bluuebunny/crossref_metadata_2025 huggingface dataset:
Differential Effects of Nitrogenous Fertilizers on Methane-Consuming Microbes in Rice Field and Forest Soils
Authors: Mohanty Santosh R. , Bodelier Paul L. E. , Floris Virgilio , Conrad Ralf
Published: February 2006

The impact of environmental perturbation (e.g., nitrogenous fertilizers) on the dynamics of methane fluxes from soils and wetland systems is poorly understood. Results of fertilizer studies are often contradictory, even within similar ecosystems. In the present study the hypothesis of whether these contradictory results may be explained by the composition of the methane-consuming microbial community and hence whether methanotrophic diversity affects methane fluxes was investigated. To this end, rice field and forest soils were incubated in microcosms and supplemented with different nitrogenous fertilizers and methane concentrations. By labeling the methane with
13
C, diversity and function could be coupled by analyses of phospholipid-derived fatty acids (PLFA) extracted from the soils at different time points during incubation. In both rice field and forest soils, the activity as well as the growth rate of methane-consuming bacteria was affected differentially. For type I methanotrophs, fertilizer application stimulated the consumption of methane and the subsequent growth, while type II methanotrophs were generally inhibited. Terminal restriction fragment length polymorphism analyses of the
pmoA
gene supported the PLFA results. Multivariate analyses of stable-isotope-probing PLFA profiles indicated that in forest and rice field soils,
Methylocystis
(type II) species were affected by fertilization. The type I methanotrophs active in forest soils (
Methylomicrobium/Methylosarcina
related) differed from the active species in rice field soils (
Methylobacter/Methylomonas
related). Our results provide a case example showing that microbial community structure indeed matters, especially when assessing and predicting the impact of environmental change on biodiversity loss and ecosystem functioning.
Host-Symbiont Relationships in Hydrothermal Vent Gastropods of the Genus
Alviniconcha
from the Southwest Pacific
Authors: Suzuki Yohey , Kojima Shigeaki , Sasaki Takenori , Suzuki Masae , Utsumi Takashi , Watanabe Hiromi , Urakawa Hidetoshi , Tsuchida Shinji , Nunoura Takuro , Hirayama Hisako , Takai Ken , Nealson Kenneth H. , Horikoshi Koki
Published: February 2006

Hydrothermal vent gastropods of the genus
Alviniconcha
are unique among metazoans in their ability to derive their nutrition from chemoautotrophic γ- and ε-proteobacterial endosymbionts. Although host-symbiont relationships in
Alviniconcha
gastropods from the Central Indian Ridge in the Indian Ocean and the Mariana Trough in the Western Pacific have been studied extensively, host-symbiont relationships in
Alviniconcha
gastropods from the Southwest Pacific remain largely unknown. Phylogenetic analysis using mitochondrial cytochrome
c
oxidase subunit I gene sequences of host gastropods from the Manus, North Fiji, and Lau Back-Arc Basins in the Southwest Pacific has revealed a new host lineage in a
Alviniconcha
gastropod from the Lau Basin and the occurrence of the host lineage
Alviniconcha
sp. type 2 in the Manus Basin. Based on 16S rRNA gene sequences of bacterial endosymbionts, two γ-proteobacterial lineages and one ε-proteobacterial lineage were identified in the present study. The carbon isotopic compositions of the biomass and fatty acids of the gastropod tissues suggest that the γ- and ε-proteobacterial endosymbionts mediate the Calvin-Benson cycle and the reductive tricarboxylic acid cycle, respectively, for their chemoautotrophic growth. Coupling of the host and symbiont lineages from the three Southwest Pacific basins revealed that each of the
Alviniconcha
lineages harbors different bacterial endosymbionts belonging to either the γ- or ε-
Proteobacteria
. The host specificity exhibited in symbiont selection provides support for the recognition of each of the host lineages as a distinct species. The results from the present study also suggest the possibility that
Alviniconcha
sp. types 1 and 2 separately inhabit hydrothermal vent sites approximately 120 m apart in the North Fiji Basin and 500 m apart in the Manus Basin.
The Thin Pili of
Acinetobacter
sp. Strain BD413 Mediate Adhesion to Biotic and Abiotic Surfaces
Authors: Gohl Olivia , Friedrich Alexandra , Hoppert Michael , Averhoff Beate
Published: February 2006

Two structurally different appendages, thin and thick pili, are found in members of the genus
Acinetobacter
. The presence of pilus structures correlates with different phenotypes, such as adherence to surfaces, a trait not only observed in pathogenic
Acinetobacter
species, as well as motility. However, their distinct individual roles were unknown. To characterize the role of different pili in the physiology of
Acinetobacter
, we isolated the thin pili from the cell surface of
Acinetobacter
sp. strain BD413 (recently recognized as representative of
Acinetobacter baylyi
), a soil bacterium that rapidly takes up naked DNA from its environment. Electron microcopy revealed that the pilus has an external diameter of 2 to 3 nm for single filaments. The filaments are packed into right-handed bundles. The major protein constituting the pilus was purified, and the encoding gene,
acuA
, was cloned. AcuA was found to be weakly related to the structural subunit of F17 pili of
Escherichia coli
. Analyses of the
acuA
flanking DNA region led to the identification of three closely associated genes,
acuD
,
acuC
, and
acuG
, whose deduced proteins are similar to chaperone, usher, and adhesin of F17-related pili, respectively. Transcriptional analyses revealed that
acuA
expression is maximal in the late-stationary-growth phase. Mutation of
acuA
led to a loss of thin pili and concomitantly loss of adhesion to polystyrene and erythrocytes but not loss of competence. Therefore, thin pili of
Acinetobacter
sp. strain BD413 are suggested to be assembled by the chaperone/usher pathway and are involved in adherence to biotic and abiotic surfaces.
Engineering the Genotype ofAcinetobactersp. Strain ADP1 To Enhance Biosynthesis of Cyanophycin
Authors: Elbahloul Yasser , Steinbüchel Alexander
Published: February 2006

