bluuebunny/crossref_metadata_2025_split huggingface dataset:
Quantitative Fluorescence
In Situ
Hybridization of Microbial Communities in the Rumens of Cattle Fed Different Diets
Authors: Kong Yunhong , He Maolong , McAlister Tim , Seviour Robert , Forster Robert 
Published: August 2010

ABSTRACT

At present there is little quantitative information on the identity and composition of bacterial populations in the rumen microbial community. Quantitative fluorescence
in situ
hybridization using newly designed oligonucleotide probes was applied to identify the microbial populations in liquid and solid fractions of rumen digesta from cows fed barley silage or grass hay diets with or without flaxseed.
Bacteroidetes
,
Firmicutes
, and
Proteobacteria
were abundant in both fractions, constituting 31.8 to 87.3% of the total cell numbers. They belong mainly to the order
Bacteroidales
(0.1 to 19.2%), hybridizing with probe BAC1080; the families
Lachnospiraceae
(9.3 to 25.5%) and
Ruminococcaceae
(5.5 to 23.8%), hybridizing with LAC435 and RUM831, respectively; and the classes
Deltaproteobacteria
(5.8 to 28.3%) and
Gammaproteobacteria
(1.2 to 8.2%). All were more abundant in the rumen communities of cows fed diets containing silage (75.2 to 87.3%) than in those of cows fed diets containing hay (31.8 to 49.5%). The addition of flaxseed reduced their abundance in the rumens of cows fed silage-based diets (to 45.2 to 58.7%) but did not change markedly their abundance in the rumens of cows fed hay-based diets (31.8 to 49.5%). Fibrolytic species, including
Fibrobacter succinogenes
and
Ruminococcus
spp., and archaeal methanogens accounted for only a small proportion (0.4 to 2.1% and 0.2 to 0.6%, respectively) of total cell numbers. Depending on diet, between 37.0 and 91.6% of microbial cells specifically hybridized with the probes used in this study, allowing them to be identified
in situ
. The identities of other microbial populations (8.4 to 63.0%) remain unknown.
Isolation and Characterization of 4-
tert
-Butylphenol-Utilizing
Sphingobium fuliginis
Strains from
Phragmites australis
Rhizosphere Sediment
Authors: Toyama Tadashi , Momotani Naonori , Ogata Yuka , Miyamori Yuji , Inoue Daisuke , Sei Kazunari , Mori Kazuhiro , Kikuchi Shintaro , Ike Michihiko 
Published: August 2010

ABSTRACT

We isolated three
Sphingobium fuliginis
strains from
Phragmites australis
rhizosphere sediment that were capable of utilizing 4-
tert
-butylphenol as a sole carbon and energy source. These strains are the first 4-
tert
-butylphenol-utilizing bacteria. The strain designated TIK-1 completely degraded 1.0 mM 4-
tert
-butylphenol in basal salts medium within 12 h, with concomitant cell growth. We identified 4-
tert
-butylcatechol and 3,3-dimethyl-2-butanone as internal metabolites by gas chromatography-mass spectrometry. When 3-fluorocatechol was used as an inactivator of
meta
-cleavage enzymes, strain TIK-1 could not degrade 4-
tert
-butylcatechol and 3,3-dimethyl-2-butanone was not detected. We concluded that metabolism of 4-
tert
-butylphenol by strain TIK-1 is initiated by hydroxylation to 4-
tert
-butylcatechol, followed by a
meta
-cleavage pathway. Growth experiments with 20 other alkylphenols showed that 4-isopropylphenol, 4-
sec
-butylphenol, and 4-
tert
-pentylphenol, which have alkyl side chains of three to five carbon atoms with α-quaternary or α-tertiary carbons, supported cell growth but that 4-
n
-alkylphenols, 4-
tert
-octylphenol, technical nonylphenol, 2-alkylphenols, and 3-alkylphenols did not. The rate of growth on 4-
tert
-butylphenol was much higher than that of growth on the other alkylphenols. Degradation experiments with various alkylphenols showed that strain TIK-1 cells grown on 4-
tert
-butylphenol could degrade 4-alkylphenols with variously sized and branched side chains (ethyl,
n
-propyl, isopropyl,
n
-butyl,
sec
-butyl,
tert
-butyl,
n
-pentyl,
tert
-pentyl,
n
-hexyl,
n
-heptyl,
n
-octyl,
tert
-octyl,
n
-nonyl, and branched nonyl) via a
meta
-cleavage pathway but not 2- or 3-alkylphenols. Along with the degradation of these alkylphenols, we detected methyl alkyl ketones that retained the structure of the original alkyl side chains. Strain TIK-1 may be useful in the bioremediation of environments polluted by 4-
tert
-butylphenol and various other 4-alkylphenols.
Sensitization of Microcin J25-Resistant Strains by a Membrane-Permeabilizing Peptide
Authors: Pomares María Fernanda , Delgado Mónica A. , Corbalán Natalia S. , Farías Ricardo N. , Vincent Paula A. 
Published: August 2010

