bluuebunny/crossref_metadata_2025_split huggingface dataset:
Assessment of Photodynamic Destruction of
Escherichia coli
O157:H7 and
Listeria monocytogenes
by Using ATP Bioluminescence
Authors: Romanova N. A. , Brovko L. Y. , Moore L. , Pometun E. , Savitsky A. P. , Ugarova N. N. , Griffiths M. W. 
Published: November 2003

ABSTRACT

Antimicrobial photodynamic therapy was shown to be effective against a wide range of bacterial cells, as well as for fungi, yeasts, and viruses. It was shown previously that photodestruction of yeast cells treated with photosensitizers resulted in cell destruction and leakage of ATP. Three photosensitizers were used in this study: tetra(
N
-methyl-4-pyridyl)porphine tetratosylate salt (TMPyP), toluidine blue O (TBO), and methylene blue trihydrate (MB). A microdilution method was used to determine MICs of the photosensitizers against both
Escherichia coli
O157:H7 and
Listeria monocytogenes
. To evaluate the effects of photodestruction on
E. coli
and
L. monocytogenes
cells, a bioluminescence method for detection of ATP leakage and a colony-forming assay were used. All tested photosensitizers were effective for photodynamic destruction of both bacteria. The effectiveness of photosensitizers (in microgram-per-milliliter equivalents) decreased in the order TBO > MB > TMPyP for both organisms. The MICs were two- to fourfold higher for
E. coli
O157:H7 than for
L. monocytogenes
. The primary effects of all of the photosensitizers tested on live bacterial cells were a decrease in intracellular ATP and an increase in extracellular ATP, accompanied by elimination of viable cells from the sample. The time courses of photodestruction and intracellular ATP leakage were different for
E. coli
and
L. monocytogenes
. These results show that bioluminescent ATP-metry can be used for investigation of the first stages of bacterial photodestruction.
Innovative Approach for Improvement of an Antibiotic-Overproducing Industrial Strain of
Streptomyces albus
Authors: Tamehiro Norimasa , Hosaka Takeshi , Xu Jun , Hu Haifeng , Otake Noboru , Ochi Kozo 
Published: November 2003

ABSTRACT

Working with a
Streptomyces albus
strain that had previously been bred to produce industrial amounts (10 mg/ml) of salinomycin, we demonstrated the efficacy of introducing drug resistance-producing mutations for further strain improvement. Mutants with enhanced salinomycin production were detected at a high incidence (7 to 12%) among spontaneous isolates resistant to streptomycin (Str
r
), gentamicin, or rifampin (Rif
r
). Finally, we successfully demonstrated improvement of the salinomycin productivity of the industrial strain by 2.3-fold by introducing a triple mutation. The Str
r
mutant was shown to have a point mutation within the
rpsL
gene (encoding ribosomal protein S12). Likewise, the Rif
r
mutant possessed a mutation in the
rpoB
gene (encoding the RNA polymerase β subunit). Increased productivity of salinomycin in the Str
r
mutant (containing the K88R mutation in the S12 protein) may be a result of an aberrant protein synthesis mechanism. This aberration may manifest itself as enhanced translation activity in stationary-phase cells, as we have observed with the poly(U)-directed cell-free translation system. The K88R mutant ribosome was characterized by increased 70S complex stability in low Mg
2+
concentrations. We conclude that this aberrant protein synthesis ability in the Str
r
mutant, which is a result of increased stability of the 70S complex, is responsible for the remarkable salinomycin production enhancement obtained.
Nitrogen-Regulated Hypermutator Strain of
Synechococcus
sp. for Use in In Vivo Artificial Evolution
Authors: Emlyn-Jones Daniel , Price G. Dean , Andrews T. John 
Published: November 2003