To study the importance of arginine provision and phosphate limitation for synthesis and accumulation of cyanophycin (CGP) inAcinetobactersp. strain ADP1, genes encoding the putative arginine regulatory protein (argR) and the arginine succinyltransferase (astA) were inactivated, and the effects of these mutations on CGP synthesis were analyzed. The inactivation of these genes resulted in a 3.5- or 7-fold increase in CGP content, respectively, when the cells were grown on glutamate. Knockout mutations in both genes led to a better understanding of the effect of the addition of other substrates to arginine on CGP synthesis during growth of the cells ofAcinetobactersp. strain ADP1. Overexpression of ArgF (ornithine carbamoyltransferase), CarA-CarB (small and large subunits of carbamoylphosphate synthetase), and PepC (phosphoenolpyruvate carboxylase) triggered synthesis of CGP if amino acids were used as a carbon source whereas it was not triggered by gluconate or other sugars. Cells ofAcinetobactersp. strain ADP1, which is largely lacking genes for carbohydrate metabolism, showed a significant increase in CGP contents when grown on mineral medium supplemented with glutamate, aspartate, or arginine. TheAcinetobactersp.ΔastA(pYargF) strain is unable to utilize arginine but synthesizes more arginine, resulting in CGP contents as high as 30% and 25% of cell dry matter when grown on protamylasse or Luria-Bertani medium, respectively. This recombinant strain overcame the bottleneck of the costly arginine provision where it produces about 75% of the CGP obtained from the parent cells grown on mineral medium containing pure arginine as the sole source of carbon. Phosphate starvation is the only known trigger for CGP synthesis in this bacterium, which possesses the PhoB/PhoR phosphate regulon system. Overexpression ofphoBcaused an 8.6-fold increase in CGP content in comparison to the parent strain at a nonlimiting phosphate concentration.
First Insights into the Evolution of
Streptococcus uberis
: a Multilocus Sequence Typing Scheme That Enables Investigation of Its Population Biology
Authors: Coffey Tracey J. , Pullinger Gillian D. , Urwin Rachel , Jolley Keith A. , Wilson Stephen M. , Maiden Martin C. , Leigh James A.
Published: February 2006

Intramammary infection with
Streptococcus uberis
is a common cause of bovine mastitis throughout the world. Several procedures to differentiate
S. uberis
isolates have been proposed. However, all are prone to interlaboratory variation, and none is suitable for the description of the population structure. We describe here the development of a multilocus sequence typing (MLST) scheme for
S. uberis
to help address these issues. The sequences of seven housekeeping gene fragments from each of 160 United Kingdom milk isolates of
S. uberis
were determined. Between 5 and 17 alleles were obtained per locus, giving the potential to discriminate between 1.3 × 10
7
sequence types. In this study, 57 sequence types (STs) were identified. Statistical comparisons between the maximum-likelihood trees constructed by using the seven housekeeping gene fragments showed that the congruence was no better than that between each tree and trees of random topology, indicating there had been significant recombination within these loci. The population contained one major lineage (designated the ST-5 complex). This dominated the population, containing 24 STs and representing 112 isolates. The other 33 STs were not assigned to any clonal complex. All of the isolates in the ST-5 lineage carried
hasA
, a gene that is essential for capsule production. There was no clear association between ST or clonal complex and disease. The
S. uberis
MLST system offers researchers a valuable tool that allows further investigation of the population biology of this organism and insights into the epidemiology of this disease on a global scale.
Application of
Streptococcus uberis
Multilocus Sequence Typing: Analysis of the Population Structure Detected among Environmental and Bovine Isolates from New Zealand and the United Kingdom
Authors: Pullinger Gillian D. , López-Benavides Mario , Coffey Tracey J. , Williamson John H. , Cursons Ray T. , Summers Emma , Lacy-Hulbert Jane , Maiden Martin C. , Leigh James A.
Published: February 2006

We recently developed a multilocus sequence typing (MLST) scheme to differentiate
S. uberis
isolates and facilitate an understanding of the population biology of this pathogen. The scheme was initially used to study a collection of 160 bovine milk isolates from the United Kingdom and showed that the majority of isolates were from one clonal complex (designated the ST-5 complex). Here we describe the MLST analysis of a collection of New Zealand isolates. These were obtained from diverse sources, including bovine milk, other bovine anatomical sites, and environmental sources. The complete allelic profiles of 253 isolates were determined. The collection was highly diverse and included 131 different sequence types (STs). The New Zealand and United Kingdom populations were distinct, since none of the 131 STs were represented within the previously studied collection of 160 United Kingdom
S. uberis
isolates. However, seven of the STs were members of the ST-5 clonal complex, the major complex within the United Kingdom collection. Two new clonal complexes were identified: ST-143 and ST-86. All three major complexes were isolated from milk, other bovine sites, and the environment. Carriage of the
hasA
gene, which is necessary for capsule formation, correlated with clonal complex and isolation from clinical cases of mastitis.
In Situ and In Vitro Gene Expression by
Vibrio vulnificus
during Entry into, Persistence within, and Resuscitation from the Viable but Nonculturable State
Authors: Smith Ben , Oliver James D.
Published: February 2006

