regulation, inhibition by bromoethylamine, and potential dependence on copper of this process are described.
Effect of sodium chloride on transport of bacteria in a saturated aquifer material
Authors: Gannon J , Tan Y H , Baveye P , Alexander M
Published: January 2020

Determinations were made of the influence of NaCl concentration, cell density, and flow velocity on the transport of Pseudomonas sp. strain KL2 through columns of aquifer sand under saturated conditions. A pulse-type boundary condition was used. The experiments were conducted by using 0.3-m-long Plexiglas columns with an internal diameter of 0.05 m. When a 1-h pulse of a 0.01 M NaCl solution containing 10(8) cells per ml was added at a flow rate of 10(-4) m s-1, the bacterial density in the effluent never exceeded 2.2% of the density of cells added, and only 1.5% of the bacteria passed through the aquifer material. In contrast, when the bacteria were applied in distilled water, the relative cell density in the effluent approached 100%, and 60% of the bacteria were transported through the aquifer solids. Under these conditions, the breakthrough of Pseudomonas sp. strain KL2 was slower than chloride. When the flow rate was 2.0 x 10(-4) m s-1, the cell density in the effluent reached 7.3% of that added in 0.01 M NaCl solution, but only 3.9% of the bacteria were transported through the aquifer particles. On the other hand, the density in the effluent approached 100% of that added in deionized water, and 77% of the added bacteria were recovered. When the density of added cells was 10(9) cells per ml at a flow rate of 10(-4) m s-1, the densities in the effluent reached 70 and 100% of those added in salt solution and deionized water, respectively, and 44 and 57% of the bacteria were transported through the aquifer solids.(ABSTRACT TRUNCATED AT 250 WORDS)
Important factors for testing barrier materials with surrogate viruses
Authors: Lytle C D , Truscott W , Budacz A P , Venegas L , Routson L B , Cyr W H
Published: January 2020

This study evaluated bacteriophages phi X174, T7, PRD1, and phi 6 as possible surrogates for pathogenic human viruses to challenge barrier materials and demonstrated some important factors for their use. Chemical incompatibility with test material was demonstrated when lipid-enveloped phi 6 was inactivated by an aqueous eluate of vinyl gloves, but 0.5% calf serum protected phi 6 from the eluate. Low concentrations (2%) of calf serum also prevented the exaggerated binding of the bacteriophages to filters. Recovery of viruses from surfaces decreased with increasing time before recovery. Penetration through punctures displayed different types of kinetics. The combined data indicate that (i) some bacteriophages may serve as surrogate viruses, (ii) experimental conditions determine whether a particular virus is appropriate as a challenge, and (iii) phi X174 is an excellent choice as a surrogate virus to test barrier materials. The data further indicate that before barrier materials are challenged with viruses, adequate tests should be performed to ensure that the virus is compatible with the test material and test conditions, so that meaningful data will result.
Characterization and overexpression of the Lactococcus lactis pepN gene and localization of its product, aminopeptidase N
Authors: van Alen-Boerrigter I J , Baankreis R , de Vos W M
Published: January 2020

The chromosomal pepN gene encoding lysyl-aminopeptidase activity in Lactococcus lactis has been identified in a lambda EMBL3 library in Escherichia coli by using an immunological screening with antiserum against a purified aminopeptidase fraction. The pepN gene was localized and subcloned in E. coli on the basis of its expression and hybridization to a mixed-oligonucleotide probe for the previously determine N-terminal amino acid sequence of lysyl-aminopeptidase (P. S. T. Tan and W. N. Konings, Appl. Environ. Microbiol. 56:526-532, 1990). The L. lactis pepN gene appeared to complement an E. coli strain carrying a mutation in its pepN gene. High-level expression of the pepN gene in E. coli was obtained by using the T7 system. The overproduction of the 95-kDa aminopeptidase N could be visualized on sodium dodecyl sulfate-polyacrylamide gels and immunoblots. Cloning of the pepN gene on a multicopy plasmid in L. lactis resulted in a 20-fold increase in lysyl-aminopeptidase activity that corresponded to several percent of total protein. Nucleotide sequence analysis of the 5' region of the pepN gene allowed a comparison between the deduced and determined amino-terminal primary sequences of aminopeptidase N. The results show that the amino terminus of PepN is not processed and does not possess the characteristics of consensus signal sequences, indicating that aminopeptidase N is probably an intracellular protein. The intracellular location of aminopeptidase N in L. lactis was confirmed by immunogold labeling of lactococcal cells.
Design and testing of a functional group-specific DNA probe for the study of natural populations of acetogenic bacteria
Authors: Lovell C R , Hui Y
Published: January 2020

The acetogens, although phylogenetically diverse, can be characterized by their possession of the acetyl coenzyme A (acetyl-CoA) pathway for autotrophic CO2 fixation. The gene encoding formyltetrahydrofolate synthetase, a key enzyme of the acetyl-CoA pathway, was previously cloned from the thermophilic acetogen Clostridium thermoaceticum and has now been tested as a group-specific probe for acetogens. Stable hybrids were formed between the probe and single DNA fragments from eight known acetogens representing six genera. A hybrid was also formed between the probe and a DNA fragment from one sulfate reducer known to be capable of both autotrophic CO2 fixation and acetate catabolism. No such hybrid was formed between the probe and DNA from a homoacetate fermenter not known to use the acetyl-CoA pathway, with two known formyltetrahydrofolate synthetase-producing purine fermenters, or with DNA from 27 other species representing 16 genera of organisms that do not use the acetyl-CoA pathway. DNA purified from cells extracted from horse manure was also screened with the acetogen probe. Six hybrids, indicating at least six detectable acetogen "strains," were observed.
Use of the polymerase chain reaction in detection of culturable and nonculturable Vibrio vulnificus cells
Authors: Brauns L A , Hudson M C , Oliver J D
Published: January 2020