ABSTRACT

Artificially evolved variants of proteins with roles in photosynthesis may be selected most conveniently by using a photosynthetic organism, such as a cyanobacterium, whose growth depends on the function of the target protein. However, the limited transformation efficiency of even the most transformable cyanobacteria wastes much of the diversity of mutant libraries of genes produced in vitro, impairing the coverage of sequence space. This highlights the advantages of an in vivo approach for generating diversity in the selection organism itself. We constructed two different hypermutator strains of
Synechococcus
sp. strain PCC 7942 by insertionally inactivating or nutritionally repressing the DNA mismatch repair gene,
mutS
. Inactivation of
mutS
greatly increases the mutation rate of the cyanobacterium's genes, leading to an up-to-300-fold increase in the frequency of resistance to the antibiotics rifampin and spectinomycin. In order to control the rate of mutation and to limit cellular damage resulting from prolonged hypermutation, we placed the uninterrupted
mutS
gene in the cyanobacterial chromosome under the transcriptional control of the cyanobacterial
nirA
promoter, which is repressed in the presence of NH
4
+
as an N source and derepressed in its absence. By removing or adding this substrate, hypermutation was activated or repressed as required. As expected, hypermutation caused by repression in P
nir
A-
mut
S transformants led to an accumulation of spectinomycin resistance mutations during growth.
Glycolytic Breakdown of Sulfoquinovose in Bacteria: a Missing Link in the Sulfur Cycle
Authors: Roy Alexander B. , Hewlins Michael J. E. , Ellis Andrew J. , Harwood John L. , White Graham F. 
Published: November 2003

ABSTRACTSulfoquinovose (6-deoxy-6-sulfo-d-glucopyranose), formed by the hydrolysis of the plant sulfolipid, is a major component of the biological sulfur cycle. However, pathways for its catabolism are poorly delineated. We examined the hypothesis that mineralization of sulfoquinovose to inorganic sulfate is initiated by reactions of the glycolytic and/or Entner-Doudoroff pathways in bacteria. Metabolites of [U-13C]sulfoquinovose were identified by13C-nuclear magnetic resonance (NMR) in strains ofKlebsiellaandAgrobacteriumpreviously isolated for their ability to utilize sulfoquinovose as a sole source of carbon and energy for growth, and cell extracts were analyzed for enzymes diagnostic for the respective pathways.Klebsiellasp. strain ABR11 grew rapidly on sulfoquinovose, with major accumulations of sulfopropandiol (2,3-dihydroxypropanesulfonate) but no detectable release of sulfate. Later, when sulfoquinovose was exhausted and growth was very slow, sulfopropandiol disappeared and inorganic sulfate and small amounts of sulfolactate (2-hydroxy-3-sulfopropionate) were formed. InAgrobacteriumsp. strain ABR2, growth and sulfoquinovose disappearance were again coincident, though slower than that inKlebsiellasp. Release of sulfate was still late but was faster than that inKlebsiellasp., and no metabolites were detected by13C-NMR. Extracts of both strains grown on sulfoquinovose contained phosphofructokinase activities that remained unchanged when fructose 6-phosphate was replaced in the assay mixture with either glucose 6-phosphate or sulfoquinovose. The results were consistent with the operation of the Embden-Meyerhoff-Parnas (glycolysis) pathway for catabolism of sulfoquinovose. Extracts ofKlebsiellabut notAgrobacteriumalso contained an NAD+-dependent sulfoquinovose dehydrogenase activity, indicating that the Entner-Doudoroff pathway might also contribute to catabolism of sulfoquinovose.
A New
Bacteroides
Conjugative Transposon That Carries an
ermB
Gene
Authors: Gupta Anamika , Vlamakis Hera , Shoemaker Nadja , Salyers Abigail A. 
Published: November 2003