Isolation of
Vibrio vulnificus
during winter months is difficult due to the entrance of these cells into the viable but nonculturable (VBNC) state. While several studies have investigated in vitro gene expression upon entrance into and persistence within the VBNC state, to our knowledge, no in situ studies have been reported. We incubated clinical and environmental isolates of
V. vulnificus
in estuarine waters during winter months to monitor the expression of several genes during the VBNC state and compared these to results from in vitro studies.
katG
(periplasmic catalase) was down-regulated during the VBNC state in vitro and in situ compared to the constitutively expressed gene
tufA
. Our results indicate that the loss of catalase activity we previously reported is a direct result of
katG
repression, which likely accounts for the VBNC response of this pathogen. While expression of
vvhA
(hemolysin) was detectable in environmental strains during in situ incubation, it ceased in all cases by ca. 1 h. These results suggest that the natural role of hemolysin in
V. vulnificus
may be in osmoprotection and/or the cold shock response. Differences in expression of the capsular genes
wza
and
wzb
were observed in the two recently reported genotypes of this species. Expression of
rpoS
, encoding the stress sigma factor RpoS, was continuous upon entry into the VBNC state during both in situ and in vitro studies. We found the half-life of mRNA to be less than 60 minutes, confirming that mRNA detection in these VBNC cells is a result of de novo RNA synthesis.
Paenibacillus
sp. Strain JDR-2 and XynA
1
: a Novel System for Methylglucuronoxylan Utilization
Authors: StJohn Franz J. , Rice John D. , Preston James F.
Published: February 2006

Environmental and economic factors predicate the need for efficient processing of renewable sources of fuels and chemicals. To fulfill this need, microbial biocatalysts must be developed to efficiently process the hemicellulose fraction of lignocellulosic biomass for fermentation of pentoses. The predominance of methylglucuronoxylan (MeGAX
n
), a β-1,4 xylan in which 10% to 20% of the xylose residues are substituted with α-1,2-4-
O
-methylglucuronate residues, in hemicellulose fractions of hardwood and crop residues has made this a target for processing and fermentation. A
Paenibacillus
sp. (strain JDR-2) has been isolated and characterized for its ability to efficiently utilize MeGAX
n
. A modular xylanase (XynA
1
) of glycosyl hydrolase family 10 (GH 10) was identified through DNA sequence analysis that consists of a triplicate family 22 carbohydrate binding module followed by a GH 10 catalytic domain followed by a single family 9 carbohydrate binding module and concluding with C-terminal triplicate surface layer homology (SLH) domains. Immunodetection of the catalytic domain of XynA
1
(XynA
1
CD) indicates that the enzyme is associated with the cell wall fraction, supporting an anchoring role for the SLH modules. With MeGAX
n
as substrate, XynA
1
CD generated xylobiose and aldotetrauronate (MeGAX
3
) as predominant products. The inability to detect depolymerization products in medium during exponential growth of
Paenibacillus
sp. strain JDR-2 on MeGAX
n
, as well as decreased growth rate and yield with XynA
1
CD-generated xylooligosaccharides and aldouronates as substrates, indicates that XynA
1
catalyzes a depolymerization process coupled to product assimilation. This depolymerization/assimilation system may be utilized for development of biocatalysts to efficiently convert MeGAX
n
to alternative fuels and biobased products.
Use of PMA1 as a Housekeeping Biomarker for Assessment of Toxicant-Induced Stress in
Saccharomyces cerevisiae
Authors: Schmitt Marcel , Schwanewilm Petra , Ludwig Jost , Lichtenberg-Fraté Hella
Published: February 2006

The brewer's yeast
Saccharomyces cerevisiae
has emerged as a versatile and robust model system for laboratory use to study toxic effects of various substances. In this study, toxicant-induced stresses of pure compounds were investigated in
Saccharomyces cerevisiae
utilizing a destabilized version of the green fluorescent protein optimized for expression in yeast (yEGFP3) under control of the promoter of the housekeeping plasma membrane ATPase gene
PMA1
. The responses of the biomarker upon increasing test compound concentrations were monitored by determining the decrease in fluorescence. The reporter assay deployed a simple and robust protocol for the rapid detection of toxic effects within a 96-well microplate format. Fluorescence emissions were normalized to cell growth determined by absorption and were correlated to internal reference standards. The results were expressed as effective concentrations (EC
20
). Dose-response experiments were conducted in which yeast cells were exposed in minimal medium and in the presence of 20% fetal calf serum to sublethal concentrations of an array of heavy metals, salt, and a number of stress-inducing compounds (Diclofenac, Lindane, methyl-
N
-nitro-
N
-nitrosoguanidine [MNNG], hydroxyurea, and caffeine). Long-term exposure (7 h) played a considerable role in the adaptive response to intoxication compared to early responses at 4 h exposure. The data obtained after 4 h of exposure and expressed as EC
20
were compared to 50% inhibitory concentration values derived from cell line and ecotoxicological tests. This study demonstrates the versatility of the novel biomarker to complement existing test batteries to assess contaminant exposure and effects.
Comparative Genomics of DNA Fragments from Six Antarctic Marine Planktonic Bacteria
Authors: Grzymski Joseph J. , Carter Brandon J. , DeLong Edward F. , Feldman Robert A. , Ghadiri Amir , Murray Alison E.
Published: February 2006

