bluuebunny/crossref_metadata_2025_split huggingface dataset:
Molecular Cloning, Characterization, and Differential Expression of a Glucoamylase Gene from the Basidiomycetous Fungus
Lentinula edodes
Authors: Zhao J. , Chen Y. H. , Kwan H. S. 
Published: July 2002

ABSTRACT

The complete nucleotide sequence of putative glucoamylase gene
gla1
from the basidiomycetous fungus
Lentinula edodes
strain L54 is reported. The coding region of the genomic glucoamylase sequence, which is preceded by eukaryotic promoter elements CAAT and TATA, spans 2,076 bp. The
gla1
gene sequence codes for a putative polypeptide of 571 amino acids and is interrupted by seven introns. The open reading frame sequence of the
gla1
gene shows strong homology with those of other fungal glucoamylase genes and encodes a protein with an N-terminal catalytic domain and a C-terminal starch-binding domain. The similarity between the Gla1 protein and other fungal glucoamylases is from 45 to 61%, with the region of highest conservation found in catalytic domains and starch-binding domains. We compared the kinetics of glucoamylase activity and levels of gene expression in
L. edodes
strain L54 grown on different carbon sources (glucose, starch, cellulose, and potato extract) and in various developmental stages (mycelium growth, primordium appearance, and fruiting body formation). Quantitative reverse transcription PCR utilizing pairs of primers specific for
gla1
gene expression shows that expression of
gla1
was induced by starch and increased during the process of fruiting body formation, which indicates that glucoamylases may play an important role in the morphogenesis of the basidiomycetous fungus.
Phylogenetic Diversity among Geographically Dispersed
Chlamydiales
Endosymbionts Recovered from Clinical and Environmental Isolates of
Acanthamoeba
spp
Authors: Fritsche Thomas R. , Horn Matthias , Wagner Michael , Herwig Russell P. , Schleifer Karl-Heinz , Gautom Romesh K. 
Published: July 2002

ABSTRACT

The recently proposed reorganization of the order
Chlamydiales
and description of new taxa are broadening our perception of this once narrowly defined taxon. We have recovered four strains of gram-negative cocci endosymbiotic in
Acanthamoeba
spp., representing 5% of the
Acanthamoeba
sp. isolates examined, which displayed developmental life cycles typical of members of the
Chlamydiales
. One of these endosymbiont strains was found stably infecting an amoebic isolate recovered from a case of amoebic keratitis in North America, with three others found in acanthamoebae recovered from environmental sources in North America (two isolates) and Europe (one isolate). Analyses of nearly full-length 16S rRNA gene sequences of these isolates by neighbor joining, parsimony, and distance matrix methods revealed their clustering with other members of the
Chlamydiales
but in a lineage separate from those of the genera
Chlamydia
,
Chlamydophila
,
Simkania
, and
Waddlia
(sequence similarities, <88%) and including the recently described species
Parachlamydia acanthamoebae
(sequence similarities, 91.2 to 93.1%). With sequence similarities to each other of 91.4 to 99.4%, these four isolates of intra-amoebal endosymbionts may represent three distinct species and, perhaps, new genera within the recently proposed family
Parachlamydiaceae
. Fluorescently labeled oligonucleotide probes targeted to 16S rRNA signature regions were able to readily differentiate two groups of intra-amoebal endosymbionts which corresponded to two phylogenetic lineages. These results reveal significant phylogenetic diversity occurring among the
Chlamydiales
in nontraditional host species and supports the existence of a large environmental reservoir of related species. Considering that all described species of
Chlamydiales
are known to be pathogenic, further investigation of intra-amoebal parachlamydiae as disease-producing agents is warranted.
Isolation of Tannin-Degrading Lactobacilli from Humans and Fermented Foods
Authors: Osawa Ro , Kuroiso Keiko , Goto Satoshi , Shimizu Akira 
Published: July 2002