Vibrio vulnificus is a human pathogen associated with consumption of raw oysters. During the colder months the organism apparently enters a viable but nonculturable state and thus cannot be cultured by ordinary bacteriological methods. For this reason, another means of detecting this bacterium is necessary. In the present study we utilized the polymerase chain reaction (PCR) to detect V. vulnificus DNA, thus eliminating the problem of nonculturability. DNA from both culturable and nonculturable cells of V. vulnificus was amplified by PCR with primers flanking a 340-bp fragment of the cytotoxin-hemolysin gene. As little as 72 pg of DNA from culturable cells and 31 ng of DNA from nonculturable cells could be detected. Fifty cycles of a two-step reaction (30 s [each] at 94 and 65 degrees C) were found to be optimal as well as more time efficient than the three-step PCR. The total procedure from the point of DNA extraction to observation on a gel required less than 8 h. Possible reasons for the difficulties encountered in amplifying DNA from nonculturable cells, e.g., gene rearrangement or loss of the hemolysin gene, are discussed.
Phosphatidylinositol-specific phospholipase C activity as a marker to distinguish between pathogenic and nonpathogenic Listeria species
Authors: Notermans S H , Dufrenne J , Leimeister-W�chter M , Domann E , Chakraborty T
Published: January 2020

In this study, 468 Listeria strains were checked for the presence of phosphatidylinositol-specific phospholipase C (PI-PLC) activity by using a simple assay that consisted of overlaying colonies formed on agar plates with L-alpha-phosphatidylinositol as substrate. In this assay, PI-PLC-active colonies show turbid halos around the colonies as a result of the release of insoluble diacylglycerol from the substrate. This activity was detected only in the pathogenic species Listeria monocytogenes and was not present in any of the 167 strains of Listeria seeligeri, Listeria welshimeri, Listeria innocua, Listeria murrayi, and Listeria grayi tested. Hence, screening for PI-PLC activity permits discrimination between pathogenic and nonpathogenic Listeria species. In particular, the hemolytic but nonpathogenic species L. seeligeri can now be separated from the hemolytic and pathogenic species L. monocytogenes and L. ivanovii. The use of this assay will improve the specific detection and/or isolation of pathogenic Listeria species from clinical samples or food enrichment cultures.
Purification and properties of 4-hydroxybutyrate coenzyme A transferase from Clostridium aminobutyricum
Authors: Scherf U , Buckel W
Published: January 2020

A new coenzyme A (CoA)-transferase from the anaerobe Clostridium aminobutyricum catalyzing the formation of 4-hydroxybutyryl-CoA from 4-hydroxybutyrate and acetyl-CoA is described. The enzyme was purified to homogeneity by standard techniques, including fast protein liquid chromatography under aerobic conditions. Its molecular mass was determined to be 110 kDa, and that of the only subunit was determined to be 54 kDa, indicating a homodimeric structure. Besides acetate and acetyl-CoA, the following substrates were detected (in order of decreasing kcat/Km): 4-hydroxybutyryl-CoA, butyryl-CoA and propionyl-CoA, vinyl-acetyl-CoA (3-butenoyl-CoA), and 5-hydroxyvaleryl-CoA. In an indirect assay the corresponding acids were also found to be substrates; however, DL-lactate, DL-2-hydroxybutyrate, DL-3-hydroxybutyrate, crotonate, and various dicarboxylates were not.
Conjugal transfer of Tn916, Tn916 delta E, and pAM beta 1 from Enterococcus faecalis to Butyrivibrio fibrisolvens strains
Authors: Hespell R B , Whitehead T R
Published: January 2020

Anaerobic filter matings of Butyrivibrio fibrisolvens H17c, CF3, D1, or GS113, representing different DNA relatedness groups, were done with Enterococcus faecalis CG110, which contains chromosomally inserted Tn916. Tetracycline-resistant transconjugants were obtained with each mating pair at average frequencies of 4.4 x 10(-6) (per recipient) and 5.2 x 10(-6) (per donor). The transfer frequencies of Tn916 into B. fibrisolvens varied 5- to 10-fold with mating time, strain, and growth stage. By using Southern hybridization with pAM120 as the probe, Tn916 was shown to insert at one or more separate chromosomal sites for each strain of B. fibrisolvens. Retransfer of Tn916 from B. fibrisolvens H17c or CF3 to E. faecalis OG1-X or JH 2-2 or to B. fibrisolvens D1 or GS113 could not be shown. Matings of E. faecalis RH110, which contains chromosomally inserted Tn916 delta E, with B. fibrisolvens 49, H17c, D1, CF3, GS113, or VV-1 resulted in erythromycin-resistant transconjugants at average frequencies of 5.3 x 10(-7) (per recipient) and 2.5 x 10(-7) (per donor). Tn916 delta E was shown by Southern hybridization with pAM120 to insert at one or more sites in the chromosome of each strain. B. fibrisolvens H17c was anaerobically filter mated with E. faecalis JH 2-SS, which contains pAM beta 1. Erythromycin-resistant transconjugants were obtained at frequencies of 2 x 10(-5) (per recipient) and 6 x 10(-5) (per donor). The presence of pAM beta 1 in these transconjugants could not be shown by agarose gel electrophoresis of plasmid minilysates but could be shown by Southern hybridization analysis.(ABSTRACT TRUNCATED AT 250 WORDS)
Abundance of Viruses in Marine Waters: Assessment by Epifluorescence and Transmission Electron Microscopy
Authors: Hara Shigemitsu , Terauchi Kazuki , Koike Isao
Published: January 2020