Six
environmental fosmid clones from Antarctic coastal water
bacterioplankton were completely sequenced. The genome fragments
harbored small-subunit rRNA genes that were between 85 and 91% similar
to those of their nearest cultivated relatives. The six fragments span
four phyla, including the
Gemmatimonadetes
,
Proteobacteria
(α and γ),
Bacteroidetes
, and high-G+C gram-positive bacteria.
Gene-finding and annotation analyses identified 244 total open reading
frames. Amino acid comparisons of 123 and 113 Antarctic
bacterial amino acid sequences to mesophilic homologs from
G+C-specific and SwissProt/UniProt databases, respectively,
revealed widespread adaptation to the cold. The most significant
changes in these Antarctic bacterial protein sequences included a
reduction in salt-bridge-forming residues such as arginine, glutamic
acid, and aspartic acid, reduced proline contents, and a reduction in
stabilizing hydrophobic clusters. Stretches of disordered amino acids
were significantly longer in the Antarctic sequences than in the
mesophilic sequences. These characteristics were not specific to any
one phylum, COG role category, or G+C content and imply that
underlying genotypic and biochemical adaptations to the cold are
inherent to life in the permanently subzero Antarctic
waters.
Multiple-Locus Sequence Typing Analysis of
Bacillus cereus
and
Bacillus thuringiensis
Reveals Separate Clustering and a Distinct Population Structure of Psychrotrophic Strains
Authors: Sorokin Alexei , Candelon Benjamin , Guilloux Kévin , Galleron Nathalie , Wackerow-Kouzova Natalia , Ehrlich S. Dusko , Bourguet Denis , Sanchis Vincent
Published: February 2006

We
used multilocus sequence typing (MLST) to characterize phylogenetic
relationships for a collection of
Bacillus cereus
group
strains isolated from forest soil in the Paris area during a mild
winter. This collection contains multiple strains isolated from the
same soil sample and strains isolated from samples from different
sites. We characterized 115 strains of this collection and 19 other
strains based on the sequences of the
clpC
,
dinB
,
gdpD
,
panC
,
purF
, and
yhfL
loci.
The number of alleles ranged from 36 to 53, and a total of 93 allelic
profiles or sequence types were distinguished. We identified three
major strain clusters—C, T, and W—based on the
comparison of individual gene sequences or concatenated sequences. Some
less representative clusters and subclusters were also distinguished.
Analysis of the MLST data using the concept of clonal complexes led to
the identification of two, five, and three such groups in clusters C,
T, and W, respectively. Some of the forest isolates were closely
related to independently isolated psychrotrophic strains. Systematic
testing of the strains of this collection showed that almost all the
strains that were able to grow at a low temperature (6°C)
belonged to cluster W. Most of these strains, including three
independently isolated strains, belong to two clonal complexes and are
therefore very closely related genetically. These clonal complexes
represent strains corresponding to the previously identified species
Bacillus weihenstephanensis
. Most of the other strains of our
collection, including some from the W cluster, are not psychrotrophic.
B. weihenstephanensis
(cluster W) strains appear to comprise
an effectively sexual population, whereas
Bacillus
thuringiensis
(cluster T) and
B. cereus
(cluster C) have
clonal population
structures.
Multitiered Approach Using Quantitative PCR To Track Sources of Fecal Pollution Affecting Santa Monica Bay, California
Authors: Noble Rachel T. , Griffith John F. , Blackwood A. Denene , Fuhrman Jed A. , Gregory Jason B. , Hernandez Ximena , Liang Xiaolin , Bera Angie A. , Schiff Kenneth
Published: February 2006

The ubiquity of fecal indicator bacteria such as
Escherichia coli
and
Enterococcus
spp. in urban environments makes tracking of fecal contamination extremely challenging. A multitiered approach was used to assess sources of fecal pollution in Ballona Creek, an urban watershed that drains to the Santa Monica Bay (SMB) near Los Angeles, Calif. A mass-based design at six main-stem sites and four major tributaries over a 6-h period was used (i) to assess the flux of
Enterococcus
spp. and
E. coli
by using culture-based methods (tier 1); (ii) to assess levels of
Enterococcus
spp. by using quantitative PCR and to detect and/or quantify additional markers of human fecal contamination, including a human-specific
Bacteroides
sp. marker and enterovirus, using quantitative reverse transcriptase PCR (tier 2); and (iii) to assess the specific types of enterovirus genomes found via sequence analysis (tier 3). Sources of fecal indicator bacteria were ubiquitous, and concentrations were high, throughout Ballona Creek, with no single tributary dominating fecal inputs. The flux of
Enterococcus
spp. and
E. coli
averaged 10
9
to 10
10
cells h
−1
and was as high at the head of the watershed as at the mouth prior to discharge into the SMB. In addition, a signal for the human-specific
Bacteroides
marker was consistently detected: 86% of the samples taken over the extent during the study period tested positive. Enteroviruses were quantifiable in 14 of 36 samples (39%), with the highest concentrations at the site furthest upstream (Cochran). These results indicated the power of using multiple approaches to assess and quantify fecal contamination in freshwater conduits to high-use, high-priority recreational swimming areas.
Enhanced Benzaldehyde Tolerance in
Zymomonas mobilis
Biofilms and the Potential of Biofilm Applications in Fine-Chemical Production
Authors: Li Xuan Zhong , Webb Jeremy S. , Kjelleberg Staffan , Rosche Bettina
Published: February 2006

