bluuebunny/crossref_metadata_2025_split huggingface dataset:
Growth of Hyperthermophilic Archaeon
Pyrococcus furiosus
on Chitin Involves Two Family 18 Chitinases
Authors: Gao Jun , Bauer Michael W. , Shockley Keith R. , Pysz Marybeth A. , Kelly Robert M. 
Published: June 2003

ABSTRACT

Pyrococcus furiosus
was found to grow on chitin, adding this polysacharide to the inventory of carbohydrates utilized by this hyperthermophilic archaeon. Accordingly, two open reading frames (
chiA
[Pf1234] and
chiB
[Pf1233]) were identified in the genome of
P. furiosus
, which encodes chitinases with sequence similarity to proteins from the glycosyl hydrolase family 18 in less-thermophilic organisms. Both enzymes contain multiple domains that consist of at least one binding domain and one catalytic domain. ChiA (ca. 39 kDa) contains a putative signal peptide, as well as a binding domain (ChiA
BD
), that is related to binding domains associated with several previously studied bacterial chitinases.
chiB
, separated by 37 nucleotides from
chiA
and in the same orientation, encodes a polypeptide with two different proline-threonine-rich linker regions (6 and 3 kDa) flanking a chitin-binding domain (ChiB
BD
[11 kDa]), followed by a catalytic domain (ChiB
cat
[35 kDa]). No apparent signal peptide is encoded within
chiB
. The two chitinases share little sequence homology to each other, except in the catalytic region, where both have the catalytic glutamic acid residue that is conserved in all family 18 bacterial chitinases. The genes encoding ChiA, without its signal peptide, and ChiB were cloned and expressed in
Escherichia coli.
ChiA exhibited no detectable activity toward chitooligomers smaller than chitotetraose, indicating that the enzyme is an endochitinase. Kinetic studies showed that ChiB followed Michaelis-Menten kinetics toward chitotriose, although substrate inhibition was observed for larger chitooligomers. Hydrolysis patterns on chitooligosaccharides indicated that ChiB is a chitobiosidase, processively cleaving off chitobiose from the nonreducing end of chitin or other chitooligomers. Synergistic activity was noted for the two chitinases on colloidal chitin, indicating that these two enzymes work together to recruit chitin-based substrates for
P. furiosus
growth. This was supported by the observed growth on chitin as the sole carbohydrate source in sulfur-free media.
Ammonia-Oxidizing Bacteria along Meadow-to-Forest Transects in the Oregon Cascade Mountains
Authors: Mintie A. T. , Heichen R. S. , Cromack, K. , Myrold D. D. , Bottomley P. J. 
Published: June 2003

ABSTRACTAlthough nitrification has been well studied in coniferous forests of Western North America, communities of NH3-oxidizing bacteria in these forests have not been characterized. Studies were conducted along meadow-to-forest transects at two sites (Lookout and Carpenter) in the H. J. Andrews Experimental Forest, located in the Cascade Mountains of Oregon. Soil samples taken at 10- or 20-m intervals along the transects showed that several soil properties, including net nitrogen mineralization and nitrification potential rates changed significantly between vegetation zones. Nonetheless, terminal restriction fragment length polymorphism (T-RFLP) analysis of the PCR-amplified NH3monooxygenase subunit A gene (amoA) showed the same DNA fragments (TaqI [283 bp],CfoI [66 bp], andAluI [392 bp]) to dominate ≥45 of 47 soil samples recovered from both sites. Two fragments (491-bpAluI [AluI491] andCfoI135) were found more frequently in meadow and transition zone soil samples than in forest samples at both sites. At the Lookout site the combinationAluI491-CfoI135 was found primarily in meadow samples expressing the highest N mineralization rates. Four uniqueamoAsequences were identified among 15 isolates recovered into pure culture from various transect locations. Six isolates possessed the most common T-RFLPamoAfingerprint of the soil samples (TaqI283-AluI392-CfoI66), and theiramoAsequences shared 99.8% similarity with a cultured species,Nitrosospirasp.strain Ka4 (cluster 4). The other threeamoAsequences were most similar to sequences ofNitrosospirasp.strain Nsp1 andNitrosospira briensis(cluster 3). 16S ribosomal DNA sequence analysis confirmed the affiliation of these isolates withNitrosospiraclusters 3 and 4. TwoamoAclone sequences matched T-RFLP fingerprints found in soil, but they were not found among the isolates.
Identification ofListeria monocytogenesGenes Involved in Salt and Alkaline-pH Tolerance
Authors: Gardan Rozenn , Cossart Pascale ,   The European Listeria Genome Consortium, Labadie Jean 
Published: June 2003