ABSTRACTMicrocin J25 (MccJ25) is a plasmid-encoded, 21-amino-acid, antibacterial peptide produced byEscherichia coli. MccJ25 inhibits RNA polymerase and the membrane respiratory chain. MccJ25 uptake intoE. coli-sensitive strains is mediated by the outer membrane receptor FhuA and the inner membrane proteins TonB, ExbB, ExbD, and SbmA. This peptide is active on someE. coli,Salmonella, andShigellaspecies strains, while other Gram-negative bacteria, such as clinical isolates ofEnterobacter cloacae,Citrobacter freundii,Klebsiella pneumoniae,Pseudomonas aeruginosa,Acinetobacter baumannii,Moraxella catarrhalis, andSalmonella entericaserovar Typhimurium, are completely resistant. In the present work, we demonstrated that the membrane-permeabilizing peptide (KFF)3K made some resistant strains sensitive to MccJ25, among themS.Typhimurium, where the antibiotic inhibitsin vitrocell growth and bacterial replication within macrophages. The results demonstrate that the membrane permeabilization induced by (KFF)3K allows MccJ25 penetration in an FhuA and SbmA-independent manner and suggest that the combination of both peptides could be considered as a therapeutic agent against pathogenicSalmonellastrains.
Use of Purified
Clostridium difficile
Spores To Facilitate Evaluation of Health Care Disinfection Regimens
Authors: Lawley Trevor D. , Clare Simon , Deakin Laura J. , Goulding David , Yen Jennifer L. , Raisen Claire , Brandt Cordelia , Lovell Jon , Cooke Fiona , Clark Taane G. , Dougan Gordon 
Published: August 2010

ABSTRACT

Clostridium difficile
is a major cause of antibiotic-associated diarrheal disease in many parts of the world. In recent years, distinct genetic variants of
C. difficile
that cause severe disease and persist within health care settings have emerged. Highly resistant and infectious
C. difficile
spores are proposed to be the main vectors of environmental persistence and host transmission, so methods to accurately monitor spores and their inactivation are urgently needed. Here we describe simple quantitative methods, based on purified
C. difficile
spores and a murine transmission model, for evaluating health care disinfection regimens. We demonstrate that disinfectants that contain strong oxidizing active ingredients, such as hydrogen peroxide, are very effective in inactivating pure spores and blocking spore-mediated transmission. Complete inactivation of 10
6
pure
C. difficile
spores on indicator strips, a six-log reduction, and a standard measure of stringent disinfection regimens require at least 5 min of exposure to hydrogen peroxide vapor (HPV; 400 ppm). In contrast, a 1-min treatment with HPV was required to disinfect an environment that was heavily contaminated with
C. difficile
spores (17 to 29 spores/cm
2
) and block host transmission. Thus, pure
C. difficile
spores facilitate practical methods for evaluating the efficacy of
C. difficile
spore disinfection regimens and bringing scientific acumen to
C. difficile
infection control.
Characterization of
Lactococcus lactis
Phage 949 and Comparison with Other Lactococcal Phages
Authors: Samson Julie E. , Moineau Sylvain 
Published: August 2010

