bluuebunny/crossref_metadata_2025_split huggingface dataset:
Cost-Effective Application of Pulsed-Field Gel Electrophoresis to Typing of
Salmonella enterica
Serovar Typhimurium
Authors: Doran Geraldine , Morris Dearbhaile , O'Hare Colette , DeLappe Niall , Bradshaw Bernard , Corbett-Feeney Geraldine , Cormican Martin 
Published: December 2005

ABSTRACT

Salmonella enterica
serovar Typhimurium is frequently isolated from humans and animals. Phage typing is historically the first-line reference typing technique in Europe. It is rapid and convenient for laboratories with appropriate training and experience, and costs of consumables are low. Phage typing and pulsed-field gel electrophoresis (PFGE) were performed on 503 isolates of serovar Typhimurium. Twenty-nine phage types and 53 PFGE patterns were observed. Most isolates of phage types DT104, DT104b, and U310 are not distinguishable from other members of their phage type by PFGE. By contrast, PFGE of isolates of phage types DT193 and U302 shows great heterogeneity. Analysis of experience with PFGE and phage typing can facilitate the selective application of PFGE to maximize the yield of epidemiologically relevant additional information while controlling costs.
Multiple Reductive-Dehalogenase-Homologous Genes Are Simultaneously Transcribed during Dechlorination by
Dehalococcoides-
Containing Cultures
Authors: Waller Alison S. , Krajmalnik-Brown Rosa , Löffler Frank E. , Edwards Elizabeth A. 
Published: December 2005

ABSTRACT

Degenerate primers were used to amplify 14 distinct reductive-dehalogenase-homologous (RDH) genes from the
Dehalococcoides
-containing mixed culture KB1. Most of the corresponding predicted proteins were highly similar (97 to >99% amino acid identity) to previously reported
Dehalococcoides
reductive dehalogenases. To examine the differential transcription of these RDH genes, KB1 was split into five subcultures amended with either trichloroethene,
cis-
1,2-dichloroethene, vinyl chloride, 1,2-dichlorethane, or no chlorinated electron acceptor. Total RNA was extracted following the onset of reductive dechlorination, and RDH transcripts were reverse transcribed and amplified using degenerate primers. The results indicate that the transcription of RDH genes requires the presence of a chlorinated electron acceptor, and for all treatments, multiple RDH genes were simultaneously transcribed, with transcripts of two of the genes being present under all four electron-accepting conditions. Two of the transcribed sequences were highly similar to reported vinyl chloride reductase genes, namely,
vcrA
from
Dehalococcoides
sp. strain VS and
bvcA
from
Dehalococcoides
sp. strain BAV1. These findings suggest that multiple RDH genes are induced by a single chlorinated substrate and that multiple reductive dehalogenases contribute to chloroethene degradation in KB1.
A Broad-Host-Range, Generalized Transducing Phage (SN-T) Acquires 16S rRNA Genes from Different Genera of Bacteria
Authors: Beumer Amy , Robinson Jayne B. 
Published: December 2005

ABSTRACT

Genomic analysis has revealed heterogeneity among bacterial 16S rRNA gene sequences within a single species; yet the cause(s) remains uncertain. Generalized transducing bacteriophages have recently gained recognition for their abundance as well as their ability to affect lateral gene transfer and to harbor bacterial 16S rRNA gene sequences. Here, we demonstrate the ability of broad-host-range, generalized transducing phages to acquire 16S rRNA genes and gene sequences. Using PCR and primers specific to conserved regions of the 16S rRNA gene, we have found that generalized transducing phages (D3112, UT1, and SN-T), but not specialized transducing phages (D3), acquired entire bacterial 16S rRNA genes. Furthermore, we show that the broad-host-range, generalized transducing phage SN-T is capable of acquiring the 16S rRNA gene from two different genera:
Sphaerotilus natans
, the host from which SN-T was originally isolated, and
Pseudomonas aeruginosa
. In sequential infections, SN-T harbored only 16S rRNA gene sequences of the final host as determined by restriction fragment length polymorphism analysis. The frequency of 16S rRNA gene sequences in SN-T populations was determined to be 1 × 10
−9
transductants/PFU. Our findings further implicate transduction in the horizontal transfer of 16S rRNA genes between different species or genera of bacteria.
Detection of Genetic Markers of Fecal Indicator Bacteria in Lake Michigan and Determination of Their Relationship to
Escherichia coli
Densities Using Standard Microbiological Methods
Authors: Bower Patricia A. , Scopel Caitlin O. , Jensen Erika T. , Depas Morgan M. , McLellan Sandra L. 
Published: December 2005