ABSTRACTThe capacity ofListeria monocytogenesto tolerate salt and alkaline stresses is of particular importance, as this pathogen is often exposed to such environments during food processing and food preservation. We screened a library of Tn917-lacZinsertional mutants in order to identify genes involved in salt and/or alkaline tolerance. We isolated six mutants sensitive to salt stress and 12 mutants sensitive to salt and alkaline stresses. The position of the insertion of the transposon was located in 15 of these mutants. In six mutants the transposon was inserted in intergenic regions, and in nine mutants it was inserted in genes. Most of the genes have unknown functions, but sequence comparisons indicated that they encode putative transporters.
Vector Systems Allowing Efficient Autonomous or Integrative Gene Cloning inMicromonosporasp. Strain 40027
Authors: Li Xiaohua , Zhou Xiufen , Deng Zixin 
Published: June 2003

ABSTRACTVector systems allowing autonomous or site-specific integrative gene cloning were developed forMicromonosporasp. strain 40027, a producer of the antibiotic fortimicin A. The autonomous system depends on the discovery of a low-copy-number, self-transmissible covalently closed circular plasmid, pJTU112 (ca. 14.1 kb), which was shown to be present in the progenitor strain in both integrated and autonomous states. The copy numbers of both wild-type pJTU112 and three derivatives of it can be amplified at least sixfold by addition of streptomycin to the culture medium. The integrative system was developed by the use of a pBR322-derivedEscherichia coliplasmid vector, pSET152, mediated by theattP site of theStreptomycesphage ΦC31. Both vectors can be transferred by conjugation fromE. coliintoMicromonosporasp. strain 40027. The heterologous cloning and expression of thedndgene cluster originating fromStreptomyces lividans1326 intoMicromonosporasp. strain 40027 demonstrated the use of the two systems.
Aerobic Denitrifying Bacteria That Produce Low Levels of Nitrous Oxide
Authors: Takaya Naoki , Catalan-Sakairi Maria Antonina B. , Sakaguchi Yasushi , Kato Isao , Zhou Zhemin , Shoun Hirofumi 
Published: June 2003

ABSTRACT

Most denitrifiers produce nitrous oxide (N
2
O) instead of dinitrogen (N
2
) under aerobic conditions. We isolated and characterized novel aerobic denitrifiers that produce low levels of N
2
O under aerobic conditions. We monitored the denitrification activities of two of the isolates, strains TR2 and K50, in batch and continuous cultures. Both strains reduced nitrate (NO
3
−
) to N
2
at rates of 0.9 and 0.03 μmol min
−1
unit of optical density at 540 nm
−1
at dissolved oxygen (O
2
) (DO) concentrations of 39 and 38 μmol liter
−1
, respectively. At the same DO level, the typical denitrifier
Pseudomonas stutzeri
and the previously described aerobic denitrifier
Paracoccus denitrificans
did not produce N
2
but evolved more than 10-fold more N
2
O than strains TR2 and K50 evolved. The isolates denitrified NO
3
−
with concomitant consumption of O
2
. These results indicated that strains TR2 and K50 are aerobic denitrifiers. These two isolates were taxonomically placed in the β subclass of the class
Proteobacteria
and were identified as
P. stutzeri
TR2 and
Pseudomonas
sp. strain K50. These strains should be useful for future investigations of the mechanisms of denitrifying bacteria that regulate N
2
O emission, the single-stage process for nitrogen removal, and microbial N
2
O emission into the ecosystem.
A Multiplex Reverse Transcription-PCR Method for Detection of Human Enteric Viruses in Groundwater
Authors: Fout G. Shay , Martinson Beth C. , Moyer Michael W. N. , Dahling Daniel R. 
Published: June 2003

ABSTRACT
Untreated groundwater is responsible for about half of the waterborne disease outbreaks in the United States. Human enteric viruses are thought to be leading etiological agents of many of these outbreaks, but there is relatively little information on the types and levels of viruses found in groundwater. To address this problem, monthly samples from 29 groundwater sites were analyzed for 1 year for enteroviruses, hepatitis A virus, Norwalk virus, reoviruses, and rotaviruses by multiplex reverse transcription-PCR (RT-PCR). A procedure with which to remove environmental RT-PCR inhibitors from groundwater samples was developed. The procedure allowed an average of 71 liters of the original groundwater to be assayed per RT-PCR, with an average virus recovery rate of 74%, based on seeded samples. Human enteric viruses were detected in 16% of the groundwater samples analyzed, with reoviruses being the most frequently detected virus group.
Identification of Intermediate and Branch Metabolites Resulting from Biotransformation of 2-Benzoxazolinone by
Fusarium verticillioides
Authors: Glenn A. E. , Meredith F. I. , Morrison W. H. , Bacon C. W. 
Published: June 2003