ABSTRACT

The virulent
Lactococcus lactis
phage 949 was isolated in 1975 from cheese whey in New Zealand. This phage is a member of the
Siphoviridae
family and of a rare lactococcal phage group that bears its name (949 group). It has an icosahedral capsid (79-nm diameter) and a very long noncontractile tail (length, 500 nm; width, 12 nm). It infected 7 of 59 tested
L. lactis
strains, a somewhat expanded host range for a rare lactococcal phage. The abortive phage infection defense mechanisms AbiQ and AbiT strongly inhibited the multiplication of phage 949, but AbiK and AbiV did not. Its double-stranded DNA (dsDNA) genome of 114,768 bp is, to date, the largest among lactococcal phages. Its GC content was calculated at 32.7%, which is the lowest reported for a lactococcal phage. Its 154 open reading frames (ORFs) share limited identity with database sequences. In addition, terminal redundancy was observed as well as the presence of six tRNAs, one group I intron, and putative recombinases. SDS-PAGE coupled with mass spectrometry identified 13 structural proteins. The genomes of the members of the 10 currently known
L. lactis
phage groups were used to construct a proteomic tree. Each
L. lactis
phage group separated into distinct genetic clusters, validating the current classification scheme. Of note, members of the polythetic P335 groups were clearly separated into subgroups.
Acquisition of Iron by Alkaliphilic
Bacillus
Species
Authors: McMillan Duncan G. G. , Velasquez Imelda , Nunn Brook L. , Goodlett David R. , Hunter Keith A. , Lamont Iain , Sander Sylvia G. , Cook Gregory M. 
Published: August 2010

ABSTRACT

The biochemical and molecular mechanisms used by alkaliphilic bacteria to acquire iron are unknown. We demonstrate that alkaliphilic (pH > 9)
Bacillus
species are sensitive to artificial iron (Fe
3+
) chelators and produce iron-chelating molecules. These alkaliphilic siderophores contain catechol and hydroxamate moieties, and their synthesis is stimulated by manganese(II) salts and suppressed by FeCl
3
addition. Purification and mass spectrometric characterization of the siderophore produced by
Caldalkalibacillus thermarum
failed to identify any matches to previously observed fragmentation spectra of known siderophores, suggesting a novel structure.
Effect of Estuarine Wetland Degradation on Transport of
Toxoplasma gondii
Surrogates from Land to Sea
Authors: Shapiro Karen , Conrad Patricia A. , Mazet Jonna A. K. , Wallender Wesley W. , Miller Woutrina A. , Largier John L. 
Published: August 2010

ABSTRACT

The flux of terrestrially derived pathogens to coastal waters presents a significant health risk to marine wildlife, as well as to humans who utilize the nearshore for recreation and seafood harvest. Anthropogenic changes in natural habitats may result in increased transmission of zoonotic pathogens to coastal waters. The objective of our work was to evaluate how human-caused alterations of coastal landscapes in California affect the transport of
Toxoplasma gondii
to estuarine waters.
Toxoplasma gondii
is a protozoan parasite that is excreted in the feces of infected felids and is thought to reach coastal waters in contaminated runoff. This zoonotic pathogen causes waterborne toxoplasmosis in humans and is a significant cause of death in threatened California sea otters. Surrogate particles that mimic the behavior of
T. gondii
oocysts in water were released in transport studies to evaluate if the loss of estuarine wetlands is contributing to an increased flux of oocysts into coastal waters. Compared to vegetated sites, more surrogates were recovered from unvegetated mudflat habitats, which represent degraded wetlands. Specifically, in Elkhorn Slough, where a large proportion of otters are infected with
T. gondii
, erosion of 36% of vegetated wetlands to mudflats may increase the flux of oocysts by more than 2 orders of magnitude. Total degradation of wetlands may result in increased
Toxoplasma
transport of 6 orders of magnitude or more. Destruction of wetland habitats along central coastal California may thus facilitate pathogen pollution in coastal waters with detrimental health impacts to wildlife and humans.
Proteolytic Enzyme Production by Strains of the Insect Pathogen
Xenorhabdus
and Characterization of an Early-Log-Phase-Secreted Protease as a Potential Virulence Factor
Authors: Massaoud Mustafa K. , Marokházi Judit , Fodor András , Venekei István 
Published: August 2010