ABSTRACT

The erythromycin resistance gene
ermB
has been found in a variety of gram-positive bacteria. This gene has also been found in
Bacteroides
species but only in six recently isolated strains; thus, the gene seems to have entered this genus only recently. One of the six
Bacteroides ermB
-containing isolates, WH207, could transfer
ermB
to
Bacteroides thetaiotaomicron
strain BT4001 by conjugation. WH207 was identified as a
Bacteroides uniformis
strain based on the sequence of its 16S rRNA gene. Results of pulsed-field gel electrophoresis experiments demonstrated that the transferring element was normally integrated into the
Bacteroides
chromosome. The element was estimated from pulsed-field gel data to be about 100 kb in size. Since the element appeared to be a conjugative transposon (CTn), it was designated CTnBST. CTnBST was able to mobilize coresident plasmids and the circular form of the mobilizable transposon NBU1 to
Bacteroides
and
Escherichia coli
recipients. A 13-kb segment that contained
ermB
was cloned and sequenced. Most of the open reading frames in this region had little similarity at the amino acid sequence level to any proteins in the sequence databases, but a 1,723-bp DNA segment that included a 950-bp segment downstream of
ermB
had a DNA sequence that was virtually identical to that of a segment of DNA found previously in a
Clostridium perfringens
strain. This finding, together with the finding that
ermB
is located on a CTn, supports the hypothesis that CTnBST could have entered
Bacteroides
from some other genus, possibly from gram-positive bacteria. Moreover, this finding supports the hypothesis that many transmissible antibiotic resistance genes in
Bacteroides
are carried on CTns.
Low-Temperature Isolation of Disease-Suppressive Bacteria and Characterization of a Distinctive Group of Pseudomonads
Authors: Johansson P. Maria , Wright Sandra A. I. 
Published: November 2003

ABSTRACT

The influence of environmental factors during isolation on the composition of potential biocontrol isolates is largely unknown. Bacterial isolates that efficiently suppressed wheat seedling blight caused by
Fusarium culmorum
were found by isolating psychrotrophic, root-associated bacteria and by screening them in a bioassay that mimicked field conditions. The impact of individual isolation factors on the disease-suppressive index (DSI) of almost 600 isolates was analyzed. The bacteria originated from 135 samples from 62 sites in Sweden and Switzerland. The isolation factors that increased the probability of finding isolates with high DSIs were sampling from arable land, Swiss origin of samples, and origination of isolates from plants belonging to the family Brassicaceae. The colony morphology of the isolates was characterized and compared to DSIs, which led to identification of a uniform morphological group containing 57 highly disease-suppressive isolates. Isolates in this group were identified as
Pseudomonas
sp.; they were fluorescent on King's medium B and had characteristic crystalline structures in their colonies. These isolates were morphologically similar to seven strains that had previously been selected for suppression of barley net blotch caused by
Drechslera teres
. Members of this morphological group grow at 1.5°C and produce an antifungal polyketide (2,3-deepoxy-2,3-didehydrorhizoxin [DDR]). They have similar two-dimensional polyacrylamide gel electrophoresis protein profiles, phenotypic characteristics, and in vitro inhibition spectra of pathogens. In summary, in this paper we describe some isolation factors that are important for obtaining disease-suppressive bacteria in our system, and we describe a novel group of biocontrol pseudomonads.
Methicillin (Oxacillin)-Resistant
Staphylococcus aureus
Strains Isolated from Major Food Animals and Their Potential Transmission to Humans
Authors: Lee John Hwa 
Published: November 2003