Abundance of bacteria and tiny DNA-associated particles in the upper layer of Japanese coastal and offshore waters was evaluated by epifluorescence microscopy with 0.015-μm-pore-size Nuclepore filters. The number of tiny DNA-associated particles was compared with the abundance of virus particles estimated by transmission electron microscopy. Although a large variation in virus abundance (1.2 � 10
6
to 35 � 10
6
ml
−1
) was obtained with the transmission electron microscopy method, the ratio of 4′,6-diamidino-2-phenylindole-reactive tiny particles to viruses was in a rather narrow range (1.0 to 1.6), indicating that the majority of the tiny DNA-associated particles identified by epifluorescence microscopy were actually virus particles. This result implies the possibility of using epifluorescence microscopy for the evaluation of virus abundance in marine environments.
Characterization of Vibrio anguillarum and closely related species isolated from farmed fish in Norway
Authors: Myhr E , Larsen J L , Lillehaug A , Gudding R , Heum M , H�stein T
Published: January 2020

A total of 264 bacterial strains tentatively or definitely classified as Vibrio anguillarum were examined. The strains were isolated from diseased or healthy Norwegian fish after routine autopsy. With the exception of five isolates from wild saithe (Pollachius virens), the strains originated from nine different species of farmed fish. The bacteria were subjected to morphological, physiological, and biochemical studies, numerical taxonomical analyses, serotyping by slide agglutination and enzyme-linked immunosorbent assay, DNA-plasmid profiling, and in vitro antimicrobial drug susceptibility testing. The results of the microbiological studies were correlated to anamnestic information. The bacterial strains were identified as V. anguillarum serovar O1 (n = 132), serovar O2 (n = 89), serovar O4 (n = 2), serovar O8 (n = 1), and not typeable (n = 1) as well as Vibrio splendidus biovar I (n = 36) and biovar II (n = 1), Vibrio tubiashii (n = 1), and Vibrio fischerii (n = 1). V. anguillarum serovar O1 or O2 was isolated in 176 out of 179 cases of clinical vibriosis in Atlantic salmon (Salmo salar). V. anguillarum serovar O1 was the only serovar isolated from salmonid fish species other than Atlantic salmon, while V. anguillarum serovar O2 was isolated from all marine fish suffering from vibriosis. A 48-Mda plasmid was isolated from all V. anguillarum serovar O1 isolates examined. Serovar O2 isolates did not harbor any plasmids. Resistance against commonly used antibiotic compounds was not demonstrated among V. anguillarum isolates. Neither V. splendidus biovar I nor other V. anguillarum-related species appeared to be of clinical importance among salmonid fish.(ABSTRACT TRUNCATED AT 250 WORDS)
Resuscitation effects of catalase on airborne bacteria
Authors: Marthi B , Shaffer B T , Lighthart B , Ganio L
Published: January 2020

Catalase incorporation into enumeration media caused a significant increase (greater than 63%) in the colony-forming abilities of airborne bacteria. Incubation for 30 to 60 min of airborne bacteria in collection fluid containing catalase caused a greater than 95% increase in colony-forming ability. However, catalase did not have any effects on enumeration at high relative humidities (80 to 90%).
Effects of Nutrients on Specific Growth Rate of Bacterioplankton in Oligotrophic Lake Water Cultures
Authors: Coveney Michael F. , Wetzel Robert G.
Published: January 2020

The effects of organic and inorganic nutrient additions on the specific growth rates of bacterioplankton in oligotrophic lake water cultures were investigated. Lake water was first passed through 0.8-μm-pore-size filters (prescreening) to remove bacterivores and to minimize confounding effects of algae. Specific growth rates were calculated from changes in both bacterial cell numbers and biovolumes over 36 h. Gross specific growth rates in unmanipulated control samples were estimated through separate measurements of grazing losses by use of penicillin. The addition of mixed organic substrates alone to prescreened water did not significantly increase bacterioplankton specific growth rates. The addition of inorganic phosphorus alone significantly increased one or both specific growth rates in three of four experiments, and one experiment showed a secondary stimulation by organic substrates. The stimulatory effects of phosphorus addition were greatest concurrently with the highest alkaline phosphatase activity in the lake water. Because bacteria have been shown to dominate inorganic phosphorus uptake in other P-deficient systems, the demonstration that phosphorus, rather than organic carbon, can limit bacterioplankton growth suggests direct competition between phytoplankton and bacterioplankton for inorganic phosphorus.
Purification and Partial Characterization of Lacticin 481, a Lanthionine-Containing Bacteriocin Produced by
Lactococcus lactis
subsp.
lactis
CNRZ 481
Authors: Piard J.-C. , Muriana P. M. , Desmazeaud M. J. , Klaenhammer T. R.
Published: January 2020