Biotransformation plays an increasingly important role in the industrial production of fine chemicals due to its high product specificity and low energy requirement. One challenge in biotransformation is the toxicity of substrates and/or products to biocatalytic microorganisms and enzymes. Biofilms are known for their enhanced tolerance of hostile environments compared to planktonic free-living cells.
Zymomonas mobilis
was used in this study as a model organism to examine the potential of surface-associated biofilms for biotransformation of chemicals into value-added products.
Z. mobilis
formed a biofilm with a complex three-dimensional architecture comprised of microcolonies with an average thickness of 20 μm, interspersed with water channels. Microscopic analysis and metabolic activity studies revealed that
Z. mobilis
biofilm cells were more tolerant to the toxic substrate benzaldehyde than planktonic cells were. When exposed to 50 mM benzaldehyde for 1 h, biofilm cells exhibited an average of 45% residual metabolic activity, while planktonic cells were completely inactivated. Three hours of exposure to 30 mM benzaldehyde resulted in sixfold-higher residual metabolic activity in biofilm cells than in planktonic cells. Cells inactivated by benzaldehyde were evenly distributed throughout the biofilm, indicating that the resistance mechanism was different from mass transfer limitation. We also found that enhanced tolerance to benzaldehyde was not due to the conversion of benzaldehyde into less toxic compounds. In the presence of glucose,
Z. mobilis
biofilms in continuous cultures transformed 10 mM benzaldehyde into benzyl alcohol at a steady rate of 8.11 g (g dry weight)
−1
day
−1
with a 90% molar yield over a 45-h production period.
GIP2, a Putative Transcription Factor That Regulates the Aurofusarin Biosynthetic Gene Cluster in
Gibberella zeae
Authors: Kim Jung-Eun , Jin Jianming , Kim Hun , Kim Jin-Cheol , Yun Sung-Hwan , Lee Yin-Won
Published: February 2006

Gibberella zeae
(anamorph:
Fusarium graminearum
) is an important pathogen of maize, wheat, and rice. Colonies of
G. zeae
produce yellow-to-tan mycelia with the white-to-carmine red margins. In this study, we focused on nine putative open reading frames (ORFs) closely linked to
PKS12
and
GIP1
, which are required for aurofusarin biosynthesis in
G. zeae
. Among them is an ORF designated
GIP2
(for Gi
bberella zeae p
igment gene
2
), which encodes a putative protein of 398 amino acids that carries a Zn(II)
2
Cys
6
binuclear cluster DNA-binding domain commonly found in transcription factors of yeasts and filamentous fungi. Targeted gene deletion and complementation analyses confirmed that
GIP2
is required for aurofusarin biosynthesis. Expression of
GIP2
in carrot medium correlated with aurofusarin production by
G. zeae
and was restricted to vegetative mycelia. Inactivation of the 10 contiguous genes in the Δ
GIP2
strain delineates an aurofusarin biosynthetic gene cluster. Overexpression of
GIP2
in both the Δ
GIP2
and the wild-type strains increases aurofusarin production and reduces mycelial growth. Thus,
GIP2
is a putative positive regulator of the aurofusarin biosynthetic gene cluster, and aurofusarin production is negatively correlated with vegetative growth by
G. zeae
.
Biodiversity of Methanogenic and Other
Archaea
in the Permanently Frozen Lake Fryxell, Antarctica
Authors: Karr Elizabeth A. , Ng Joshua M. , Belchik Sara M. , Sattley W. Matthew , Madigan Michael T. , Achenbach Laurie A.
Published: February 2006

Archaea
were detected in molecular diversity studies of the permanently frozen Lake Fryxell, Antarctica. Two clusters of methanogens were detected in the sediments, and another cluster of possibly methanotrophic
Euryarchaeota
was detected in the anoxic water column just above the sediments. One crenarchaeote was detected in water just below the oxycline. The
Archaea
present in Lake Fryxell are likely involved in the major biogeochemical cycles that occur there.
A
Bacillus megaterium
Plasmid System for the Production, Export, and One-Step Purification of Affinity-Tagged Heterologous Levansucrase from Growth Medium
Authors: Malten Marco , Biedendieck Rebekka , Gamer Martin , Drews Ann-Christin , Stammen Simon , Buchholz Klaus , Dijkhuizen Lubbert , Jahn Dieter
Published: February 2006

A multiple vector system for the production and export of recombinant affinity-tagged proteins in
Bacillus megaterium
was developed. Up to 1 mg/liter of a His
6
-tagged or Strep-tagged
Lactobacillus reuteri
levansucrase was directed into the growth medium, using the
B. megaterium
esterase LipA signal peptide, and recovered by one-step affinity chromatography.
Cumulative Effects of Short-Term Polymetal Contamination on Soil Bacterial Community Structure
Authors: Ranjard L. , Lignier L. , Chaussod R.
Published: February 2006

In this study we evaluated the short-term effects of copper, cadmium, and mercury, added singly or in combination at different doses, on soil bacterial community structure using the bacterial automated ribosomal intergenic spacer analysis (B-ARISA) fingerprinting technique. Principal-component analysis of B-ARISA profiles allowed us to deduce the following order of impact: (Cu + Cd + Hg) >> Hg ≥ Cd > Cu. These results demonstrated that there was a cumulative effect of metal toxicity. Furthermore, the trend of modifications was consistent with the “hump-backed” relationships between biological diversity and disturbance described by Giller et al. (K. E. Giller, E. Witler, and S. P. McGrath, Soil Biol. Biochem.
30:
1389-1414, 1998).
Probing In Vivo Promoter Activities in
Mycoplasma pneumoniae
: a System for Generation of Single-Copy Reporter Constructs
Authors: Halbedel Sven , Stülke Jörg
Published: February 2006

The nucleotide sequences that control transcription initiation and regulation in
Mycoplasma pneumoniae
are poorly understood. In this work, we developed a
lacZ
-based reporter plasmid that can be used to integrate fusions of promoter fragments to a promoterless
lacZ
gene into the chromosome of
M. pneumoniae
.
Diversity of Microorganisms within Rock Varnish in the Whipple Mountains, California
Authors: Kuhlman K. R. , Fusco W. G. , La Duc M. T. , Allenbach L. B. , Ball C. L. , Kuhlman G. M. , Anderson R. C. , Erickson I. K. , Stuecker T. , Benardini J. , Strap J. L. , Crawford R. L.
Published: February 2006