ABSTRACT

From May 2001 to April 2003, various types of specimens from cattle, pigs, and chickens were collected and examined for the presence of methicillin (oxacillin)-resistant
Staphylococcus aureus
(MRSA).
S. aureus
was isolated and positively identified by using Gram staining, colony morphology, tests for coagulase and urease activities, and an API Staph Ident system. Among 1,913 specimens collected from the animals, 421 contained
S. aureus
; of these, 28 contained
S. aureus
resistant to concentrations of oxacillin higher than 2 μg/ml. Isolates from 15 of the 28 specimens were positive by PCR for the
mecA
gene. Of the 15
mecA
-positive MRSA isolates, 12 were from dairy cows and 3 were from chickens. Antimicrobial susceptibility tests of
mecA
-positive MRSA strains were performed by the disk diffusion method. All isolates were resistant to members of the penicillin family, such as ampicillin, oxacillin, and penicillin. All isolates were also susceptible to amikacin, vancomycin, and trimethoprim-sulfamethoxazole. To determine molecular epidemiological relatedness of these 15 animal MRSA isolates to isolates from humans, random amplified polymorphic DNA (RAPD) patterns were generated by arbitrarily primed PCR. The RAPD patterns of six of the isolates from animals were identical to the patterns of certain isolates from humans. The antibiotypes of the six animal isolates revealed types similar to those of the human isolates. These data suggested that the genomes of the six animal MRSA isolates were very closely related to those of some human MRSA isolates and were a possible source of human infections caused by consuming contaminated food products made from these animals.
Construction of
Escherichia coli
Strains for Conversion of Nitroacetophenones to
ortho
-Aminophenols
Authors: Kadiyala Venkateswarlu , Nadeau Lloyd J. , Spain Jim C. 
Published: November 2003

ABSTRACT

The predominant bacterial pathway for nitrobenzene (NB) degradation uses an NB nitroreductase and hydroxylaminobenzene (HAB) mutase to form the ring-fission substrate
ortho
-aminophenol. We tested the hypothesis that constructed strains might accumulate the aminophenols from nitroacetophenones and other nitroaromatic compounds. We constructed a recombinant plasmid carrying NB nitroreductase (
nbzA
) and HAB mutase A (
habA
) genes, both from
Pseudomonas pseudoalcaligenes
JS45, and expressed the enzymes in
Escherichia coli
JS995. IPTG (isopropyl-β-
d
-thiogalactopyranoside)-induced cells of strain JS995 rapidly and stoichiometrically converted NB to 2-aminophenol, 2-nitroacetophenone (2NAP) to 2-amino-3-hydroxyacetophenone (2AHAP), and 3-nitroacetophenone (3NAP) to 3-amino-2-hydroxyacetophenone (3AHAP). We constructed another recombinant plasmid containing the nitroreductase gene (
nfs1
) from
Enterobacter cloacae
and
habA
from strain JS45 and expressed the enzymes in
E. coli
JS996. Strain JS996 converted NB to 2-aminophenol, 2-nitrotoluene to 2-amino-3-methylphenol, 3-nitrotoluene to 2-amino-4-methylphenol, 4-nitrobiphenyl ether to 4-amino-5-phenoxyphenol, and 1-nitronaphthalene to 2-amino-1-naphthol. In larger-scale biotransformations catalyzed by strain JS995, 75% of the 2NAP transformed was converted to 2AHAP, whereas 3AHAP was produced stoichiometrically from 3NAP. The final yields of the aminophenols after extraction and recovery were >64%. The biocatalytic synthesis of
ortho
-aminophenols from nitroacetophenones suggests that strain JS995 may be useful in the biocatalytic production of a variety of substituted
ortho
-aminophenols from the corresponding nitroaromatic compounds.
Population Structure of
Alexandrium
(Dinophyceae) Cyst Formation-Promoting Bacteria in Hiroshima Bay, Japan
Authors: Adachi Masao , Kanno Tsukasa , Okamoto Ryo , Itakura Shigeru , Yamaguchi Mineo , Nishijima Toshitaka 
Published: November 2003

