bluuebunny/crossref_metadata_2025_split huggingface dataset:
Widespread Distribution of a Tet W Determinant among Tetracycline-Resistant Isolates of the Animal PathogenArcanobacterium pyogenes
Authors: Billington Stephen J. , Songer J. Glenn , Jost B. Helen 
Published: July 2002

ABSTRACTTetracycline resistance is common among isolates of the animal commensal and opportunistic pathogenArcanobacterium pyogenes. The tetracycline resistance determinant cloned from two bovine isolates ofA. pyogeneswas highly similar at the DNA level (92% identity) to thetet(W) gene, encoding a ribosomal protection tetracycline resistance protein, from the rumen bacteriumButyrivibrio fibrisolvens. Thetet(W) gene was found in all 20 tetracycline-resistant isolates tested, indicating that it is a widely distributed determinant of tetracycline resistance in this organism. In 25% of tetracycline-resistant isolates, thetet(W) gene was associated with amobgene, encoding a functional mobilization protein, and an origin of transfer, suggesting that the determinant may be transferable to other bacteria. In fact, low-frequency transfer oftet(W) was detected frommob+A. pyogenesisolates to a tetracycline-sensitiveA. pyogenesrecipient. The mobile nature of this determinant and the presence ofA. pyogenesin the gastrointestinal tract of cattle and pigs suggest thatA. pyogenesmay have inherited this determinant within the gastrointestinal tracts of these animals.
The Presence of Humic Substances and DNA in RNA Extracts Affects Hybridization Results
Authors: Alm Elizabeth Wheeler , Zheng Dandan , Raskin Lutgarde 
Published: July 2002

ABSTRACT
RNA extracts obtained from environmental samples are frequently contaminated with coextracted humic substances and DNA. It was demonstrated that the response in rRNA-targeted oligonucleotide probe hybridizations decreased as the concentrations of humic substances and DNA in RNA extracts increased. The decrease in hybridization signal in the presence of humic substances appeared to be due to saturation of the hybridization membrane with humic substances, resulting in a lower amount of target rRNA bound to the membrane. The decrease in hybridization response in the presence of low amounts of DNA may be the result of reduced rRNA target accessibility. The presence of high amounts of DNA in RNA extracts resulted in membrane saturation. Consistent with the observations for DNA contamination, the addition of poly(A) to RNA extracts, a common practice used to prepare RNA dilutions for membrane blotting, also reduced hybridization signals, likely because of reduced target accessibility and membrane saturation effects.
Succession of Pelagic Marine Bacteria during Enrichment: a Close Look at Cultivation-Induced Shifts
Authors: Eilers Heike , Pernthaler Jakob , Amann Rudolf 
Published: July 2002

ABSTRACT

Enrichment experiments with North Sea bacterioplankton were performed to test if rapid incubation-induced changes in community structure explain the frequent isolation of members of a few particular bacterial lineages or if readily culturable bacteria are common in the plankton but in a state of dormancy. A metabolic inhibitor of cell division (nalidixic acid [NA]) was added to substrate-amended (S+) and unamended (S−) grazer-free seawater samples, and shifts in community composition and per cell DNA and protein content were compared with untreated controls. In addition, starvation survival experiments were performed on selected isolates. Incubations resulted in rapid community shifts towards typical culturable genera rather than in the activation of either dormant cells or the original DNA-rich bacterial fraction.
Vibrio
spp. and members of the
Alteromonas/Colwellia
cluster (A/C) were selectively enriched in S+ and S−, respectively, and this trend was even magnified by the addition of NA. These increases corresponded with the rise of cell populations with distinctively different but generally higher protein and DNA content in the various treatments. Uncultured dominant γ-proteobacteria affiliating with the SAR86 cluster and members of the culturable genus
Oceanospirillum
were not enriched or activated, but there was no indication of substrate-induced cell death, either. Strains of
Vibrio
and A/C maintained high ribosome levels in pure cultures during extended periods of starvation, whereas
Oceanospirillum
spp. did not. The life strategy of rapidly enriched culturable γ-proteobacteria could thus be described as a “feast and famine” existence involving different activation levels of substrate concentration.
Integrative Food-Grade Expression System Based on the Lactose Regulon of
Lactobacillus casei
Authors: Gosalbes María José , Esteban Carlos David , Galán José Luis , Pérez-Martínez Gaspar 
Published: July 2002