ABSTRACT

Lactobacilli with tannase activity were isolated from human feces and fermented foods. A PCR-based taxonomic assay revealed that the isolates belong to
Lactobacillus plantarum
,
L. paraplantarum
, and
L. pentosus
. Additional studies on a range of
Lactobacillus
species from established culture collections confirmed that this enzymatic activity is a phenotypic property common to these three species.
Physiological and Regulatory Effects of Controlled Overproduction of Five Cold Shock Proteins of
Lactococcus lactis
MG1363
Authors: Wouters Jeroen A. , Mailhes Marielle , Rombouts Frank M. , de Vos Willem M. , Kuipers Oscar P. , Abee Tjakko 
Published: July 2002

ABSTRACT

The physiological and regulatory effects of overproduction of five cold shock proteins (CSPs) of
Lactococcus lactis
were studied. CspB, CspD, and CspE could be overproduced at high levels (up to 19% of the total protein), whereas for CspA and CspC limited overproduction (0.3 to 0.5% of the total protein) was obtained. Northern blot analysis revealed low abundance of the
cspC
transcript, indicating that the stability of
cspC
mRNA is low. The limited overproduction of CspA is likely to be caused by low stability of CspA since when there was an Arg-Pro mutation at position 58, the level of CspA production increased. Using two-dimensional gel electrophoresis, it was found that upon overproduction of the CSPs several proteins, including a number of cold-induced proteins of
L. lactis
, were induced. Strikingly, upon overproduction of CspC induction of CspB, putative CspF, and putative CspG was also observed. Overproduction of CspB and overproduction of CspE result in increased survival when
L. lactis
is frozen (maximum increases, 10- and 5-fold, respectively, after 4 freeze-thaw cycles). It is concluded that in
L. lactis
CSPs play a regulatory role in the cascade of events that are initiated by cold shock treatment and that they either have a direct protective effect during freezing (e.g., RNA stabilization) or induce other factors involved in the freeze-adaptive response or both.
Polyhydroxyalkanoate Degradation Is Associated with Nucleotide Accumulation and Enhances Stress Resistance and Survival ofPseudomonas oleovoransin Natural Water Microcosms
Authors: Ruiz Jimena A. , López Nancy I. , Fernández Rubén O. , Méndez Beatriz S. 
Published: July 2002

ABSTRACTPseudomonas oleovoransGPo1 and its polyhydroxyalkanoic acid (PHA) depolymerization-minus mutant, GPo500phaZ, residing in natural water microcosms, were utilized to asses the effect of PHA availability on survival and resistance to stress agents. The wild-type strain showed increased survival compared to the PHA depolymerase-minus strain. The appearance of a round cellular shape, characteristic of bacteria growing under starvation conditions, was delayed in the wild type in comparison to the mutant strain. Percent survival at the end of ethanol and heat challenges was always higher in GPo1 than in GPo500. Based on these results and on early experiments (H. Hippe, Arch. Mikrobiol. 56:248–277, 1967) that suggested an association of PHA utilization with respiration and oxidative phosphorylation, we investigated the association between PHA degradation and nucleotide accumulation. ATP and guanosine tetraphosphate (ppGpp) production was analyzed under culture conditions leading to PHA depolymerization. A rise in the ATP and ppGpp levels appeared concomitant with PHA degradation, while this phenomenon was not observed in the mutant strain unable to degrade the polymer. Complementation of thephaZmutation restored the wild-type phenotype.
Clonal Populations of Thermotolerant
Enterobacteriaceae
in Recreational Water and Their Potential Interference with Fecal
Escherichia coli
Counts
Authors: McLellan Sandra L. , Daniels Annette D. , Salmore Alissa K. 
Published: July 2002