Lacticin 481, a bacteriocin produced during the growth of
Lactococcus lactis
subsp.
lactis
CNRZ 481, was purified sequentially by ammonium sulfate precipitation, gel filtration, and preparative and analytical reversed-phase high-pressure liquid chromatography. Ammonium sulfate precipitations resulted in a 455-fold increase in total lacticin 481 activity. The entire purification protocol led to a 107, 506-fold increase in the specific activity of lacticin 481. On the basis of its electrophoretic pattern in sodium dodecyl sulfate-polyacrylamide gels, lacticin 481 appeared as a single peptide band of 1.7 kDa. However, dimers of 3.4 kDa also exhibiting lacticin activity were detected. Derivatives of the lacticin-producing strain which did not produce lacticin 481 (Bac
-
) were sensitive to this bacteriocin (Bac
s
) and failed to produce the 1.7-kDa band. Amino acid composition analysis of purified lacticin 481 revealed the presence of lanthionine residues, suggesting that lacticin 481 is a member of the lantibiotic family of antimicrobial peptides. Seven residues (K G G S G V I) were sequenced from the N-terminal portion of lacticin 481, and these did not shown any homology with nisin or other known bacteriocin sequences.
Competitive Ability and Survival in Soil of
Pseudomonas
Strain 679-2, a Dominant, Nonobligate Bacterial Predator of Bacteria
Authors: Casida L. E.
Published: January 2020

A copper-resistant, nonobligate, bacterial predator of bacteria was isolated from soil. It was a
Pseudomonas
species, designated strain 679-2. It attacked most other nonobligate bacterial predators and hence could control their predatory and other activities in nature. It also inhibited various fungi. It attached to prey cells and produced a toxic, copper-related, growth initiation factor like that produced by
Cupriavidus necator
. In addition, it produced a second, novel compound that was both antibacterial and antifungal. Strain 679-2 appeared to have only a very limited natural occurrence. It was found only in the soil from one small area in one field. It was absent on the leaves of the plant species that were examined. Regardless of its rarity, however, it was highly competitive in soil. An inoculum consisting of only a few cells added to soil multiplied rapidly to become a major component of the soil microflora within 24 h. A small amount of glutamic acid could be added along with the cells to stimulate production of the toxic compounds noted above, but this was not necessary. After this multiplication, or when large numbers of cells were added to soil, the numbers decreased only slowly during the next several months. Cell survival also was good on plant leaves. The survival in soil and on plant leaves occurred in both laboratory and field experiments. Other than desiccation, the natural mechanism for controlling the numbers or activities of strain 679-2 in soil is not known. The various characteristics of this bacterium, as noted above, are of particular interest because they indicate a possible use of the cells or inhibitor compounds for controlling organisms in soil or on plant surfaces.
Isolation of 2,4-Diacetylphloroglucinol from a Fluorescent Pseudomonad and Investigation of Physiological Parameters Influencing Its Production
Authors: Shanahan Phil , O'Sullivan Daniel J. , Simpson Paul , Glennon Jeremy D. , O'Gara Fergal
Published: January 2020

Pseudomonas
sp. strain F113 was isolated from the rhizosphere of sugar beets and shown to inhibit a range of plant pathogenic fungi by producing an antibioticlike compound. An antibiotic-negative mutant strain, F113G22, was generated by transposon mutagenesis. This mutant has lost the ability to inhibit both bacterial and fungal microorganisms on high-iron medium. The antibioticlike compound was subsequently identified as 2,4-diacetylphloroglucinol (DAPG), and a high-pressure liquid chromatographic assay was developed for to detect it quantitatively in growth culture media and soil. The growth temperature had a direct bearing on DAPG production by strain F113, with maximum production at 12°C. The iron concentration, pH, and oxygen had no influence on DAPG production by strain F113 under the assay conditions used. However, a low ratio of culture volume to surface area available to the microbe in the growth container was critical for optimum DAPG production. Different types of carbon sources influenced DAPG production by strain F113 to various degrees. For example, sucrose, fructose, and mannitol promoted high yields of DAPG by strain F113, whereas glucose and sorbose resulted in very poor DAPG production.
Anaerobic Production of Extracellular Polysaccharide by
Butyrivibrio fibrisolvens
nyx
Authors: Wachenheim Daniel E. , Patterson John A.
Published: January 2020

Anaerobic production of extracellular polysaccharide (EP) was examined, using a previously uncharacterized, obligately anaerobic rumen isolate,
Butyrivibrio fibrisolvens
nyx, which produced an EP that was rheologically similar to xanthan gum. The main objectives were to determine the nutritional requirements and conditions which promoted EP production by strain nyx. Strain nyx was grown anaerobically in defined and semidefined media. In addition to carbohydrate and nitrogen sources, strain nyx required acetic acid, folic acid, biotin, and pyridoxamine. Strain nyx produced similar amounts of EP at 35 to 40°C. Conditions that improved growth usually improved EP production. Of the carbohydrates tested, glucose supported the fastest growth and most EP production, followed by sucrose, xylose, and lactose. Strain nyx utilized ammonium sulfate, urea, or vitamin-free casein hydrolysate as nitrogen sources for growth and EP production. At 2 and 20 g/liter, respectively, ammonium sulfate and vitamin-free casein hydrolysate supported about the same rates of growth and EP production. EP was not produced in the lag or stationary phases, and EP production was exponential during exponential cell growth. Based on the results of this work, anaerobic EP production with
B. fibrisolvens
nyx could reduce energy costs for industrial EP production compared with the cost of aerated systems. Finally, this work demonstrated that, under appropriate growth conditions, a gastrointestinal tract (ruminal) microorganism produced high levels of EP.
Novel assay for screening rifamycin B-producing mutants
Authors: Vohra R M
Published: January 2020