ABSTRACT

Lake Michigan surface waters impacted by fecal pollution were assessed to determine the occurrence of genetic markers for
Bacteroides
and
Escherichia coli
. Initial experiments with sewage treatment plant influent demonstrated that total
Bacteroides
spp. could be detected by PCR in a 25- to 125-fold-higher dilution series than
E. coli
and human-specific
Bacteroides
spp., which were both found in similar dilution ranges. The limit of detection for the human-specific genetic marker ranged from 0.2 CFU/100 ml to 82 CFU/100 ml culturable
E. coli
for four wastewater treatment plants in urban and rural areas. The spatial and temporal distributions of these markers were assessed following major rain events that introduced urban storm water, agricultural runoff, and sewage overflows into Lake Michigan.
Bacteroides
spp. were detected in all of these samples by PCR, including those with <1 CFU/100 ml
E. coli
. Human-specific
Bacteroides
spp. were detected as far as 2 km into Lake Michigan during sewage overflow events, with variable detection 1 to 9 days postoverflow, whereas the cow-specific
Bacteroides
spp. were detected in only highly contaminated samples near the river outflow. Lake Michigan beaches were also assessed throughout the summer season for the same markers.
Bacteroides
spp. were detected in all beach samples, including 28 of the 74 samples that did not exceed 235 CFU/100 ml of
E. coli
. Human-specific
Bacteroides
spp. were detected at three of the seven beaches; one of the sites demonstrating positive results was sampled during a reported sewage overflow, but
E. coli
levels were below 235 CFU/100 ml. This study demonstrates the usefulness of non-culture-based microbial-source tracking approaches and the prevalence of these genetic markers in the Great Lakes, including freshwater coastal beaches.
Listeria monocytogenes
PerR Mutants Display a Small-Colony Phenotype, Increased Sensitivity to Hydrogen Peroxide, and Significantly Reduced Murine Virulence
Authors: Rea Rosemarie , Hill Colin , Gahan Cormac G. M. 
Published: December 2005

ABSTRACT

Deletion of
perR
in
Listeria monocytogenes
results in a small-colony phenotype (Δ
perR
sm
) that is slow growing and exhibits increased sensitivity to H
2
O
2
. At a relatively high frequency, large-colony variants (Δ
perR
lg
) arise, which are more resistant to H
2
O
2
than the wild-type and ultimately dominate the culture. Transcriptional analysis revealed that the
kat
gene (catalase) is up-regulated in both types of mutants and that the highest level is apparent in Δ
perR
sm
mutants, demonstrating PerR regulation of this gene. Overexpression of the catalase gene in the wild-type background resulted in a slower-growing strain with a smaller colony size similar to that of Δ
perR
sm
. By combining a bioinformatic approach with experimental evidence, other PerR-regulated genes were identified, including
fur
, lmo0641,
fri
, lmo1604,
hemA
, and
trxB
. The transcriptional profile of these genes in both mutant backgrounds was similar to that of catalase in that a higher level of expression was observed in Δ
perR
sm
than in the wild type or Δ
perR
lg
. Murine studies revealed that the virulence potential of the Δ
perR
sm
mutant is substantially reduced compared to that of the wild-type and Δ
perR
lg
strains. Collectively, the data demonstrate that the Δ
perR
sm
mutant represents the true phenotype associated with the absence of PerR, which is linked to overexpression of regulated genes that negatively affect bacterial homeostasis both in vitro and in vivo. A subsequent secondary mutation occurred at a high frequency, which resulted in phenotypic reversion to a large-colony phenotype with increased fitness that may have obstructed the analysis of the role of PerR in the physiology of the bacterial cell.
Influence of Inorganic Nitrogen Management Regime on the Diversity of Nitrite-Oxidizing Bacteria in Agricultural Grassland Soils
Authors: Freitag Thomas E. , Chang Lisa , Clegg Christopher D. , Prosser James I. 
Published: December 2005