ABSTRACT

Bacterial strains were isolated from beach water samples using the original Environmental Protection Agency method for
Escherichia coli
enumeration and analyzed by pulsed-field gel electrophoresis (PFGE). Identical PFGE patterns were found for numerous isolates from 4 of the 9 days sampled, suggesting environmental replication. 16S rRNA gene sequencing, API 20E biochemical testing, and the absence of β-glucuronidase activity revealed that these clonal isolates were
Klebsiella
,
Citrobacter
, and
Enterobacter
spp. In contrast, 82% of the nonclonal isolates from water samples were confirmed to be
E. coli
, and 16% were identified as other fecal coliforms. These nonclonal isolates produced a diverse range of PFGE patterns similar to those of isolates obtained directly from untreated sewage and gull droppings. β-Glucuronidase activity was critical in distinguishing
E. coli
from other fecal coliforms, particularly for the clonal isolates. These findings demonstrate that
E. coli
is a better indicator of fecal pollution than fecal coliforms, which may replicate in the environment and falsely elevate indicator organism levels.
Accumulation of Polyhydroxyalkanoic Acid Containing Large Amounts of Unsaturated Monomers in
Pseudomonas fluorescens
BM07 Utilizing Saccharides and Its Inhibition by 2-Bromooctanoic Acid
Authors: Lee Ho-Joo , Choi Mun Hwan , Kim Tae-Un , Yoon Sung Chul 
Published: July 2002

ABSTRACT

A psychrotrophic bacterium,
Pseudomonas fluorescens
BM07, which is able to accumulate polyhydroxyalkanoic acid (PHA) containing large amounts of 3-hydroxy-
cis
-5-dodecenoate unit up to 35 mol% in the cell from unrelated substrates such as fructose, succinate, etc., was isolated from an activated sludge in a municipal wastewater treatment plant. When it was grown on heptanoic acid (C
7
) to hexadecanoic acid (C
16
) as the sole carbon source, the monomer compositional characteristics of the synthesized PHA were similar to those observed in other fluorescent pseudomonads belonging to rRNA homology group I. However, growth on stearic acid (C
18
) led to no PHA accumulation, but instead free stearic acid was stored in the cell. The existence of the linkage between fatty acid de novo synthesis and PHA synthesis was confirmed by using inhibitors such as acrylic acid and two other compounds, 2-bromooctanoic acid and 4-pentenoic acid, which are known to inhibit β-oxidation enzymes in animal cells. Acrylic acid completely inhibited PHA synthesis at a concentration of 4 mM in 40 mM octanoate-grown cells, but no inhibition of PHA synthesis occurred in 70 mM fructose-grown cells in the presence of 1 to 5 mM acrylic acid. 2-Bromooctanoic acid and 4-pentenoic acid were found to much inhibit PHA synthesis much more strongly in fructose-grown cells than in octanoate-grown cells over concentrations ranging from 1 to 5 mM. However, 2-bromooctanoic acid and 4-pentenoic acid did not inhibit cell growth at all in the fructose media. Especially, with the cells grown on fructose, 2-bromooctanoic acid exhibited a steep rise in the percent PHA synthesis inhibition over a small range of concentrations below 100 μM, a finding indicative of a very specific inhibition, whereas 4-pentenoic acid showed a broad, featureless concentration dependence, suggesting a rather nonspecific inhibition. The apparent inhibition constant
K

i

(the concentration for 50% inhibition of PHA synthesis) for 2-bromooctanoic acid was determined to be 60 μM, assuming a single-site binding of the inhibitor at a specific inhibition site. Thus, it seems likely that a coenzyme A thioester derivative of 2-bromooctanoic acid specifically inhibits an enzyme linking the two pathways, fatty acid de novo synthesis and PHA synthesis. We suggest that 2-bromooctanoic acid can substitute for the far more expensive (2,000 times) and cell-growth-inhibiting PHA synthesis inhibitor, cerulenin.
Enzyme-Linked Immunosorbent Assay Specific for (1→6) Branched, (1→3)-β-
d
-Glucan Detection in Environmental Samples
Authors: Milton Donald K. , Alwis K. Udeni , Fisette Leslie , Muilenberg Michael 
Published: July 2002