ABSTRACT

Detoxification of the maize (
Zea mays
) antimicrobial compound 2-benzoxazolinone by the fungal endophyte
Fusarium verticillioides
involves two genetic loci,
FDB1
and
FDB2
, and results in the formation of
N
-(2-hydroxyphenyl)malonamic acid. Intermediate and branch metabolites were previously suggested to be part of the biotransformation pathway. Evidence is presented here in support of 2-aminophenol as the intermediate metabolite and 2-acetamidophenol as the branch metabolite, which was previously designated as BOA-X. Overall, 2-benzoxazolinone metabolism involves hydrolysis (
FDB1
) to produce 2-aminophenol, which is then modified (
FDB2
) by addition of a malonyl group to produce
N
-(2-hydroxyphenyl)malonamic acid. If the modification is prevented due to genetic mutation (
fbd2
), then 2-acetamidophenol may accumulate as a result of addition of an acetyl group to 2-aminophenol. This study resolves the overall chemistry of the 2-benzoxazolinone detoxification pathway, and we hypothesize that biotransformation of the related antimicrobial 6-methoxy-2-benzoxazolinone to produce
N
-(2-hydroxy-4-methoxyphenyl)malonamic acid also occurs via the same enzymatic modifications. Detoxification of these antimicrobials by
F. verticillioides
apparently is not a major virulence factor but may enhance the ecological fitness of the fungus during colonization of maize stubble and field debris.
Intraspecific Diversity of
Vibrio vulnificus
in Galveston Bay Water and Oysters as Determined by Randomly Amplified Polymorphic DNA PCR
Authors: Lin Meilan , Payne Deborah A. , Schwarz John R. 
Published: June 2003

ABSTRACT

Randomly amplified polymorphic DNA (RAPD) PCR was used to analyze the temporal and spatial intraspecific diversity of 208
Vibrio vulnificus
strains isolated from Galveston Bay water and oysters at five different sites between June 2000 and June 2001.
V. vulnificus
was not detected during the winter months (December through February). The densities of
V. vulnificus
in water and oysters were positively correlated with water temperature. Cluster analysis of RAPD PCR profiles of the 208
V. vulnificus
isolates revealed a high level of intraspecific diversity among the strains. No correlation was found between the intraspecific diversity among the isolates and sampling site or source of isolation. After not being detected during the winter months, the genetic diversity of
V. vulnificus
strains first isolated in March was 0.9167. Beginning in April, a higher level of intraspecific diversity (0.9933) and a major shift in population structure were observed among
V. vulnificus
isolates. These results suggest that a great genetic diversity of
V. vulnificus
strains exists in Galveston Bay water and oysters and that the population structure of this species is linked to changes in environmental conditions, especially temperature.
Bacteriophage Ecology in Commercial Sauerkraut Fermentations
Authors: Lu Z. , Breidt F. , Plengvidhya V. , Fleming H. P. 
Published: June 2003

ABSTRACT

Knowledge of bacteriophage ecology in vegetable fermentations is essential for developing phage control strategies for consistent and high quality of fermented vegetable products. The ecology of phages infecting lactic acid bacteria (LAB) in commercial sauerkraut fermentations was investigated. Brine samples were taken from four commercial sauerkraut fermentation tanks over a 60- or 100-day period in 2000 and 2001. A total of 171 phage isolates, including at least 26 distinct phages, were obtained. In addition, 28 distinct host strains were isolated and identified as LAB by restriction analysis of the intergenic transcribed spacer region and 16S rRNA sequence analysis. These host strains included
Leuconostoc
,
Weissella
, and
Lactobacillus
species. It was found that there were two phage-host systems in the fermentations corresponding to the population shift from heterofermentative to homofermentative LAB between 3 and 7 days after the start of the fermentations. The data suggested that phages may play an important role in the microbial ecology and succession of LAB species in vegetable fermentations. Eight phage isolates, which were independently obtained two or more times, were further characterized. They belonged to the family
Myoviridae
or
Siphoviridae
and showed distinct host ranges and DNA fingerprints. Two of the phage isolates were found to be capable of infecting two
Lactobacillus
species. The results from this study demonstrated for the first time the complex phage ecology present in commercial sauerkraut fermentations, providing new insights into the bioprocess of vegetable fermentations.
Population Genetics of
Vibrio vulnificus
: Identification of Two Divisions and a Distinct Eel-Pathogenic Clone
Authors: Gutacker Michaela , Conza Nadine , Benagli Cinzia , Pedroli Ambra , Bernasconi Marco Valerio , Permin Lise , Aznar Rosa , Piffaretti Jean-Claude 
Published: June 2003