A simple method that allows easy identification of rifamycin B-producing strains is described. This method involves the use of an enzyme, rifamycin oxidase, which converts inactive rifamycin B to active rifamycin S. In this method, colonies to be tested are grown in pairs. The two colonies are then transferred to two plates seeded with a sensitive strain of Staphylococcus aureus, one plate of which contains the enzyme rifamycin oxidase. All paired colonies which show a larger inhibition zone diameter on the enzyme-containing plate are identified as rifamycin B producers.
Solubilization and Mineralization of Lignin by White Rot Fungi
Authors: Boyle C. David , Kropp Bradley R. , Reid Ian D.
Published: January 2020

The white rot fungi
Lentinula edodes, Phanerochaete chrysosporium, Pleurotus sajor-caju, Flammulina velutipes
, and
Schizophyllum commune
were grown in liquid media containing
14
C-lignin-labelled wood, and the formation of water-soluble
14
C-labelled products and
14
CO
2
, the growth of the fungi, and the activities of extracellular lignin peroxidase, manganese peroxidase, and laccase were measured. Conditions that affect the rate of lignin degradation were imposed, and both long-term (0- to 16-day) and short-term (0- to 72-h) effects on the production of the two types of product and on the activities of the enzymes were monitored. The production of
14
CO
2
-labelled products from the aqueous ones was also investigated. The short-term studies showed that the different conditions had different effects on the production of the two products and on the activities of the enzymes. Nitrogen sources inhibited the production of both products by all species when differences in growth could be discounted. Medium pH and manganese affected lignin degradation by the different species differently. With
P. chrysosporium
, the results were consistent, with lignin peroxidase playing a role in lignin solubilization and manganese peroxidase being important in subsequent CO
2
production.
Controlled Integration into the
Lactococcus
Chromosome of the pCI829-Encoded Abortive Infection Gene from
Lactococcus lactis
subsp.
lactis
UC811
Authors: Casey Judy , Daly Charles , Fitzgerald Gerald F.
Published: January 2020

The phage insensitivity gene of lactococcal plasmid pCI829 which encodes an abortive infection defense mechanism (Abi) was inserted into the
Lactococcus lactis
subsp.
lactis
CH919 chromosome by utilizing the integration plasmid pCI194, which contains 4.2 kb of homology with the conjugative transposon Tn
919.
Chloramphenicol-resistant transformants expressed phage insensitivity to the prolate-headed phage c2 and the small isometric-headed phage 712, and hybridization analysis indicated that transformants contained pCI194 integrated in single copy. The level of phage insensitivity expressed by the transformants was reduced from that observed when the
abi
gene was located on a replicating plasmid, as determined by plaque assay and burst size analysis. Amplification of the integrated structure after growth in increased concentrations of chloramphenicol resulted in an increase in the expression of phage insensitivity. Hybridization analysis revealed that while pCI194 was stably maintained in an integrated state over 100 generations in the absence of selective pressure, the ability to express phage insensitivity was lost. Hybridization analysis also revealed that DNA flanking the
abi
gene contains homology to the CH919 chromosome.
Competition for Ammonium between Nitrifying and Heterotrophic Bacteria in Continuously Percolated Soil Columns
Authors: Verhagen Frank J. M. , Duyts Hendrik , Laanbroek Hendrikus J.
Published: January 2020

Although the absence of nitrate formation in grassland soils rich in organic matter has often been reported, low numbers of nitrifying bacteria are still found in these soils. To obtain more insight into these observations, we studied the competition for limiting amounts of ammonium between the chemolithotrophic ammonium-oxidizing species
Nitrosomonas europaea
and the heterotrophic species
Arthrobacter globiformis
in the presence of
Nitrobacter winogradskyi
with soil columns containing calcareous sandy soil. The soil columns were percolated continuously at a dilution rate of 0.007 h
-1
, based on liquid volumes, with medium containing 5 mM ammonium and different amounts of glucose ranging from 0 to 12 mM.
A. globiformis
was the most competitive organism for limiting amounts of ammonium. The numbers of
N. europaea
and
N. winogradskyi
cells were lower at higher glucose concentrations, and the potential ammonium-oxidizing activities in the uppermost 3 cm of the soil columns were nonexistent when at least 10 mM glucose was present in the reservoir, although 10
7
nitrifying cells per g of dry soil were still present. This result demonstrated that there was no correlation between the numbers of nitrifying bacteria and their activities. The numbers and activities of
N. winogradskyi
cells decreased less than those of
N. europaea
cells in all layers of the soil columns, probably because of heterotrophic growth of the nitrite-oxidizing bacteria on organic substrates excreted by the heterotrophic bacteria or because of nitrate reduction at reduced oxygen concentrations by the nitrite-oxidizing bacteria. Our conclusion was that the nitrifying bacteria were less competitive than the heterotrophic bacteria for ammonium in soil columns but that they survived as viable inactive cells. Inactive nitrifying bacteria may also be found in the rhizosphere of grassland plants, which is rich in organic carbon. They are possibly reactivated during periods of net mineralization.
Evaluation of synergism among Bacillus thuringiensis toxins
Authors: Tabashnik B E
Published: January 2020