ABSTRACT

(1→3)-β-
d
-Glucans have been recognized as a potential causative agent responsible for bioaerosol-induced respiratory symptoms observed in both indoor and occupational environments. A specific enzyme immunoassay was developed to quantify (1→6) branched, (1→3)-β-
d
-glucans in environmental samples. The assay was based on the use of a high-affinity receptor (galactosyl ceramide) specific for (1→3)-β-
d
-glucans as a capture reagent and a monoclonal antibody specific for fungal cell wall β-
d
-glucans as a detector reagent. The assay was highly specific for (1→6) branched, (1→3)-β-
d
-glucans (such as that from
Saccharomyces
cerevisiae
) and did not show any response at 200 ng/ml to curdlan, laminarin, pustulan, dextran, mannan, carboxymethyl cellulose, and endotoxins. The detection level was 0.8 ng/ml for baker's yeast glucan and Betafectin. A coefficient of variation of 7.8% was obtained for (1→3)-β-
d
-glucans in house dust samples. Metal working fluids spiked with (1→3)-β-
d
-glucans inhibited the glucan assay. Because the assay is specific for (1→6) branched, (1→3)-β-
d
-glucans and is sensitive and reproducible, it will be useful for the investigation of health effects from exposure to this class of biologically active molecules.
Polysaccharide Lyase: Molecular Cloning, Sequencing, and Overexpression of the Xanthan Lyase Gene of
Bacillus
sp. Strain GL1
Authors: Hashimoto Wataru , Miki Hikaru , Tsuchiya Noriaki , Nankai Hirokazu , Murata Kousaku 
Published: July 2002

ABSTRACT

When grown on xanthan as a carbon source, the bacterium
Bacillus
sp. strain GL1 produces extracellular xanthan lyase (75 kDa), catalyzing the first step of xanthan depolymerization (H. Nankai, W. Hashimoto, H. Miki, S. Kawai, and K. Murata, Appl. Environ. Microbiol. 65:2520–2526, 1999). A gene for the lyase was cloned, and its nucleotide sequence was determined. The gene contained an open reading frame consisting of 2,793 bp coding for a polypeptide with a molecular weight of 99,308. The polypeptide had a signal peptide (2 kDa) consisting of 25 amino acid residues preceding the N-terminal amino acid sequence of the enzyme and exhibited significant homology with hyaluronidase of
Streptomyces griseus
(identity score, 37.7%).
Escherichia coli
transformed with the gene without the signal peptide sequence showed a xanthan lyase activity and produced intracellularly a large amount of the enzyme (400 mg/liter of culture) with a molecular mass of 97 kDa. During storage at 4°C, the purified enzyme (97 kDa) from
E. coli
was converted to a low-molecular-mass (75-kDa) enzyme with properties closely similar to those of the enzyme (75 kDa) from
Bacillus
sp. strain GL1, specifically in optimum pH and temperature for activity, substrate specificity, and mode of action. Logarithmically growing cells of
Bacillus
sp. strain GL1 on the medium with xanthan were also found to secrete not only xanthan lyase (75 kDa) but also a 97-kDa protein with the same N-terminal amino acid sequence as that of xanthan lyase (75 kDa). These results suggest that, in
Bacillus
sp. strain GL1, xanthan lyase is first synthesized as a preproform (99 kDa), secreted as a precursor (97 kDa) by a signal peptide-dependent mechanism, and then processed into a mature form (75 kDa) through excision of a C-terminal protein fragment with a molecular mass of 22 kDa.
The
adnA
Transcriptional Factor Affects Persistence and Spread of
Pseudomonas fluorescens
under Natural Field Conditions
Authors: Marshall Bonnie , Robleto Eduardo A. , Wetzler Richard , Kulle Peter , Casaz Paul , Levy Stuart B. 
Published: July 2002