Rock varnish from Arizona's Whipple Mountains harbors a microbial community containing about 108microorganisms g−1of varnish. Analyses of varnish phospholipid fatty acids and rRNA gene libraries reveal a community comprised of mostlyProteobacteriabut also includingActinobacteria, eukaryota, and a few members of theArchaea. Rock varnish represents a significant niche for microbial colonization.
Characterization of an
Agrobacterium tumefaciens
d
-Psicose 3-Epimerase That Converts
d
-Fructose to
d
-Psicose
Authors: Kim Hye-Jung , Hyun Eun-Kyung , Kim Yeong-Su , Lee Yong-Joo , Oh Deok-Kun
Published: February 2006

The noncharacterized gene previously proposed as the
d
-tagatose 3-epimerase gene from
Agrobacterium tumefaciens
was cloned and expressed in
Escherichia coli
. The expressed enzyme was purified by three-step chromatography with a final specific activity of 8.89 U/mg. The molecular mass of the purified protein was estimated to be 132 kDa of four identical subunits. Mn
2+
significantly increased the epimerization rate from
d
-fructose to
d
-psicose. The enzyme exhibited maximal activity at 50°C and pH 8.0 with Mn
2+
. The turnover number (
k
cat
) and catalytic efficiency (
k
cat
/

K
m

) of the enzyme for
d
-psicose were markedly higher than those for
d
-tagatose, suggesting that the enzyme is not
d
-tagatose 3-epimerase but
d
-psicose 3-epimerase. The equilibrium ratio between
d
-psicose and
d
-fructose was 32:68 at 30°C.
d
-Psicose was produced at 230 g/liter from 700-g/liter
d
-fructose at 50°C after 100 min, corresponding to a conversion yield of 32.9%.
Identification of an
endo
-β-1,4-
d
-Xylanase from
Magnaporthe grisea
by Gene Knockout Analysis, Purification, and Heterologous Expression
Authors: Wu Sheng-Cheng , Halley Jeffrey E. , Luttig Christopher , Fernekes Linda M. , Gutiérrez-Sanchez Gerardo , Darvill Alan G. , Albersheim Peter
Published: February 2006

Magnaporthe grisea
, a destructive ascomycetous pathogen of rice, secretes cell wall-degrading enzymes into a culture medium containing purified rice cell walls as the sole carbon source. From
M. grisea
grown under the culture conditions described here, we have identified an expressed sequenced tag,
XYL-6
, a gene that is also expressed in
M. grisea
-infected rice leaves 24 h postinoculation with conidia. This gene encodes a protein about 65% similar to
endo
-β-1,4-
d
-glycanases within glycoside hydrolase family GH10. A
M. grisea
knockout mutant for
XYL-6
was created, and it was shown to be as virulent as the parent strain in infecting the rice host. The proteins secreted by the parent strain and by the
xyl-6
Δ mutant were each fractionated by liquid chromatography, and the collected fractions were assayed for
endo
-β-1,4-
d
-glucanase or
endo
-β-1,4-
d
-xylanase activities. Two protein-containing peaks with
endo
-β-1,4-
d
-xylanase activity secreted by the parent strain are not detectable in the column eluant of the proteins secreted by the mutant. The two endoxylanases (XYL-6α and XYL-6β) from the parent were each purified to homogeneity. N-terminal amino acid sequencing indicated that XYL-6α is a fragment of XYL-6β and that XYL-6β is identical to the deduced protein sequence encoded by the
XYL-6
gene. Finally,
XYL-6
was introduced into
Pichia pastoris
for heterologous expression, which resulted in the purification of a fusion protein, XYL-6H, from the
Pichia pastoris
culture filtrate. XYL-6H is active in cleaving arabinoxylan. These experiments unequivocally established that the
XYL-6
gene encodes a secreted
endo
-β-1,4-
d
-xylanase.
Reliable and Rapid Identification of
Listeria monocytogenes
and
Listeria
Species by Artificial Neural Network-Based Fourier Transform Infrared Spectroscopy
Authors: Rebuffo Cecilia A. , Schmitt Jürgen , Wenning Mareike , von Stetten Felix , Scherer Siegfried
Published: February 2006

Differentiation of the species within the genus
Listeria
is important for the food industry but only a few reliable methods are available so far. While a number of studies have used Fourier transform infrared (FTIR) spectroscopy to identify bacteria, the extraction of complex pattern information from the infrared spectra remains difficult. Here, we apply artificial neural network technology (ANN), which is an advanced multivariate data-processing method of pattern analysis, to identify
Listeria
infrared spectra at the species level. A hierarchical classification system based on ANN analysis for
Listeria
FTIR spectra was created, based on a comprehensive reference spectral database including 243 well-defined reference strains of
Listeria monocytogenes
,
L. innocua
,
L. ivanovii
,
L. seeligeri
, and
L. welshimeri
. In parallel, a univariate FTIR identification model was developed. To evaluate the potentials of these models, a set of 277 isolates of diverse geographical origins, but not included in the reference database, were assembled and used as an independent external validation for species discrimination. Univariate FTIR analysis allowed the correct identification of 85.2% of all strains and of 93% of the
L. monocytogenes
strains. ANN-based analysis enhanced differentiation success to 96% for all
Listeria
species, including a success rate of 99.2% for correct
L. monocytogenes
identification. The identity of the 277-strain test set was also determined with the standard phenotypical API
Listeria
system. This kit was able to identify 88% of the test isolates and 93% of
L. monocytogenes
strains. These results demonstrate the high reliability and strong potential of ANN-based FTIR spectrum analysis for identification of the five
Listeria
species under investigation. Starting from a pure culture, this technique allows the cost-efficient and rapid identification of
Listeria
species within 25 h and is suitable for use in a routine food microbiological laboratory.
Differential strength of association of child injury prevention attitudes and beliefs on practices: a case for audience segmentation
Authors: Vladutiu C J , Nansel T R , Weaver N L , Jacobsen H A , Kreuter M W
Published: February 2006