ABSTRACT

A total of 31 bacterial isolates that have potential
Alexandrium
cyst formation-promoting activity (Alex-CFPB) were isolated from Hiroshima Bay (Japan), which is characterized by seasonal blooms of the toxic dinoflagellate
Alexandrium tamarense
. The population structure of Alex-CFPB was analyzed by means of restriction fragment length polymorphism analysis of the 16S rRNA genes (16S rDNA). Fourteen ribotypes, A to N, were observed among the 31 isolates of Alex-CFPB by using four restriction enzymes,
Mbo
I,
Hha
I,
Rsa
I and
Bst
UI. Among them, seven isolates, which were obtained from the seawater samples taken during the peak and termination periods of the
A. tamarense
bloom in 1998, belonged to ribotype A. This result suggests that bacterial strains of ribotype A may be dominant in the Alex-CFPB assemblages during these periods. The partial 16S rDNA-based phylogenetic tree of 10 ribotypes studied showed that nine of them fell into the
Rhodobacter
group of the α subclass of the
Proteobacteria
. Eight of nine ribotypes of the
Rhodobacter
group fell into the lineage of the
Roseobacter
subgroup, and one fell into the
Rhodobacter
subgroup. The non-
Rhodobacter
group type fell into the
Marinobacterium
-
Neptunomonas
-
Pseudomonas
group of the γ-
Proteobacteria
. Isolates of Alex-CFPB ribotypes A and C do not have clear growth-promoting activities but have strong cyst formation-promoting activities (CFPAs) under our laboratory conditions. These results show that the Alex-CFPB assemblage may consist of various bacteria that belong mainly to the
Roseobacter
group and have strong CFPAs. These results suggest that not only the
Alexandrium
cyst formation-inhibiting bacteria (Alex-CFIB) reported previously but also Alex-CFPB, especially bacteria of ribotype A, may play significant roles in the process of encystment and bloom dynamics of
Alexandrium
in the natural environment.
Diversity and Structure of Bacterial Communitiesin Arctic versus Antarctic PackIce
Authors: Brinkmeyer Robin , Knittel Katrin , Jürgens Jutta , Weyland Horst , Amann Rudolf , Helmke Elisabeth 
Published: November 2003

ABSTRACT

A
comprehensive assessment of bacterial diversity and community
composition in arctic and antarctic pack ice was conducted through
cultivation and cultivation-independent molecular techniques. We
sequenced 16S rRNA genes from 115 and 87 pure cultures of bacteria
isolated from arctic and antarctic pack ice, respectively. Most of the
33 arctic phylotypes were >97% identical to previously
described antarctic species or to our own antarctic isolates. At both
poles, the α- and γ-proteobacteria and the
Cytophaga
-
Flavobacterium
group were the dominant
taxonomic bacterial groups identified by cultivation as well as by
molecular methods. The analysis of 16S rRNA gene clone libraries from
multiple arctic and antarctic pack ice samples revealed a high
incidence of closely overlapping 16S rRNA gene clone and isolate
sequences. Simultaneous analysis of environmental samples with
fluorescence in situ hybridization (FISH) showed that∼
95% of 4′,6′-diamidino-2-phenylindole
(DAPI)-stained cells hybridized with the general bacterial probe
EUB338. More than 90% of those were further assignable.
Approximately 50 and 36% were identified asγ
-proteobacteria in arctic and antarctic samples,respectively. Approximately 25% were identified asα
-proteobacteria, and 25% were identified as belonging
to the
Cytophaga
-
Flavobacterium
group. For the
quantification of specific members of the sea ice community, new
oligonucleotide probes were developed which target the genera
Octadecabacter
,
Glaciecola
,
Psychrobacter
,
Marinobacter
,
Shewanella
, and
Polaribacter
.
High FISH detection rates of these groups as well as high viable counts
corroborated the overlap of clone and isolate sequences. A terrestrial
influence on the arctic pack ice community was suggested by the
presence of limnic
phylotypes.
Physicochemical Conditions and Microbial Activities in the Highly Alkaline Gut of the Humus-Feeding Larva of
Pachnoda ephippiata
(Coleoptera: Scarabaeidae)
Authors: Lemke Thorsten , Stingl Ulrich , Egert Markus , Friedrich Michael W. , Brune Andreas 
Published: November 2003