ABSTRACT

The lactose operon from
Lactobacillus casei
is regulated by very tight glucose repression and substrate induction mechanisms, which made it a tempting candidate system for the expression of foreign genes or metabolic engineering. An integrative vector was constructed, allowing stable gene insertion in the chromosomal lactose operon of
L. casei
. This vector was based on the nonreplicative plasmid pRV300 and contained two DNA fragments corresponding to the 3′ end of
lacG
and the complete
lacF
gene. Four unique restriction sites were created, as well as a ribosome binding site that would allow the cloning and expression of new genes between these two fragments. Then, integration of the cloned genes into the lactose operon of
L. casei
could be achieved via homologous recombination in a process that involved two selection steps, which yielded highly stable food-grade mutants. This procedure has been successfully used for the expression of the
E. coli gusA
gene and the
L. lactis ilvBN
genes in
L. casei
. Following the same expression pattern as that for the lactose genes, β-glucuronidase activity and diacetyl production were repressed by glucose and induced by lactose. This integrative vector represents a useful tool for strain improvement in
L. casei
that could be applied to engineering fermentation processes or used for expression of genes for clinical and veterinary uses.
PCR Bias in Ecological Analysis: a Case Study for Quantitative
Taq
Nuclease Assays in Analyses of Microbial Communities
Authors: Becker Sven , Böger Peter , Oehlmann Ralfh , Ernst Anneliese 
Published: July 2002

ABSTRACT

Succession of ecotypes, physiologically diverse strains with negligible rRNA sequence divergence, may explain the dominance of small, red-pigmented (phycoerythrin-rich) cyanobacteria in the autotrophic picoplankton of deep lakes (C. Postius and A. Ernst, Arch. Microbiol. 172:69–75, 1999). In order to test this hypothesis, it is necessary to determine the abundance of specific ecotypes or genotypes in a mixed background of phylogenetically similar organisms. In this study, we examined the performance of
Taq
nuclease assays (TNAs), PCR-based assays in which the amount of an amplicon is monitored by hydrolysis of a labeled oligonucleotide (TaqMan probe) when hybridized to the amplicon. High accuracy and a 7-order detection range made the real-time TNA superior to the corresponding end point technique. However, in samples containing mixtures of homologous target sequences, quantification can be biased due to limited specificity of PCR primers and probe oligonucleotides and due to accumulation of amplicons that are not detected by the TaqMan probe. A decrease in reaction efficiency, which can be recognized by direct monitoring of amplification, provides experimental evidence for the presence of such a problem and emphasizes the need for real-time technology in quantitative PCR. Use of specific primers and probes and control of amplification efficiency allow correct quantification of target DNA in the presence of an up to 10
4
-fold excess of phylogenetically similar DNA and of an up to 10
7
-fold excess of dissimilar DNA.
Bacterial Biodegradation of Extractives and Patterns of Bordered Pit Membrane Attack in Pine Wood
Authors: Burnes Todd A. , Blanchette Robert A. , Farrell Roberta L. 
Published: July 2002

ABSTRACT

Wood extractives, commonly referred to as pitch, cause major problems in the manufacturing of pulp and paper. Treatment of nonsterile southern yellow pine chips for 14 days with
Pseudomonas fluorescens
,
Pseudomonas
sp.,
Xanthomonas campestris
, and
Serratia marcescens
reduced wood extractives by as much as 40%. Control treatments receiving only water lost 11% of extractives due to the growth of naturally occurring microorganisms. Control treatments were visually discolored after the 14-day incubation, whereas bacterium-treated wood chips were free of dark staining. Investigations using
P. fluorescens
NRRL B21432 showed that all individual resin and fatty acid components of the pine wood extractives were substantially reduced. Micromorphological observations showed that bacteria were able to colonize resin canals, ray parenchyma cells, and tracheids. Tracheid pit membranes within bordered pit chambers were degraded after treatment with
P. fluorescens
NRRL B21432.
P. fluorescens
and the other bacteria tested appear to have the potential for biological processing to substantially reduce wood extractives in pine wood chips prior to the paper making process so that problems associated with pitch in pulp mills can be controlled.
Efficient Improvement of Silage Additives by Using Genetic Algorithms
Authors: Davies Zoe S. , Gilbert Richard J. , Merry Roger J. , Kell Douglas B. , Theodorou Michael K. , Griffith Gareth W. 
Published: July 2002