ABSTRACT

A soil plot was inoculated with a mixture of
Pseudomonas fluorescens
Pf0-2, the wild type, and Pf0-5a, a Tn
5
insertion mutant in
adnA
, at 7.84 log CFU/g of soil. Over a period of 231 days, culturable populations of both strains were measured at selected times below and away from the point of inoculation. Pf0-5a did not spread as fast and attained significantly lower populations than Pf0-2. At sample depths below the inoculation site, the
adnA
mutant showed a significant decrease in CFU/g of soil as compared to Pf0-2. Pf0-2 was first detected at the 1.5-cm annular site at 3 days after inoculation, whereas Pf0-5a required 7 days to travel the same distance. At this distance, the wild-type strain could be detected at a 21.5- to 25-cm depth, whereas Pf0-5a could be detected only as deep as 15.5 to 18 cm. At 4.5 cm from the site of inoculation and in soil fractions corresponding to 13 to 18 cm, Pf0-2 was the only strain detected. These results suggest that the transcription factor AdnA provides a fitness advantage in
P. fluorescens
, allowing it to spread and survive in soil under field conditions.
Different Mechanisms of Resistance to
Bacillus thuringiensis
Toxins in the Indianmeal Moth
Authors: Herrero Salvador , Oppert Brenda , Ferré Juan 
Published: July 2002

ABSTRACT

Susceptibility to protoxin and toxin forms of Cry1Ab and the binding of
125
I-labeled Cry1Ab and Cry1Ac has been examined in three
Plodia interpunctella
colonies, one susceptible (688
s
) and two resistant (198
r
and Dpl
r
) to
Bacillus thuringiensis
. Toxicological studies showed that the 198
r
colony was 11-fold more resistant to Cry1Ab protoxin than to Cry1Ab activated toxin, whereas the Dpl
r
colony was 4-fold more resistant to protoxin versus toxin. Binding results with
125
I-labeled toxins indicated the occurrence of two different binding sites for Cry1Ab in the susceptible insects, one of them shared with Cry1Ac. Cry1Ab binding was found to be altered in insects from both resistant colonies, though in different ways. Compared with the susceptible colony, insects from the Dpl
r
colony showed a drastic reduction in binding affinity (60-fold higher

K
d

), although they had similar concentrations of binding sites. Insects from the 198
r
colony showed a slight reduction in both binding affinity and binding site concentration (five-fold-higher

K
d

and ca. three-fold-lower

R
t

compared with the 688
s
colony). No major difference in Cry1Ac binding was found among the three colonies. The fact that the 198
r
colony also has a protease-mediated mechanism of resistance (B. Oppert, R. Hammel, J. E. Throne, and K. J. Kramer, J. Biol. Chem. 272:23473–23476, 1997) is in agreement with our toxicological data in which this colony has a different susceptibility to the protoxin and toxin forms of Cry1Ab. It is noteworthy that the three colonies used in this work derived originally from ca. 100 insects, which reflects the high variability and high frequency of
B. thuringiensis
resistance genes occurring in natural populations.
Molecular Characterization of
Cryptosporidium
Oocysts in Samples of Raw Surface Water and Wastewater
Authors: Xiao Lihua , Singh Ajaib , Limor Josef , Graczyk Thaddeus K. , Gradus Steve , Lal Altaf 
Published: July 2002

ABSTRACT

Recent molecular characterizations of
Cryptosporidium
parasites make it possible to differentiate the human-pathogenic
Cryptosporidium
parasites from those that do not infect humans and to track the source of
Cryptosporidium
oocyst contamination in the environment. In this study, we used a small-subunit rRNA-based PCR-restriction fragment length polymorphism (RFLP) technique to detect and characterize
Cryptosporidium
oocysts in 55 samples of raw surface water collected from several areas in the United States and 49 samples of raw wastewater collected from Milwaukee, Wis.
Cryptosporidium
parasites were detected in 25 surface water samples and 12 raw wastewater samples.
C. parvum
human and bovine genotypes were the dominant
Cryptosporidium
parasites in the surface water samples from sites where there was potential contamination by humans and cattle, whereas
C. andersoni
was the most common parasite in wastewater. There may be geographic differences in the distribution of
Cryptosporidium
genotypes in surface water. The PCR-RFLP technique can be a useful alternative method for detection and differentiation of
Cryptosporidium
parasites in water.
Long-Range1H-15N Heteronuclear Shift Correlation at Natural Abundance: a Tool To Study Benzothiazole Biodegradation by TwoRhodococcusStrains
Authors: Besse Pascale , Combourieu Bruno , Boyse Gaëlle , Sancelme Martine , De Wever Heleen , Delort Anne-Marie 
Published: July 2002