ABSTRACT

The soil macrofauna plays an important role in the carbon and nitrogen cycle of terrestrial ecosystems. In order to gain more insight into the role of the intestinal microbiota in transformation and mineralization of organic matter during gut passage, we characterized the physicochemical conditions, microbial activities, and community structure in the gut of our model organism, the humus-feeding larva of the cetoniid beetle
Pachnoda ephippiata
. Microsensor measurements revealed an extreme alkalinity in the midgut, with highest values (pH > 10) between the second and third crown of midgut ceca. Both midgut and hindgut were largely anoxic, but despite the high pH, the redox potential of the midgut content was surprisingly high even in the largest instar. However, reducing conditions prevailed in the hindgut paunch of all instars (E
h
∼ −100 mV). Both gut compartments possessed a pronounced gut microbiota, with highest numbers in the hindgut, and microbial fermentation products were present in high concentrations. The stimulation of hindgut methanogenesis by exogenous electron donors, such as H
2
, formate, and methanol, together with considerable concentrations of formate in midgut and hemolymph, suggests that midgut fermentations are coupled to methanogenesis in the hindgut by an intercompartmental transfer of reducing equivalents via the hemolymph. The results of a cultivation-based enumeration of the major metabolic groups in midgut and hindgut, which yielded high titers of lactogenic, propionigenic, and acetogenic bacteria, are in good agreement not only with the accumulation of microbial fermentation products in the respective compartments but also with the results of a cultivation-independent characterization of the bacterial communities reported in the companion paper (M. Egert, B. Wagner, T. Lemke, A. Brune, and M. W. Friedrich, Appl. Environ. Microbiol. 69:6659-6668, 2003).
Microbial Community Structure in Midgut and Hindgut of the Humus-Feeding Larva of
Pachnoda ephippiata
(Coleoptera: Scarabaeidae)
Authors: Egert Markus , Wagner Bianca , Lemke Thorsten , Brune Andreas , Friedrich Michael W. 
Published: November 2003

ABSTRACT

The guts of soil-feeding macroinvertebrates contain a complex microbial community that is involved in the transformation of ingested soil organic matter. In a companion paper (T. Lemke, U. Stingl, M. Egert, M. W. Friedrich, and A. Brune, Appl. Environ. Microbiol. 69:6650-6658, 2003), we show that the gut of our model organism, the humivorous larva of the cetoniid beetle
Pachnoda ephippiata
, is characterized by strong midgut alkalinity, high concentrations of microbial fermentation products, and the presence of a diverse, yet unstudied microbial community. Here, we report on the community structure of bacteria and archaea in the midgut, hindgut, and food soil of
P. ephippiata
larvae, determined with cultivation-independent techniques. Clone libraries and terminal restriction fragment length polymorphism analysis of 16S rRNA genes revealed that the intestines of
P. ephippiata
larvae contain a complex gut microbiota that differs markedly between midgut and hindgut and that is clearly distinct from the microbiota in the food soil. The bacterial community is dominated by phylogenetic groups with a fermentative metabolism (
Lactobacillales
,
Clostridiales
,
Bacillales
, and
Cytophaga-Flavobacterium-Bacteroides
[CFB] phylum), which is corroborated by high lactate and acetate concentrations in the midgut and hindgut and by the large numbers of lactogenic and acetogenic bacteria in both gut compartments reported in the companion paper. Based on 16S rRNA gene frequencies,
Actinobacteria
dominate the alkaline midgut, while the hindgut is dominated by members of the CFB phylum. The archaeal community, however, is less diverse. 16S rRNA genes affiliated with mesophilic
Crenarchaeota
, probably stemming from the ingested soil, were most frequent in the midgut, whereas
Methanobacteriaceae
-related 16S rRNA genes were most frequent in the hindgut. These findings agree with the reported restriction of methanogenesis to the hindgut of
Pachnoda
larvae.