ABSTRACTThe enormous variety of substances which may be added to forage in order to manipulate and improve the ensilage process presents an empirical, combinatorial optimization problem of great complexity. To investigate the utility of genetic algorithms for designing effective silage additive combinations, a series of small-scale proof of principle silage experiments were performed with fresh ryegrass. Having established that significant biochemical changes occur over an ensilage period as short as 2 days, we performed a series of experiments in which we used 50 silage additive combinations (prepared by using eight bacterial and other additives, each of which was added at six different levels, including zero [i.e., no additive]). The decrease in pH, the increase in lactate concentration, and the free amino acid concentration were measured after 2 days and used to calculate a “fitness” value that indicated the quality of the silage (compared to a control silage made without additives). This analysis also included a “cost” element to account for different total additive levels. In the initial experiment additive levels were selected randomly, but subsequently a genetic algorithm program was used to suggest new additive combinations based on the fitness values determined in the preceding experiments. The result was very efficient selection for silages in which large decreases in pH and high levels of lactate occurred along with low levels of free amino acids. During the series of five experiments, each of which comprised 50 treatments, there was a steady increase in the amount of lactate that accumulated; the best treatment combination was that used in the last experiment, which produced 4.6 times more lactate than the untreated silage. The additive combinations that were found to yield the highest fitness values in the final (fifth) experiment were assessed to determine a range of biochemical and microbiological quality parameters during full-term silage fermentation. We found that these combinations compared favorably both with uninoculated silage and with a commercial silage additive. The evolutionary computing methods described here are a convenient and efficient approach for designing silage additives.
Molecular Identification of Species from the
Penicillium roqueforti
Group Associated with Spoiled Animal Feed
Authors: Boysen Marianne E. , Jacobsson Karl-Gustav , Schnürer Johan 
Published: July 2002

ABSTRACT

The
Penicillium roqueforti
group has recently been split into three species,
P. roqueforti
,
Penicillium carneum
, and
Penicillium paneum
, on the basis of differences in ribosomal DNA sequences and secondary metabolite profiles. We reevaluated the taxonomic identity of 52 livestock feed isolates from Sweden, previously identified by morphology as
P. roqueforti
, by comparing the sequences of the ribosomal internal transcribed spacer region. Identities were confirmed with random amplified polymorphic DNA analysis and secondary metabolite profiles. Of these isolates, 48 were
P. roqueforti
, 2 were
P. paneum
, and 2 were
Penicillium expansum
. No
P. carneum
isolates were found. The three species produce different mycotoxins, but no obvious relationship between mold and animal disease was detected, based on medical records.
P. roqueforti
appears to dominate in silage, but the ecological and toxicological importance of
P. carneum
and
P. paneum
as feed spoilage fungi is not clear. This is the first report of
P. expansum
in silage.
Use of Randomly Amplified Polymorphic DNA as a Means of Developing Genus- and Strain-Specific
Streptomyces
DNA Probes
Authors: Roberts Mark A. , Crawford Don L. 
Published: July 2002

ABSTRACT

We have analyzed 20 randomly amplified polymorphic DNA (RAPD) primers against 36
Streptomyces
strains, including 17 taxonomically undefined strains, 25 nonstreptomycete actinomycetes, and 12 outgroups consisting of gram-positive and -negative species. Most of the primers were useful in identifying unique DNA polymorphisms of all strains tested. We have used RAPD techniques to develop a genus-specific probe, one not necessarily targeting the ribosomal gene, for
Streptomyces
, and a strain-specific probe for the biological control agent
Streptomyces lydicus
WYEC108. In the course of these investigations, small-scale DNA isolations were also developed for efficiently isolating actinomycete DNA. Various modifications of isolation procedures for soil DNA were compared, and the reliability and specificity of the RAPD methodology were tested by specifically detecting the
S. lydicus
WYEC108 in DNA isolated from soil.
Functional Exoenzymes as Indicators of Metabolically Active Bacteria in 124,000-Year-Old Sapropel Layers of the Eastern Mediterranean Sea
Authors: Coolen Marco J. L. , Overmann Jörg 
Published: July 2002