ABSTRACTThe biodegradation of benzothiazole and 2-hydroxybenzothiazole by two strains ofRhodococcuswas monitored by reversed phase high-pressure liquid chromatography and by1H nuclear magnetic resonance (NMR). Both xenobiotics were biotransformed into a hydroxylated derivative of 2-hydroxybenzothiazole by these two strains. The chemical structure of this metabolite was determined by a new NMR methodology: long-range1H-15N heteronuclear shift correlation without any previous15N enrichment of the compound. This powerful NMR tool allowed us to assign the metabolite structure to 2,6-dihydroxybenzothiazole.
Mutations in the
csgD
Promoter Associated with Variations in Curli Expression in Certain Strains of
Escherichia coli
O157:H7
Authors: Uhlich Gaylen A. , Keen James E. , Elder Robert O. 
Published: July 2002

ABSTRACT

Single-base-pair
csgD
promoter mutations in human outbreak
Escherichia coli
O157:H7 strains ATCC 43894 and ATCC 43895 coincided with differential Congo red dye binding from curli fiber expression. Red phenotype
csgD
::
lacZ
promoter fusions had fourfold-greater expression than white promoter fusions. Cloning the red variant
csgDEFG
operon into white variants induced the red phenotype. Substrate utilization differed between red and white variants.
Attached and Unattached Microbial Communities in a Simulated Basalt Aquifer under Fracture- and Porous-Flow Conditions
Authors: Lehman R. Michael , Colwell Frederick S. , Bala Greg A. 
Published: July 2002

ABSTRACT

Bench scale column studies were used to examine the partitioning of microorganisms between groundwater and a geologic medium and to examine the effect of hydrogeology (i.e., porous- versus fracture-flow) on organism partitioning. Replicated columns were constructed with intact basalt core segments that contained natural fractures and with the same basalt crushed into particles. The columns were perfused with groundwater, and upon reaching a steady state, the columns were sacrificed and the attached and unattached communities were analyzed by multiple approaches. The analyses included the total number of cells, the phylogenetic affiliation of the cells (i.e., the α, β, and γ subclasses of the class
Proteobacteria
and gram positives with high G+C DNA content) by fluorescent in situ hybridization (FISH), number and taxonomic affiliation by fatty acid methyl ester profiles of culturable heterotrophs, most-probable-number estimates of methanotrophs and phenol oxidizers, and whole-community sole carbon source utilization patterns from Biolog GN microplates. In the packed columns, about 99% of the total biomass (per cubic centimeter of porous medium) was attached to the geologic medium. Lack of equitable units precluded a comparison of attached and unattached biomasses in the fractured columns where the attached biomass was expressed per unit of surface area. Compositional differences in the attached and unattached communities were evidenced by (i) the recovery of
Pseudomonas stutzeri
, an
Enterococcus
sp., and
Bacillus psychrophilus
from the groundwater and not from the basalt, (ii) differences between community carbon source utilization patterns, and (iii) the relative abundances of different phylogenetic groups estimated by FISH in both column types. In the packed columns, attached communities were depleted of members of the α- and β-
Proteobacteria
subclasses in comparison to those in the corresponding groundwater. In the fractured columns, attached communities were enriched in gram-positive
Bacteria
and γ-
Proteobacteria
and depleted of β-
Proteobacteria
, in comparison to those in the corresponding groundwater. Segregation of populations and their activities, possibly modified by attachment to geologic media, may influence contaminant fate and transport in the subsurface and impact other in situ applications.