ABSTRACT

To assess links between the diversity of nitrite-oxidizing bacteria (NOB) in agricultural grassland soils and inorganic N fertilizer management, NOB communities in fertilized and unfertilized soils were characterized by analysis of clone libraries and denaturing gradient gel electrophoresis (DGGE) of 16S rRNA gene fragments. Previously uncharacterized
Nitrospira-
like sequences were isolated from both long-term-fertilized and unfertilized soils, but DGGE migration patterns indicated the presence of additional sequence types in the fertilized soils. Detailed phylogenetic analysis of
Nitrospira-
like sequences suggests the existence of one newly described evolutionary group and of subclusters within previously described sublineages, potentially representing different ecotypes; the new group may represent a lineage of noncharacterized
Nitrospira
species. Clone libraries of
Nitrobacter
-like sequences generated from soils under different long-term N management regimes were dominated by sequences with high similarity to the rhizoplane isolate
Nitrobacter
sp. strain PJN1. However, the diversity of
Nitrobacter
communities did not differ significantly between the two soil types. This is the first cultivation-independent study of nitrite-oxidizing bacteria in soil demonstrating that nitrogen management practices influence the diversity of this bacterial functional group.
Genetic Variation among
Staphylococcus aureus
Strains from Norwegian Bulk Milk
Authors: Jørgensen H. J. , Mørk T. , Caugant D. A. , Kearns A. , Rørvik L. M. 
Published: December 2005

ABSTRACT

Strains of
Staphylococcus aureus
obtained from bovine (
n
= 117) and caprine (
n
= 114) bulk milk were characterized and compared with
S. aureus
strains from raw-milk products (
n
= 27), bovine mastitis specimens (
n
= 9), and human blood cultures (
n
= 39). All isolates were typed by pulsed-field gel electrophoresis (PFGE). In addition, subsets of isolates were characterized using multilocus sequence typing (MLST), multiplex PCR (m-PCR) for genes encoding nine of the staphylococcal enterotoxins (SE), and the cloverleaf method for penicillin resistance. A variety of genotypes were observed, and greater genetic diversity was found among bovine than caprine bulk milk isolates. Certain genotypes, with a wide geographic distribution, were common to bovine and caprine bulk milk and may represent ruminant-specialized
S. aureus
. Isolates with genotypes indistinguishable from those of strains from ruminant mastitis were frequently found in bulk milk, and strains with genotypes indistinguishable from those from bulk milk were observed in raw-milk products. This indicates that
S. aureus
from infected udders may contaminate bulk milk and, subsequently, raw-milk products. Human blood culture isolates were diverse and differed from isolates from other sources. Genotyping by PFGE, MLST, and m-PCR for SE genes largely corresponded. In general, isolates with indistinguishable PFGE banding patterns had the same SE gene profile and isolates with identical SE gene profiles were placed together in PFGE clusters. Phylogenetic analyses agreed with the division of MLST sequence types into clonal complexes, and isolates within the same clonal complex had the same SE gene profile. Furthermore, isolate
s
within PFGE clusters generally belonged to the same clonal complex.
Complete Sequences of Four Plasmids ofLactococcus lactissubsp.cremorisSK11 Reveal Extensive Adaptation to the Dairy Environment
Authors: Siezen Roland J. , Renckens Bernadet , van Swam Iris , Peters Sander , van Kranenburg Richard , Kleerebezem Michiel , de Vos Willem M. 
Published: December 2005

ABSTRACTLactococcus lactisstrains are known to carry plasmids encoding industrially important traits.L. lactissubsp.cremorisSK11 is widely used by the dairy industry in cheese making. Its complete plasmid complement was sequenced and found to contain the plasmids pSK11A (10,372 bp), pSK11B (13,332 bp), pSK11L (47,165 bp), and pSK11P (75,814 bp). Six highly homologousrepB-containing replicons were found, all belonging to the family of lactococcal theta-type replicons. Twenty-three complete insertion sequence elements segment the plasmids into numerous modules, many of which can be identified as functional units or containing functionally related genes. Plasmid-encoded functions previously known to reside onL. lactisSK11 plasmids were now mapped in detail, e.g., lactose utilization (lacR-lacABCDFEGX), the proteolytic system (prtM-prtP,pepO,pepF), and the oligopeptide permease system (oppDFBCA). Newly identified plasmid-encoded functions could facilitate the uptake of various cations, while thepabAandpabBgenes could be essential for folate biosynthesis. A competitive advantage could be obtained by using the putative flavin adenine dinucleotide-dependentd-lactate dehydrogenase and oxalate:formate antiporter for enhanced ATP synthesis, while the activity of the predicted α-acetolactate decarboxylase may contribute to the formation of an additional electron sink. Various stress response proteins are plasmid encoded, which could enhance strain robustness. A substantial number of these “adaptation” genes have not been described before onL. lactisplasmids. Moreover, several genes were identified for the first time inL. lactis, possibly reflecting horizontal gene transfer.
Detection and Cultivation of Soil Verrucomicrobia
Authors: Sangwan Parveen , Kovac Suzana , Davis Kathryn E. R. , Sait Michelle , Janssen Peter H. 
Published: December 2005