ABSTRACT

Hydrolytic exoenzymes as indicators of metabolically active bacteria were investigated in four consecutive sapropel layers collected from bathyal sediments of the eastern Mediterranean Sea. For comparison, the organic carbon-poor layers between the sapropels, sediment from the anoxic Urania basin, and sediments of intertidal mud flats of the German Wadden Sea were also analyzed. The sapropel layers contained up to 1.5 · 10
8
bacterial cells cm
−3
, whereas cell numbers in the intermediate layers were lower by a factor of 10. In sapropels, the determination of exoenzyme activity with fluorescently labeled substrate analogues was impaired by the strong adsorption of up to 97% of the enzymatically liberated fluorophores (4-methylumbelliferone [MUF] and 7-amino-4-methylcoumarin [MCA]) to the sediment particles. Because all established methods for the extraction of adsorbed fluorophores proved to be inadequate for sapropel sediments, we introduce a correction method which is based on the measurement of equilibrium adsorption isotherms for both compounds. Using this new approach, high activities of aminopeptidase and alkaline phosphatase were detected even in a 124,000-year-old sapropel layer, whereas the activity of β-glucosidase was low in all layers. So far, it had been assumed that the organic matter which constitutes the sapropels is highly refractory. The high potential activities of bacterial exoenzymes indicate that bacteria in Mediterranean sapropels are metabolically active and utilize part of the subfossil kerogen. Since a high adsorption capacity was determined not only for the low-molecular-weight compounds MUF and MCA but also for DNA, the extraordinarily strong adsorption of structurally different substrates to the sapropel matrix appears to be the major reason for the long-term preservation of biodegradable carbon in this environment.
Energetics of Syntrophic Propionate Oxidation in Defined Batch and Chemostat Cocultures
Authors: Scholten Johannes C. M. , Conrad Ralf 
Published: July 2002

ABSTRACT

Propionate consumption was studied in syntrophic batch and chemostat cocultures of
Syntrophobacter fumaroxidans
and
Methanospirillum hungatei
. The Gibbs free energy available for the H
2
-consuming methanogens was <−20 kJ mol of CH
4
−1
and thus allowed the synthesis of 1/3 mol of ATP per reaction. The Gibbs free energy available for the propionate oxidizer, on the other hand, was usually >−10 kJ mol of propionate
−1
. Nevertheless, the syntrophic coculture grew in the chemostat at steady-state rates of 0.04 to 0.07 day
−1
and produced maximum biomass yields of 2.6 g mol of propionate
−1
and 7.6 g mol of CH
4
−1
for
S. fumaroxidans
and
M. hungatei
, respectively. The energy efficiency for syntrophic growth of
S. fumaroxidans
, i.e., the biomass produced per unit of available Gibbs free energy was comparable to a theoretical growth yield of 5 to 12 g mol of ATP
−1
. However, a lower growth efficiency was observed when sulfate served as an additional electron acceptor, suggesting inefficient energy conservation in the presence of sulfate. The maintenance Gibbs free energy determined from the maintenance coefficient of syntrophically grown
S. fumaroxidans
was surprisingly low (0.14 kJ h
−1
mol of biomass C
−1
) compared to the theoretical value. On the other hand, the Gibbs free-energy dissipation per mole of biomass C produced was much higher than expected. We conclude that the small Gibbs free energy available in many methanogenic environments is sufficient for syntrophic propionate oxidizers to survive on a Gibbs free energy that is much lower than that theoretically predicted.
Penetration of the Coral-Bleaching Bacterium
Vibrio shiloi
into
Oculina patagonica
Authors: Banin E. , Israely T. , Kushmaro A. , Loya Y. , Orr E. , Rosenberg E. 
Published: July 2002

ABSTRACT

Inoculation of the coral-bleaching bacterium
Vibrio shiloi
into seawater containing its host
Oculina patagonica
led to adhesion of the bacteria to the coral surface via a β-
d
-galactose receptor, followed by penetration of the bacteria into the coral tissue. The internalized
V. shiloi
cells were observed inside the exodermal layer of the coral by electron microscopy and fluorescence microscopy using specific anti-
V. shiloi
antibodies to stain the intracellular bacteria. At 29°C, 80% of the bacteria bound to the coral within 8 h. Penetration, measured by the viable count (gentamicin invasion assay) inside the coral tissue, was 5.6, 20.9, and 21.7% of the initial inoculum at 8, 12, and 24 h, respectively. The viable count in the coral tissue decreased to 5.3% at 48 h, and none could be detected at 72 h. Determination of
V. shiloi
total counts (using the anti-
V. shiloi
antibodies) in the coral tissue showed results similar to viable counts for the first 12 h of infection. After 12 h, however, the total count more than doubled from 12 to 24 h and continued to rise, reaching a value 6 times that of the initial inoculum at 72 h. Thus, the intracellular
V. shiloi
organisms were transformed into a form that could multiply inside the coral tissue but did not form colonies on agar medium. Internalization of the bacteria was accompanied by the production of high concentrations of
V. shiloi
toxin P activity in the coral tissue. Internalization and multiplication of
V. shiloi
are discussed in terms of the mechanism of bacterial bleaching of corals.
Cultivation and In Situ Detection of a Thermophilic Bacterium Capable of Oxidizing Propionate in Syntrophic Association with Hydrogenotrophic Methanogens in a Thermophilic Methanogenic Granular Sludge
Authors: Imachi Hiroyuki , Sekiguchi Yuji , Kamagata Yoichi , Ohashi Akiyoshi , Harada Hideki 
Published: July 2002