ABSTRACT

Genetic relationships among 62
Vibrio vulnificus
strains of different geographical and host origins were analyzed by multilocus enzyme electrophoresis (MLEE), random amplification of polymorphic DNA (RAPD), and sequence analyses of the
recA
and
glnA
genes. Out of 15 genetic loci analyzed by MLEE, 11 were polymorphic. Cluster analysis identified 43 distinct electrophoretic types (ETs) separating the
V. vulnificus
population into two divisions (divisions Ι and ΙΙ). One ET (ET 35) included all indole-negative isolates from diseased eels worldwide (biotype 2). A second ET (ET 2) marked all of the strains from Israel isolated from patients who handled St. Peter's fish (biotype 3). RAPD analysis of the 62
V. vulnificus
isolates identified 26 different profiles separated into two divisions as well. In general, this subdivision was comparable (but not identical) to that observed by MLEE. Phylogenetic analysis of 543 bp of the
recA
gene and of 402 bp of the
glnA
gene also separated the
V. vulnificus
population into two major divisions in a manner similar to that by MLEE and RAPD. Sequence data again indicated the overall subdivision of the
V. vulnificus
population into different biotypes. In particular, indole-negative eel-pathogenic isolates (biotype 2) on one hand and the Israeli isolates (biotype 3) on the other tended to cluster together in both gene trees. None of the methods showed an association between distinct clones and human clinical manifestations. Furthermore, except for the Israeli strains, only minor clusters comprising geographically related isolates were observed. In conclusion, all three approaches (MLEE, RAPD, and DNA sequencing) generated comparable but not always equivalent results. The significance of the two divisions (divisions Ι and ΙΙ) still remains to be clarified, and a reevaluation of the definition of the biotypes is also needed.
Community Analysis of Ammonia and Nitrite Oxidizers during Start-Up of Nitritation Reactors
Authors: Egli Konrad , Langer Christian , Siegrist Hans-Ruedi , Zehnder Alexander J. B. , Wagner Michael , van der Meer Jan Roelof 
Published: June 2003

ABSTRACT

Partial nitrification of ammonium to nitrite under oxic conditions (nitritation) is a critical process for the effective use of alternative nitrogen removal technologies from wastewater. Here we investigated the conditions which promote establishment of a suitable microbial community for performing nitritation when starting from regular sewage sludge. Reactors were operated in duplicate under different conditions (pH, temperature, and dilution rate) and were fed with 50 mM ammonium either as synthetic medium or as sludge digester supernatant. In all cases, stable nitritation could be achieved within 10 to 20 days after inoculation. Quantitative in situ hybridization analysis with group-specific fluorescent rRNA-targeted oligonucleotides (FISH) in the different reactors showed that nitrite-oxidizing bacteria of the genus
Nitrospira
were only active directly after inoculation with sewage sludge (up to 4 days and detectable up to 10 days). As demonstrated by quantitative FISH and restriction fragment length polymorphism (RFLP) analyses of the
amoA
gene (encoding the active-site subunit of the ammonium monooxygenase), the community of ammonia-oxidizing bacteria changed within the first 15 to 20 days from a more diverse set of populations consisting of members of the
Nitrosomonas communis
and
Nitrosomonas oligotropha
sublineages and the
Nitrosomonas europaea-Nitrosomonas eutropha
subgroup in the inoculated sludge to a smaller subset in the reactors. Reactors operated at 30°C and pH 7.5 contained reproducibly homogeneous communities dominated by one
amoA
RFLP type from the
N. europaea-N. eutropha
group. Duplicate reactors at pH 7.0 developed into diverse communities and showed transient population changes even within the ammonia oxidizer community. Reactors at pH 7.5 and 25°C formed communities that were indistinguishable by the applied FISH probes but differing in
amoA
RFLP types. Communities in reactors fed with sludge digester supernatant exhibited a higher diversity and were constantly reinoculated with ammonium oxidizers from the supernatant. Therefore, such systems could be maintained at a higher dilution rate (0.75 day
−1
compared to 0.2 day
−1
for the synthetic wastewater reactors). Despite similar reactor performance with respect to chemical parameters, the underlying community structures were different, which may have an influence on stability during perturbations.
Cultivation-Dependent and -Independent Approaches for Determining Bacterial Diversity in Heavy-Metal-Contaminated Soil
Authors: Ellis Richard J. , Morgan Philip , Weightman Andrew J. , Fry John C. 
Published: June 2003