A simple test for synergism among toxins is described and applied to previously reported data on independent and joint toxicities of insecticidal proteins from Bacillus thuringiensis. The analysis shows synergism between a 27-kDa (CytA) toxin and 130- or 65-kDa (CryIV) toxins from B. thuringiensis subsp. israelensis against Aedes aegypti larvae. No positive synergism between 130- and 65-kDa toxins or among three CryIA toxins tested against seven species of Lepidoptera occurred. Comparisons with the original interpretations of these data show one case in which synergism occurred but was reported previously as absent and two cases that were not synergistic but were reported previously as suggestive of synergism. These results show that lack of an appropriate test for synergism can produce misleading conclusions. The methods described here can be used to test for synergistic effects of any poisons.
Novel method to extract large amounts of bacteriocins from lactic acid bacteria
Authors: Yang R , Johnson M C , Ray B
Published: January 2020

Antimicrobial peptides, bacteriocins, produced by lactic acid bacteria were adsorbed on the cells of producing strains and other gram-positive bacteria. pH was a crucial factor in determining the degree of adsorption of these peptides onto cell surfaces. In general, between 93 and 100% of the bacteriocin molecules were adsorbed at pHs near 6.0, and the lowest (< or = 5%) adsorption took place at pH 1.5 to 2.0. On the basis of this property, a novel isolation method was developed for bacteriocins from four genera of lactic acid bacteria. By using this method we made preparations of pediocin AcH, nisin, sakacin A, and leuconocin Lcm1 that were potent and concentrated. This method produced a higher yield than isolation procedures, which rely on precipitation of the bacteriocins from the cell-free culture liquor. It is simple and can be used to produce large quantities of bacteriocins from lactic acid bacteria to be used as food biopreservatives.
Bacterioplankton community structure and dynamics after large-scale release of nonindigenous bacteria as revealed by low-molecular-weight-RNA analysis
Authors: Höfle M G
Published: January 2020

A set of freshwater mesocosms (1.7 m3 each) was inoculated with large amounts of Escherichia coli, Pseudomonas putida, and their culture medium to substantially disturb the natural microbial community. To monitor microbial community dynamics, low-molecular-weight RNA (5S rRNA and tRNA) obtained directly from bacterioplankton was analyzed by using high-resolution electrophoresis. The introduced bacteria showed no significant effect on the community structure of the natural bacterial assemblage and its dynamics for 16 days. In contrast, the addition of culture medium resulted within 2 days in a reduction of community diversity due to dominance of a single 5S rRNA band from an indigenous bacterium. Partial sequencing of several 5S rRNAs demonstrated the molecular homogeneity of most of the abundant bands and enabled the identification of corresponding bacterial isolates and/or species. The dominating bacterium (around 54% of the total 5S rRNA) in the nutrient-amended mesocosms could be identified by partial sequencing as a member of the Aeromonas hydrophila complex. Another bloom of heterotrophic bacteria belonging to the Cytophaga johnsonae complex was detected in the nutrient-amended mesocosms after 13 days. The dominance of this C. johnsonae-like bacterium could even be seen in the environmental tRNAs of the bacterioplankton, where its specific tRNAs prevailed from day 13 onward. This event was also independent of the introduced nonindigenous bacteria because it occurred at the same time in all nutrient-amended mesocosms. By contrast, in the unamended experiments, a different small 5S rRNA could by observed from day 10 onward with less pronounced dominance.(ABSTRACT TRUNCATED AT 250 WORDS)
Inhibition of alkylbenzene biodegradation under denitrifying conditions by using the acetylene block technique
Authors: Hutchins S R
Published: January 2020

Addition of acetylene to microcosms simultaneously amended with nitrate and alkylbenzenes resulted in inhibition of the rate of alkylbenzene biodegradation under denitrifying conditions. Toluene, xylenes, and 1,2,4-trimethylbenzene were recalcitrant, whereas ethylbenzene was degraded at a slower rate than usual. Benzene was not degraded in either case. Addition of acetylene to microcosms preexposed to nitrate and alkylbenzenes produced similar inhibition. These data indicate that the activities of microorganisms that degrade alkylbenzenes under denitrifying conditions may be suppressed if the standard acetylene block technique is used to verify denitrifying activity.
Stable Defined Substrate for Turbidimetric Assay of Endoxylanases
Authors: Wang Ping , Broda Paul
Published: January 2020

A stable xylan suspension was prepared and characterized. Hydrolysis of the particles converts them into soluble fragments, thereby lowering the turbidity of the suspension. The small volume of the assay mixture, the short incubation time required, and the simplicity of the procedure permit the rapid analysis of many samples. Furthermore, the procedure can be used to assay xylanase activities in the presence of other reducing materials and is also useful for monitoring low-level xylanase activities.
Predicting the growth of Salmonella typhimurium on beef by using the temperature function integration technique
Authors: Dickson J S , Siragusa G R , Wray J E
Published: January 2020

Lag and generation times for the growth of Salmonella typhimurium on sterile lean beef were modeled as functions of cooling time under various carcass-chilling scenarios. Gompertz growth models were fit to the log10 colony counts over time at each of six temperatures in the range of 15 to 40 degrees C. Lag and generation times were defined as the points at which the second and first derivatives, respectively, of each growth curve attained a maximum. Generation time and lag time parameters were modeled as functions of temperature by use of exponential-decay models. The models were applied to typical beef carcass-cooling scenarios to predict the potential growth of S. typhimurium during the cooling of beef. Validation studies indicated no significant difference between the observed and predicted bacterial populations on inoculated lean and fatty beef tissues cooled at either 6 or 9 degrees C/h.
Isolation and characterization of a gene from Aspergillus parasiticus associated with the conversion of versicolorin A to sterigmatocystin in aflatoxin biosynthesis
Authors: Skory C D , Chang P K , Cary J , Linz J E
Published: January 2020