ABSTRACT

Only one isolate each of the class “
Spartobacteria
” (subdivision 2 of the phylum
Verrucomicrobia
) and of subdivision 3 of
Verrucomicrobia
have previously been reported to grow in laboratory culture. Using media that had been used successfully in other studies to isolate members of diverse groups of soil bacteria, we generated a collection of over 1,200 isolates from soil from a pasture. An oligonucleotide probe that targets the 16S rRNA genes of verrucomicrobia was used to screen this collection, and 14 new verrucomicrobia were identified. Nine of these belonged to the class “
Spartobacteria
” and were related to “
Chthoniobacter flavus
.” Five further isolates were members of subdivision 3 and were related to the only known isolate of this subdivision. The differences in the 16S rRNA gene sequences of the new isolates and previously described isolates, of up to 10%, indicated that the new isolates represent new species and genera. All but two of the verrucomicrobial isolates were from colonies that first became visible one or more months after inoculation of plates with soil, but subcultures grew more rapidly. Analysis of PCR-amplified 16S rRNA genes in the pasture soil showed that members of the class “
Spartobacteria
” were more numerous than members of subdivision 3. Isolates of subdivision 3 were only found on plates receiving an inoculum that yielded a mean of 29 colonies per plate, while members of the class “
Spartobacteria
” were only found on plates receiving a more dilute inoculum that resulted in a mean of five colonies per plate. This suggested that colony development by members of the class “
Spartobacteria
” was inhibited by other culturable bacteria.
Enzymatic Ability of
Bifidobacterium animalis
subsp.
lactis
To Hydrolyze Milk Proteins: Identification and Characterization of Endopeptidase O
Authors: Janer C. , Arigoni F. , Lee B. H. , Peláez C. , Requena T. 
Published: December 2005

ABSTRACT

The proteolytic system of
Bifidobacterium animalis
subsp.
lactis
was analyzed, and an intracellular endopeptidase (PepO) was identified and characterized. This work reports the first complete cloning, purification, and characterization of a proteolytic enzyme in
Bifidobacterium
spp. Aminopeptidase activities (general aminopeptidases, proline iminopeptidase, X-prolyl dipeptidylaminopeptidase) found in cell extracts of
B. animalis
subsp.
lactis
were higher for cells that had been grown in a milk-based medium than for those grown in MRS. A high specific proline iminopeptidase activity was observed in
B. animalis
subsp.
lactis
. Whole cells and cell wall-bound protein fractions showed no caseinolytic activity; however, the combined action of intracellular proteolytic enzymes could hydrolyze casein fractions rapidly. The endopeptidase activity of
B. animalis
subsp.
lactis
was examined in more detail, and the gene encoding an endopeptidase O in
B. animalis
subsp.
lactis
was cloned and overexpressed in
Escherichia coli
. The deduced amino acid sequence for
B. animalis
subsp.
lactis
PepO indicated that it is a member of the M13 peptidase family of zinc metallopeptidases and displays 67.4% sequence homology with the predicted PepO protein from
Bifidobacterium longum
. The recombinant enzyme was shown to be a 74-kDa monomer. Activity of
B. animalis
subsp.
lactis
PepO was found with oligopeptide substrates of at least 5 amino acid residues, such as met-enkephalin, and with larger substrates, such as the 23-amino-acid peptide α
s1
-casein(f1-23). The predominant peptide bond cleaved by
B. animalis
subsp.
lactis
PepO was on the N-terminal side of phenylalanine residues. The enzyme also showed a post-proline secondary cleavage site.
Gibberella xylarioides
Sensu Lato from
Coffea canephora
: a New Mating Population in the
Gibberella fujikuroi
Species Complex
Authors: Lepoint Pascale C. E. , Munaut Françoise T. J. , Maraite Henri M. M. 
Published: December 2005