ABSTRACT

As a comparison to a similar study on
Photorhabdus
strains, 15
Xenorhabdus
bacterial strains and secondary phenotypic variants of two strains were screened for proteolytic activity by five detection methods. Although the number and intensity of proteolytic activities were different, every strain was positive for proteolytic activity by several tests. Zymography following native PAGE detected two groups of activities with different substrate affinities and a higher and lower electrophoretic mobility that were distinguished as activity 1 and 2, respectively. Zymography following SDS-PAGE resolved three activities, which were provisionally named proteases A, B, and C. Only protease B, an ∼55-kDa enzyme, was produced by every strain. This enzyme exhibited higher affinity to the gelatin substrate than to the casein substrate. Of the chromogenic substrates used, three were hydrolyzed: furylacryloyl-Ala-Leu-Val-Tyr (Fua-ALVY), Fua-LGPA (LGPA is Leu-Gly-Pro-Ala) (a substrate for collagen peptidases), and succinyl-Ala-Ala-Pro-Phe-thiobenzyl (Succ-AAPF-SBzl). All but the Fua-LGPA-ase activity seemed to be from secreted enzymes. According to their substrate preference profiles and inhibitor sensitivities, at least six such proteolytic enzymes could be distinguished in the culture medium of
Xenorhabdus
strains. The proteolytic enzyme that was secreted the earliest, protease B and the Succ-AAPF-SBzl-hydrolyzing enzyme, appeared from the early logarithmic phase of growth. Protease B could also be detected in the hemolymph of
Xenorhabdus
-infected
Galleria mellonella
larvae from 15 h postinfection. The purified protease B hydrolyzed
in vitro
seven proteins in the hemolymph of
Manduca sexta
that were also cleaved by PrtA peptidase from
Photorhabdus
. The N-terminal sequence of protease B showed similarity to a 55-kDa serralysin type metalloprotease in
Xenorhabdus nematophila
, which had been identified as an orthologue of
Photorhabdus
PrtA peptidase.
Vertical Distribution of Ammonia-Oxidizing Crenarchaeota and Methanogens in the Epipelagic Waters of Lake Kivu (Rwanda-Democratic Republic of the Congo)
Authors: Llirós Marc , Gich Frederic , Plasencia Anna , Auguet Jean-Christophe , Darchambeau François , Casamayor Emilio O. , Descy Jean-Pierre , Borrego Carles 
Published: August 2010