Objective:Many injuries to children cannot be prevented without some degree of active behavior on the part of parents. A better understanding of social and cognitive determinants of parents’ injury prevention behavior and the identification of potential subgroups for targeted message delivery could advance the effectiveness of educational and behavioral interventions. This study assessed the degree to which parents’ injury prevention behavior is associated with theoretical determinants and examined whether this relation differs by age or birth order of child.

Design:Cross sectional observational study.

Setting:Three Midwestern pediatric clinics.

Subjects:594 parents of children ages 0–4 attending routine well child visits.

Measures:Injury prevention attitudes, beliefs, and practices.

Results:Overall, only modest relations were observed between injury beliefs and attitudes and injury prevention behaviors. However, these relations differed substantially by child age and birth order, with stronger associations observed for parents of older first born children. Outcome expectations and social norms were more strongly related to injury prevention behavior among parents of preschool children than among parents of infants and toddlers, while attitudes were more predictive for parents of first born children than parents of later born children.

Conclusions:These findings highlight the complexity of relations between theorized determinants and behavior, and suggest the potential utility of using audience segmentation strategies in behavioral interventions addressing injury prevention.
Accuracy of injury coding under ICD-9 for New Zealand public hospital discharges
Authors: Langley J , Stephenson S , Thorpe C , Davie G
Published: February 2006

Objective:To determine the level of accuracy in coding for injury principal diagnosis and the first external cause code for public hospital discharges in New Zealand and determine how these levels vary by hospital size.

Method:A simple random sample of 1800 discharges was selected from the period 1996–98 inclusive. Records were obtained from hospitals and an accredited coder coded the discharge independently of the codes already recorded in the national database.

Results:Five percent of the principal diagnoses, 18% of the first four digits of the E-codes, and 8% of the location codes (5th digit of the E-code), were incorrect. There were no substantive differences in the level of incorrect coding between large and small hospitals.

Conclusions:Users of New Zealand public hospital discharge data can have a high degree of confidence in the injury diagnoses coded under ICD-9-CM-A. A similar degree of confidence is warranted for E-coding at the group level (for example, fall), but not, in general, at higher levels of specificity (for example, type of fall). For those countries continuing to use ICD-9 the study provides insight into potential problems of coding and thus guidance on where the focus of coder training should be placed. For those countries that have historical data coded according to ICD-9 it suggests that some specific injury and external cause incidence estimates may need to be treated with more caution.
Specific epitopes of the structural and hypothetical proteins elicit variable humoral responses in SARS patients
Authors: Chow S C S , Ho C Y S , Tam T T Y , Wu C , Cheung T , Chan P K S , Ng M H L , Hui P K , Ng H K , Au D M Y , Lo A W I
Published: February 2006

Background:Severe acute respiratory syndrome (SARS) is an infectious disease which was caused by a novel coronavirus (SARS-CoV). SARS has caused an outbreak in the world during 2003 and 2004, with 8098 individuals being infected and a death toll of 774 in 28 regions around the world. Specific humoral responses to viral infection remain unclear.

Objective:To analyse the antigenicity of the SARS-CoV genome and identify potential antigenic epitopes in the structural proteins.

Methods:Potential antigenic epitopes were identified in the structural proteins (nucleocapsid, membrane, spike, and small envelope proteins) and hypothetical proteins (SARS3a, 3b, 6, 7a, and 9b) that are specific for SARS-CoV. A peptide chip platform was created and the profiles of antibodies to these epitopes were investigated in 59 different SARS patients’ sera obtained 6–103 days after the onset of the illness. Serial sera from five additional patients were also studied.

Results:Epitopes at the N-terminus of the membrane protein and the C-terminus of nucleocapsid protein elicited strong antibody responses. Epitopes on the spike protein were only moderately immunogenic but the effects were persistent. Antibodies were also detected for some putative proteins, noticeably the C-termini of SARS3a and SARS6.

Conclusions:Important epitopes of the SARS-CoV genome that may serve as potential markers for the viral infection are identified. These specific antigenic sites may also be important for vaccine development against this new fatal infectious disease.
Modern management of arrhythmias
Authors: Hall M C S , Todd D M
Published: February 2006

The specialist management of arrhythmias has changed significantly over the past decade. This article outlines current management strategies for atrial flutter and atrial fibrillation, with particular emphasis on curative strategies with catheter ablation and the recent data on rhythm compared with rate control strategies. The expanding role of catheter ablation in the treatment of a wide variety of supraventricular and ventricular arrhythmias is discussed. The current evidence for implantable cardioverter defibrillators in the prevention of sudden cardiac death is summarised. The article also highlights the increasing recognition of a number of inherited syndromes that predispose to sudden cardiac death (for example, Brugada and long QT syndromes).
Incidence and risk factors for development of venous thromboembolism in Indian patients undergoing major orthopaedic surgery: results of a prospective study
Authors: Bagaria V , Modi N , Panghate A , Vaidya S
Published: February 2006

Introduction

The incidence of venous thromboembolism (VTE) in Western populations undergoing major orthopaedic surgery without any thromboprophylaxis has been reported to range from 32% to 88%. There is however limited information on incidence of VTE in Indian patients and most of the Indian patients undergoing these surgeries do not receive any form of prophylaxis regardless of their risk profile.