ABSTRACT

Gibberella xylarioides
Heim & Saccas (presumed anamorph,
Fusarium xylarioides
Steyaert) is the causal agent of coffee wilt disease, an economically important tracheomycosis in Africa. In vitro crosses carried out with Congolese, Ugandan, and Tanzanian single-ascospore/conidial isolates originating from diseased
Coffea canephora
/
excelsa
demonstrated a heterothallic mating system, controlled by a single locus with two alleles,
MAT-1
and
MAT-2
. Compatible isolates produced fertile perithecia within 2 to 8 weeks after mating. Mating type (
MAT
) was characterized by PCR with primer pairs previously developed for the
Gibberella fujikuroi
species complex (GFC) and for
Fusarium oxysporum
. All strains analyzed were morphologically identical and corresponded to Booth's description of the “female”
F. xylarioides
strain. Based on crossing results and
MAT-2
/translation elongation 1-α (
tef
) sequence data,
G. xylarioides
, as currently understood, is demonstrated to encompass at least three “groups”:
G. xylarioides
sensu strictu Ia, defined hitherto by two “historical” West African strains originating from the severe 1930s to 1950s epidemic (CBS 25852 and CBS 74979);
G. xylarioides
sensu strictu Ib, defined by two “historical” Central African lowland strains (DSMZ 62457 and ATCC 15664); and
G. xylarioides
sensu lato II, containing Congolese, Ugandan, and Tanzanian
C. canephora
/
excelsa
isolates. Infertility of crosses between the coffee wilt pathogen and known GFC mating populations demonstrates that
G. xylarioides
sensu lato constitutes a new biological species within the
G. fujikuroi
complex. MUCL 44532/MUCL 43887 and MUCL 35223/MUCL 44549 are proposed as
G. xylarioides
sensu lato II
MAT-1
/
MAT-2
reference mating type tester strains.
Multilocus Sequence Type System for the Plant Pathogen
Xylella fastidiosa
and Relative Contributions of Recombination and Point Mutation to Clonal Diversity
Authors: Scally Mark , Schuenzel Erin L. , Stouthamer Richard , Nunney Leonard 
Published: December 2005

ABSTRACT

Multilocus sequence typing (MLST) identifies and groups bacterial strains based on DNA sequence data from (typically) seven housekeeping genes. MLST has also been employed to estimate the relative contributions of recombination and point mutation to clonal divergence. We applied MLST to the plant pathogen
Xylella fastidiosa
using an initial set of sequences for 10 loci (9.3 kb) of 25 strains from five different host plants, grapevine (PD strains), oleander (OLS strains), oak (OAK strains), almond (ALS strains), and peach (PP strains). An eBURST analysis identified six clonal complexes using the grouping criterion that each member must be identical to at least one other member at 7 or more of the 10 loci. These clonal complexes corresponded to previously identified phylogenetic clades; clonal complex 1 (CC1) (all PD strains plus two ALS strains) and CC2 (OLS strains) defined the
X. fastidiosa
subsp.
fastidiosa
and
X. fastidiosa
subsp.
sandyi
clades, while CC3 (ALS strains), CC4 (OAK strains), and CC5 (PP strains) were subclades of
X. fastidiosa
subsp.
multiplex
. CC6 (ALS strains) identified an
X. fastidiosa
subsp.
multiplex
-like group characterized by a high frequency of intersubspecific recombination. Compared to the recombination rate in other bacterial species, the recombination rate in
X. fastidiosa
is relatively low. Recombination between different alleles was estimated to give rise to 76% of the nucleotide changes and 31% of the allelic changes observed. The housekeeping loci
holC
,
nuoL
,
leuA
,
gltT
,
cysG
,
petC
, and
lacF
were chosen to form the basis of a public database for typing
X. fastidiosa
(
www.mlst.net
). These loci identified the same six clonal complexes using the strain grouping criterion of identity at five or more loci with at least one other member.
Structure and Biosynthesis of the BT Peptide Antibiotic from
Brevibacillus texasporus
Authors: Wu Xiaofeng , Ballard Johnathan , Jiang Yi Wei 
Published: December 2005

ABSTRACT

We isolated a novel gram-positive bacterium,
Brevibacillus texasporus
, that produces an antibiotic, BT. BT is a group of related peptides that are produced by
B. texasporus
cells in response to nutrient limitation. We report here purification and determination of the structure of the most abundant BT isomer, BT1583. Amino acid composition and tandem mass spectrometry experiments yielded a partial BT1583 structure. The presence of ornithine and
d
-form residues in the partial BT1583 structure indicated that the peptide is synthesized by a nonribosomal peptide synthetase (NRPS). The BT NRPS operon was rapidly and accurately identified by using a novel in silico NRPS operon hunting strategy that involved direct shotgun genomic sequencing rather than the unreliable cosmid library hybridization scheme. Sequence analysis of the BT NRPS operon indicated that it encodes a colinear modular NRPS with a strict correlation between the NRPS modules and the amino acid residues in the peptide. The colinear nature of the BT NRPS enabled us to utilize the genomic information to refine the BT1583 peptide sequence to Me
2
-4-methyl-4-[(E)-2-butenyl]-4,
N
-methyl-threonine-L-
d
O-I-V-V-
d
K-V-
d
L-K-
d
Y-L-V-CH
2
OH. In addition, we report the discovery of novel NRPS codons (sets of the substrate specificity-conferring residues in NRPS modules) for valine, lysine, ornithine, and tyrosine.