ABSTRACT

Four stratified basins in Lake Kivu (Rwanda-Democratic Republic of the Congo) were sampled in March 2007 to investigate the abundance, distribution, and potential biogeochemical role of planktonic archaea. We used fluorescence
in situ
hybridization with catalyzed-reported deposition microscopic counts (CARD-FISH), denaturing gradient gel electrophoresis (DGGE) fingerprinting, and quantitative PCR (qPCR) of signature genes for ammonia-oxidizing archaea (16S rRNA for marine
Crenarchaeota
group 1.1a [MCG1] and ammonia monooxygenase subunit A [
amoA
]). Abundance of archaea ranged from 1 to 4.5% of total DAPI (4′,6-diamidino-2-phenylindole) counts with maximal concentrations at the oxic-anoxic transition zone (∼50-m depth). Phylogenetic analysis of the archaeal planktonic community revealed a higher level of richness of crenarchaeal 16S rRNA gene sequences (21 of the 28 operational taxonomic units [OTUs] identified [75%]) over euryarchaeotal ones (7 OTUs). Sequences affiliated with the kingdom
Euryarchaeota
were mainly recovered from the anoxic water compartment and mostly grouped into methanogenic lineages (
Methanosarcinales
and
Methanocellales
). In turn, crenarchaeal phylotypes were recovered throughout the sampled epipelagic waters (0- to 100-m depth), with clear phylogenetic segregation along the transition from oxic to anoxic water masses. Thus, whereas in the anoxic hypolimnion crenarchaeotal OTUs were mainly assigned to the miscellaneous crenarchaeotic group, the OTUs from the oxic-anoxic transition and above belonged to
Crenarchaeota
groups 1.1a and 1.1b, two lineages containing most of the ammonia-oxidizing representatives known so far. The concomitant vertical distribution of both nitrite and nitrate maxima and the copy numbers of both MCG1 16S rRNA and
amoA
genes suggest the potential implication of
Crenarchaeota
in nitrification processes occurring in the epilimnetic waters of the lake.
Extracts of Edible and Medicinal Plants Damage Membranes of
Vibrio cholerae
Authors: Sánchez Eduardo , García Santos , Heredia Norma 
Published: August 2010

ABSTRACT

The use of natural compounds from plants can provide an alternative approach against food-borne pathogens. The mechanisms of action of most plant extracts with antimicrobial activity have been poorly studied. In this work, changes in membrane integrity, membrane potential, internal pH (pH
in
), and ATP synthesis were measured in
V
ibrio
cholerae
cells after exposure to extracts of edible and medicinal plants. A preliminary screen of methanolic, ethanolic, and aqueous extracts of medicinal and edible plants was performed. Minimal bactericidal concentrations (MBCs) were measured for extracts showing high antimicrobial activity. Our results indicate that methanolic extracts of basil (
Ocimum basilicum
L.), nopal cactus (
Opuntia ficus-indica
var. Villanueva L.), sweet acacia (
Acacia farnesiana
L.), and white sagebrush (
Artemisia ludoviciana
Nutt.) are the most active against
V. cholera
, with MBCs ranging from 0.5 to 3.0 mg/ml. Using four fluorogenic techniques, we studied the membrane integrity of
V. cholerae
cells after exposure to these four extracts. Extracts from these plants were able to disrupt the cell membranes of
V. cholerae
cells, causing increased membrane permeability, a clear decrease in cytoplasmic pH, cell membrane hyperpolarization, and a decrease in cellular ATP concentration in all strains tested. These four plant extracts could be studied as future alternatives to control
V. cholerae
contamination in foods and the diseases associated with this microorganism.
A Novel Gene,ROA, Is Required for Normal Morphogenesis and Discharge of Ascospores in Gibberella zeae
Authors: Min Kyunghun , Lee Jungkwan , Kim Jin-Cheol , Kim Sang Gyu , Kim Young Ho , Vogel Steven , Trail Frances , Lee Yin-Won 
Published: August 2010

ABSTRACTHead blight, caused byGibberella zeae, is a significant disease among cereal crops, including wheat, barley, and rice, due to contamination of grain with mycotoxins.G. zeaeis spread by ascospores forcibly discharged from sexual fruiting bodies forming on crop residues. In this study, we characterized a novel gene,ROA, which is required for normal sexual development. Deletion ofROA(Δroa) resulted in an abnormal size and shape of asci and ascospores but did not affect vegetative growth. The Δroamutation triggered round ascospores and insufficient cell division after spore delimitation. The asci of the Δroastrain discharged fewer ascospores from the perithecia but achieved a greater dispersal distance than those of the wild-type strain. Turgor pressure within the asci was calculated through the analysis of osmolytes in the epiplasmic fluid. Deletion of theROAgene appeared to increase turgor pressure in the mutant asci. The higher turgor pressure of the Δroamutant asci and the mutant spore shape contributed to the longer distance dispersal. When the Δroamutant was outcrossed with a Δmat1-2mutant, a strain that contains a green fluorescence protein (GFP) marker in place of theMAT1-2gene, unusual phenotypic segregation occurred. The ratio of GFP to non-GFP segregation was 1:1; however, all eight spores had the same shape. Taken together, the results of this study suggest thatROAplays multiple roles in maintaining the proper morphology and discharge of ascospores inG. zeae.
Comparative Analysis of the Biochemical and Functional Properties of C-Terminal Domains of Autotransporters
Authors: Marín Elvira , Bodelón Gustavo , Fernández Luis Ángel 
Published: August 2010