DNA isolated from the wild-type aflatoxin-producing (Afl+) fungus Aspergillus parasiticus NRRL 5862 was used to construct a cosmid genomic DNA library employing the homologous gene (pyrG) encoding orotidine monophosphate decarboxylase for selection of fungal transformants. The cosmid library was transformed into an Afl- mutant, A. parasiticus CS10 (ver-1 wh-1 pyrG), deficient in the conversion of the aflatoxin biosynthetic intermediate versicolorin A to sterigmatocystin. One pyrG+ Afl+ transformant was identified. DNA fragments from this transformant, recovered by marker rescue, contained part of the cosmid vector including the pyrG gene, the ampr gene, and a piece of the original genomic insert DNA. Transformation of these rescued DNA fragments into A. parasiticus CS10 resulted in production of wild-type levels of aflatoxin and abundant formation of sclerotia. The gene responsible for this complementation (ver-1) was identified by Northern RNA analysis and transformation with subcloned DNA fragments. The approximate locations of transcription initiation and polyadenylation sites of ver-1 were determined by an RNase protection assay and cDNA sequence analysis. The predicted amino acid sequence, deduced from the ver-1 genomic and cDNA nucleotide sequences, was compared with the EMBL and GenBank data bases. The search revealed striking similarity with Streptomyces ketoreductases involved in polyketide biosynthesis.
Identification of Distinct Communities of Sulfate-Reducing Bacteria in Oil Fields by Reverse Sample Genome Probing
Authors: Voordouw Gerrit , Voordouw Johanna K. , Jack Thomas R. , Foght Julia , Fedorak Phillip M. , Westlake Donald W. S.
Published: January 2020

Thirty-five different standards of sulfate-reducing bacteria, identified by reverse sample genome probing and defined as bacteria with genomes showing little or no cross-hybridization, were in part characterized by Southern blotting, using 16S rRNA and hydrogenase gene probes. Samples from 56 sites in seven different western Canadian oil field locations were collected and enriched for sulfate-reducing bacteria by using different liquid media containing one of the following carbon sources: lactate, ethanol, benzoate, decanoate, propionate, or acetate. DNA was isolated from the enrichments and probed by reverse sample genome probing using master filters containing denatured chromosomal DNAs from the 35 sulfate-reducing bacterial standards. Statistical analysis of the microbial compositions at 44 of the 56 sites indicated the presence of two distinct communities of sulfate-reducing bacteria. The discriminating factor between the two communities was the salt concentration of the production waters, which were either fresh water or saline. Of 34 standards detected, 10 were unique to the fresh water and 18 were unique to the saline oil field environment, while only 6 organisms were cultured from both communities.
Investigation of Mitochondrial Transmission in Selected Matings between Homokaryons from Commercial and Wild-Collected Isolates of
Agaricus bisporus
(=
Agaricus brunnescens
)
Authors: Jin Tianru , Sonnenberg Anton S. M. , Van Griensven L. J. L. D. , Horgen Paul A.
Published: January 2020

Ten heterokaryons of
Agaricus bisporus
(=
Agaricus brunnescens
) were shown to carry four different mitochondrial (mt) genotypes by analysis of mt restriction fragment length polymorphisms (RFLPs). Fifteen homokaryons derived from these strains were used to investigate mt inheritance in
A. bisporus.
One hundred eighty-nine pairings were performed in 25 different combinations. Pairings in 15 different combinations produced heterokaryons on the basis of nuclear RFLP analyses and/or fruiting trials. The mt genotype of each new intraspecies hybrid was examined by using mt RFLPs as genetic markers. Our results suggest the following. (i) Recombination between the mt genomes was not a common event. (ii) From most individual pairings, all heterokaryons carried the same mt genotype. (iii) Heterokaryons carrying either of the two possible mt genotypes were observed in certain crosses after modification of the pairing procedure. A biparental transmission pattern was demonstrated for some crosses, but there appears to be a preference for one of the mt genotypes to predominate in any specific pairing.
Synthesis of Poly(3-Hydroxybutyrate-Co-3-Hydroxyvalerate) from Methanol and
n
-Amyl Alcohol by the Methylotrophic Bacteria
Paracoccus denitrificans
and
Methylobacterium extorquens
Authors: Ueda Shunsaku , Matsumoto Seiji , Takagi Aya , Yamane Tsuneo
Published: January 2020

Strains of two types of methylotrophic bacteria,
Paracoccus denitrificans
and
Methylobacterium extorquens,
synthesized the copolyester poly(3-hydroxybutyrate-co-3-hydroxyvalerate) when methanol and
n
-amyl alcohol were added together to nitrogen-limited medium. The composition of the copolyester differed considerably between the two strains: the copolyester from
P. denitrificans
was comparatively rich in 3-hydroxyvalerate (3HV). The 3HV content of the copolyester synthesized by this strain increased with increasing concentrations of
n
-amyl alcohol. Its maximum content was 91.5 mol% under the conditions used. In
M. extorquens,
the maximum 3HV content was limited to 38.2 mol%. Since
n
-amyl alcohol served as a substrate for a standard methanol dehydrogenase, the enzyme was proposed to oxidize both methanol and
n
-amyl alcohol in the first step of copolyester synthesis from these substrates by methanol-grown cells.
Fungal metabolism of acenaphthene by Cunninghamella elegans
Authors: Pothuluri J V , Freeman J P , Evans F E , Cerniglia C E
Published: January 2020