ABSTRACT

The thermophilic, anaerobic, propionate-oxidizing bacterial populations present in the methanogenic granular sludge in a thermophilic (55°C) upflow anaerobic sludge blanket reactor were studied by cultivation and in situ hybridization analysis. For isolation of propionate-degrading microbes, primary enrichment was made with propionate as the sole energy source at 55°C. After several attempts to purify the microbes, a thermophilic, syntrophic, propionate-oxidizing bacterium, designated strain SI, was isolated in both pure culture and coculture with
Methanobacterium thermoautotrophicum
. Under thermophilic (55°C) conditions, strain SI oxidized propionate, ethanol, and lactate in coculture with
M. thermoautotrophicum
. In pure culture, the isolate was found to ferment pyruvate. 16S ribosomal DNA sequence analysis revealed that the strain was relatively close to members of the genus
Desulfotomaculum
, but it was only distantly related to any known species. To elucidate the abundance and spatial distribution of organisms of the strain SI type within the sludge granules, a 16S rRNA-targeted oligonucleotide probe specific for strain SI was developed and applied to thin sections of the granules. Fluorescence in situ hybridization combined with confocal laser scanning microscopy revealed that a number of rod-shaped cells were present in the middle and inner layers of the thermophilic granule sections and that they formed close associations with hydrogenotrophic methanogens. They accounted for approximately 1.1% of the total cells in the sludge. These results demonstrated that strain SI was one of the significant populations in the granular sludge and that it was responsible for propionate oxidation in the methanogenic granular sludge in the reactor.
Characterization of
Leuconostoc gasicomitatum
sp. nov., Associated with Spoiled Raw Tomato-Marinated Broiler Meat Strips Packaged under Modified-Atmosphere Conditions
Authors: Björkroth K. Johanna , Geisen Rolf , Schillinger Ulrich , Weiss Norbert , De Vos Paul , Holzapfel Wilhelm H. , Korkeala Hannu J. , Vandamme Peter 
Published: July 2002

ABSTRACT

Lactic acid bacteria (LAB) associated with gaseous spoilage of modified-atmosphere-packaged, raw, tomato-marinated broiler meat strips were identified on the basis of a restriction fragment length polymorphism (RFLP) (ribotyping) database containing DNAs coding for 16S and 23S rRNAs (rDNAs). A mixed LAB population dominated by a
Leuconostoc
species resembling
Leuconostoc gelidum
caused the spoilage of the product.
Lactobacillus sakei
,
Lactobacillus curvatus
, and a gram-positive rod phenotypically similar to heterofermentative
Lactobacillus
species were the other main organisms detected. An increase in pH together with the extreme bulging of packages suggested a rare LAB spoilage type called “protein swell.” This spoilage is characterized by excessive production of gas due to amino acid decarboxylation, and the rise in pH is attributed to the subsequent deamination of amino acids. Protein swell has not previously been associated with any kind of meat product. A polyphasic approach, including classical phenotyping, whole-cell protein electrophoresis, 16 and 23S rDNA RFLP, 16S rDNA sequence analysis, and DNA-DNA reassociation analysis, was used for the identification of the dominant
Leuconostoc
species. In addition to the RFLP analysis, phenotyping, whole-cell protein analysis, and 16S rDNA sequence homology indicated that
L. gelidum
was most similar to the spoilage-associated species. The two spoilage strains studied possessed 98.8 and 99.0% 16S rDNA sequence homology with the
L. gelidum
type strain. DNA-DNA reassociation, however, clearly distinguished the two species. The same strains showed only 22 and 34% hybridization with the
L. gelidum
type strain. These results warrant a separate species status, and we propose the name
Leuconostoc gasicomitatum
sp. nov. for this spoilage-associated
Leuconostoc
species.