Methods

A prospective study was performed on 147 patients undergoing major orthopaedic surgery for total knee replacement (TKR), total hip replacement (THR), and proximal femur fracture fixation (PFF) without any prophylaxis. These patients were profiled for presence of the known risk factors responsible for development of VTE. A duplex ultrasound on both lower limbs was done 6 to 10 days after surgery. Twenty three patients underwent THR, 22 patients underwent TKR, and 102 underwent surgery for PFF. The patients were assessed clinically for any signs of deep venous thrombosis (DVT) and pulmonary embolism (PE). A helical CT scan was done in case of suspicion of PE and a duplex ultrasound was done in case of clinical suspicion of DVT irrespective of the stage of study.

Results

The overall incidence of VTE was 6.12% and that of PE was 0.6%. The risk factors that were found to be significantly responsible for development of VTE (p < 0.05) were: immobility greater than 72 hours, malignancy, obesity, surgery lasting more than two hours.

Conclusion

The study reconfirms the belief that DVT has a lower incidence in Indian patients as compared with other ethnic groups.
Diabetic neuropathy
Authors: Bansal V , Kalita J , Misra U K
Published: February 2006

Diabetic neuropathy (DN) refers to symptoms and signs of neuropathy in a patient with diabetes in whom other causes of neuropathy have been excluded. Distal symmetrical neuropathy is the commonest accounting for 75% DN. Asymmetrical neuropathies may involve cranial nerves, thoracic or limb nerves; are of acute onset resulting from ischaemic infarction of vasa nervosa. Asymmetric neuropathies in diabetic patients should be investigated for entrapment neuropathy. Diabetic amyotrophy, initially considered to result from metabolic changes, and later ischaemia, is now attributed to immunological changes. For diagnosis of DN, symptoms, signs, quantitative sensory testing, nerve conduction study, and autonomic testing are used; and two of these five are recommended for clinical diagnosis. Management of DN includes control of hyperglycaemia, other cardiovascular risk factors; α lipoic acid and L carnitine. For neuropathic pain, analgesics, non-steroidal anti-inflammatory drugs, antidepressants, and anticonvulsants are recommended. The treatment of autonomic neuropathy is symptomatic.
Retrospective analysis of the use of inferior vena cava filters in routine hospital practice
Authors: Dovrish Z , Hadary R , Blickstein D , Shilo L , Ellis M H
Published: February 2006

Introduction

Characteristics and outcomes of patients undergoing inferior vena cava (IVC) filter insertion are not well reported. Particularly, the role of long term anticoagulation in these patients is unclear.

Aims

(1) To describe in a cohort of patients undergoing IVC filter insertion, underlying diseases, indications for filter insertion, complications, and survival. (2) To determine the effect of long term anticoagulant treatment on thromboembolism and patient survival.

Study design

A retrospective analysis of 109 consecutive patients undergoing IVC filter insertion in two university hospitals.

Results

Average age was 67.4 years. Median duration of follow up was two years. Indications for IVC filter insertion were: contraindication to anticoagulation (n = 61, 56%), prophylactic insertion (n = 29, 27%), thromboembolism while receiving adequate anticoagulation (n = 17, 15%), and non-compliance with anticoagulation (n = 2, 2%). Insertion related complications were groin haematoma in four patients (3.5%) and localised infection at the puncture site in one patient (0.9%). Fifty six patients (51.4%) died during the study period. Of these, 22 received long term anticoagulants and 34 did not. Overall and thrombosis free survival was greater in the anticoagulant treated group (median survival not reached) than in the untreated group (median survival = 12 months). Patients not receiving long term anticoagulation after IVC filter insertion were nearly 2.5-fold more likely to die or experience venous thromboembolism.

Conclusion

IVC filter insertion was a safe procedure and was performed for appropriate indications in the patients studied. In patients surviving for longer than 30 days, prolonged administration of oral anticoagulants was associated with improved survival with no significant increase in haemorrhagic complications.
Effects of topical treatment of sodium butyrate and 5-aminosalicylic acid on expression of trefoil factor 3, interleukin 1β, and nuclear factor κB in trinitrobenzene sulphonic acid induced colitis in rats
Authors: Song M , Xia B , Li J
Published: February 2006

Background and Aims

Butyrate enemas have been shown to be effective in treatment of ulcerative colitis, but the mechanism of the effects of butyrate is not totally known. This study evaluates effects of topical treatment of sodium butyrate (NaB) and 5-aminosalicylic acid (5-ASA) on the expression of trefoil factor 3 (TFF3), interleukin 1β (IL1β), and nuclear factor κB (NFκB) in trinitrobenzene sulphonic acid (TNBS) induced colitis in rats.

Methods

Distal colitis was induced in male Wistar rats by colonic administration of TNBS and colonically treated with NaB, 5-ASA, combination of NaB and 5-ASA, and normal saline for 14 consecutive days. Colonic damage score, tissue myeloperoxidase (MPO) activity, TFF3 mRNA expression, serum IL1β production, and tissue NFκB expression were determined, respectively.

Results

Treatment of NaB, 5-ASA, and the combination improved diarrhoea, colonic damage score, and MPO activities, increased TFF3 mRNA expression, and decreased serum IL1β production and tissue NFκB expression. The combination therapy of NaB and 5-ASA had better effects than any other single treatment.

Conclusions

The combination of topical treatment of NaB and 5-ASA was effective for relieving and repairing colonic inflammation and the effects were related to stimulation of TFF3 mRNA expression and down-regulation of IL1β production and NFκB expression.