The filamentous fungus Cunninghamella elegans ATCC 36112 metabolized within 72 h of incubation approximately 64% of the [1,8-14C]acenaphthene added. The radioactive metabolites were extracted with ethyl acetate and separated by thin-layer chromatography and reversed-phase high-performance liquid chromatography. Seven metabolites were identified by 1H nuclear magnetic resonance, UV, and mass spectral techniques as 6-hydroxyacenaphthenone (24.8%), 1,2-acenaphthenedione (19.9%), trans-1,2-dihydroxyacenaphthene (10.3%), 1,5-dihydroxyacenaphthene (2.7%), 1-acenaphthenol (2.4%), 1-acenaphthenone (2.1%), and cis-1,2-dihydroxyacenaphthene (1.8%). Parallel experiments with rat liver microsomes indicated that the major metabolite formed from acenaphthene by rat liver microsomes was 1-acenaphthenone. The fungal metabolism of acenaphthene was similar to bacterial and mammalian metabolism, since the primary site of enzymatic attack was on the two carbons of the five-member ring.
An uncooked vegan diet shifts the profile of human fecal microflora: computerized analysis of direct stool sample gas-liquid chromatography profiles of bacterial cellular fatty acids
Authors: Peltonen R , Ling W H , H�nninen O , Eerola E
Published: January 2020

The effect of an uncooked extreme vegan diet on fecal microflora was studied by direct stool sample gas-liquid chromatography (GLC) of bacterial cellular fatty acids and by quantitative bacterial culture by using classical microbiological techniques of isolation, identification, and enumeration of different bacterial species. Eighteen volunteers were divided randomly into two groups. The test group received an uncooked vegan diet for 1 month and a conventional diet of mixed Western type for the other month of the study. The control group consumed a conventional diet throughout the study period. Stool samples were collected. Bacterial cellular fatty acids were extracted directly from the stool samples and measured by GLC. Computerized analysis of the resulting fatty acid profiles was performed. Such a profile represents all bacterial cellular fatty acids in a sample and thus reflects its microflora and can be used to detect changes, differences, or similarities of bacterial flora between individual samples or sample groups. GLC profiles changed significantly in the test group after the induction and discontinuation of the vegan diet but not in the control group at any time, whereas quantitative bacterial culture did not detect any significant change in fecal bacteriology in either of the groups. The results suggest that an uncooked extreme vegan diet alters the fecal bacterial flora significantly when it is measured by direct stool sample GLC of bacterial fatty acids.
Ultrastructural and Immunocytochemical Studies on the H
2
O
2
-Producing Enzyme Pyranose Oxidase in
Phanerochaete chrysosporium
Grown under Liquid Culture Conditions
Authors: Daniel Geoffrey , Volc Jindrich , Kubatova Elena , Nilsson Thomas
Published: January 2020

The ultrastructural distribution of the sugar-oxidizing enzyme pyranose 2-oxidase (POD) in hyphae of
Phanerochaete chrysosporium
K-3 grown under liquid culture conditions optimal for the enzyme's production was studied by transmission electron microscopy immunocytochemistry. Using the 3-dimethylaminobenzoic acid-3-methyl-2-benzothiazolinone hydrazone hydrochloride H
2
O
2
peroxidase spectrophotometric assay, POD was detected in mycelial extracts from days 7 to 18, with maximum activity recorded on day 12. Onset of POD activity occurred in the secondary phase of hyphal development at a time of stationary growth, glucose limitation, and pH increase. POD was also detected extracellularly in the culture fluid from days 7 to 18, with maximum activity recorded on day 13. At early stages of development (3 to 4 days), using anti-POD antibodies and immunogold labeling, POD was localized in multivesicular and electron-dense bodies and in cell membrane regions. After 10 to 12 days of growth, at maximum POD activity, POD was concentrated within the periplasmic space where it was associated with membrane-bound vesicles and other membrane structures. At later stages of development (17 to 18 days), when the majority of hyphae were lysed, POD was observed associated with residual intracellular membrane systems and vesicles. Transmission electron microscopy immunocytochemical studies also demonstrated an extracellular distribution of the enzyme at the stationary growth phase, showing its association with fungal extracellular slime. In studies of ligninolytic cultures of the same fungus, POD was found to have a similar intracellular and extracellular distribution in slime as that recorded for cultures grown with cornsteep. POD's peripheral cytoplasmic distribution shows similarities to the cellular distribution of that reported previously for H
2
O
2
-dependent lignin and manganese peroxidases in
P. chrysosporium.
A lactococcal expression system for engineered nisins
Authors: Dodd H M , Horn N , Hao Z , Gasson M J
Published: January 2020

The nisin-producing Lactococcus lactis strain FI5876 has been modified and developed for use as an expression system for engineered nisin variants. Insertional inactivation of the resident nisA gene had a polar effect on downstream genes, including those involved in nisin immunity. However, subsequent chromosomal rearrangements in this region involving a newly discovered insertion element (IS905) generated a strain that was deficient in the nisA gene product but expressed those nisin determinants necessary for prenisin maturation, secretion, and immunity. Complementation of the lesion in the nisA gene by plasmid-encoded nisA genes containing site-specific mutations resulted in the exclusive production of altered nisins containing specific amino acid substitutions.