ABSTRACTAutotransporters (ATs) are the largest group of proteins secreted by Gram-negative bacteria and include many virulence factors from human pathogens. ATs are synthesized as large precursors with a C-terminal domain that is inserted in the outer membrane (OM) and is essential for the translocation of an N-terminal passenger domain to the extracellular milieu. Several mechanisms have been proposed for AT secretion. Self-translocation models suggest transport across a hydrophilic channel formed by an internal pore of the β-barrel or by the oligomerization of C-terminal domains. Alternatively, an assisted-translocation model suggests that transport employs a conserved machinery of the bacterial OM such as the Bam complex. In this work we have investigated AT secretion by carrying out a comparative study to analyze the conserved biochemical and functional features of different C-terminal domains selected from ATs of gammaproteobacteria, betaproteobacteria, alphaproteobacteria, and epsilonproteobacteria. Our results indicate that C-terminal domains having an N-terminal α-helix and a β-barrel constitute functional transport units for the translocation of peptides and immunoglobulin domains with disulfide bonds.In vivoandin vitroanalyses show that multimerization is not a conserved feature in AT C-terminal domains. Furthermore, we demonstrate that the deletion of the conserved α-helix severely impairs β-barrel folding and OM insertion and thereby blocks passenger domain secretion. These observations suggest that the AT β-barrel without its α-helix cannot form a stable hydrophilic channel in the OM for protein translocation. The implications of our data for an understanding of AT secretion are discussed.
Mechanism of Positive Regulation by DsrA and RprA Small Noncoding RNAs: Pairing Increases Translation and ProtectsrpoSmRNA from Degradation
Authors: McCullen Colleen A. , Benhammou Jihane N. , Majdalani Nadim , Gottesman Susan 
Published: August 2010

ABSTRACTSmall noncoding RNAs (sRNAs) regulate gene expression inEscherichia coliby base pairing with mRNAs and modulating translation and mRNA stability. The sRNAs DsrA and RprA stimulate the translation of the stress response transcription factor RpoS by base pairing with the 5′ untranslated region of therpoSmRNA. In the present study, we found that therpoSmRNA was unstable in the absence of DsrA and RprA and that expression of these sRNAs increased both the accumulation and the half-life of therpoSmRNA. Mutations indsrA,rprA, orrpoSthat disrupt the predicted pairing sequences and reduce translation of RpoS also destabilize therpoSmRNA. We found that therpoSmRNA accumulates in an RNase E mutant strain in the absence of sRNA expression and, therefore, is degraded by an RNase E-mediated mechanism. DsrA expression is required, however, for maximal translation even whenrpoSmRNA is abundant. This suggests that DsrA protectsrpoSmRNA from degradation by RNase E and that DsrA has a further activity in stimulating RpoS protein synthesis.rpoSmRNA is subject to degradation by an additional pathway, mediated by RNase III, which, in contrast to the RNase E-mediated pathway, occurs in the presence and absence of DsrA or RprA.rpoSmRNA and RpoS protein levels are increased in an RNase III mutant strain with or without the sRNAs, suggesting that the role of RNase III in this context is to reduce the translation of RpoS even when the sRNAs are acting to stimulate translation.