ABSTRACTIn recent years, culture-independent methods have been used in preference to traditional isolation techniques for microbial community analysis. However, it is questionable whether uncultured organisms from a given sample are important for determining the impact of anthropogenic stress on indigenous communities. To investigate this, soil samples were taken from a site with patchy metal contamination, and the bacterial community structure was assessed with a variety of approaches. There were small differences in microscopic epifluorescence bacterial counts. Denaturing gradient gel electrophoresis (DGGE) profiles of 16S rRNA gene fragments (16S-DGGE) amplified directly from soil samples were highly similar. A clone library generated from the most contaminated sample revealed a diverse bacterial community, which showed similarities to pristine soil communities from other studies. However, the proportion of bacteria from the soil samples that were culturable on standard plate-counting media varied between 0.08 and 2.2%, and these values correlated negatively with metal concentrations. The culturable communities from each sample were compared by 16S-DGGE of plate washes and by fatty acid profiling of individual isolates. Each approach indicated that there were considerable differences between the compositions of the culturable communities from each sample. DGGE bands from both culture-based and culture-independent approaches were sequenced and compared. These data indicated that metal contamination did not have a significant effect on the total genetic diversity present but affected physiological status, so that the number of bacteria capable of responding to laboratory culture and their taxonomic distribution were altered. Thus, it appears that plate counts may be a more appropriate method for determining the effect of heavy metals on soil bacteria than culture-independent approaches.
Unusual Organization for Lactose and Galactose Gene Clusters in
Lactobacillus helveticus
Authors: Fortina Maria Grazia , Ricci Giovanni , Mora Diego , Guglielmetti Simone , Manachini Pier Luigi 
Published: June 2003

ABSTRACT

The nucleotide sequences of the
Lactobacillus helveticus
lactose utilization genes were determined, and these genes were located and oriented relative to one another. The
lacLM
genes (encoding the β-galactosidase protein) were in a divergent orientation compared to
lacR
(regulatory gene) and
lacS
(lactose transporter). Downstream from
lacM
was an open reading frame (
galE
) encoding a UDP-galactose 4 epimerase, and the open reading frame had the same orientation as
lacM
. The
lacR
gene was separated from the downstream
lacS
gene by 2.0 kb of DNA containing several open reading frames that were derived from fragmentation of another permease gene (
lacS
′). Northern blot analysis revealed that
lacL
,
lacM
, and
galE
made up an operon that was transcribed in the presence of lactose from an upstream
lacL
promoter. The inducible genes
lacL
and
lacM
were regulated at the transcriptional level by the LacR repressor. In the presence of glucose and galactose
galE
was transcribed from its promoter, suggesting that the corresponding enzyme can be expressed constitutively. Lactose transport was inducible by addition of lactose to the growth medium.
Involvement of the Reserve Material Poly-β-Hydroxybutyrate inAzospirillum brasilenseStress Endurance and Root Colonization
Authors: Kadouri Daniel , Jurkevitch Edouard , Okon Yaacov 
Published: June 2003

ABSTRACTWhen grown under suboptimal conditions, rhizobacteria of the genusAzospirillumproduce high levels of poly-β-hydroxybutyrate (PHB).Azospirillum brasilensestrain Sp7 and aphbC(PHB synthase) mutant strain in which PHB production is impaired were evaluated for metabolic versatility, for the ability to endure various stress conditions, for survival in soil inoculants, and for the potential to promote plant growth. The carbon source utilization data were similar for the wild-type and mutant strains, but the generation time of the wild-type strain was shorter than that of the mutant strain with all carbon sources tested. The ability of the wild type to endure UV irradiation, heat, osmotic pressure, osmotic shock, and desiccation and to grow in the presence of hydrogen peroxide was greater than that of the mutant strain. The motility and cell aggregation of the mutant strain were greater than the motility and cell aggregation of the wild type. However, the wild type exhibited greater chemotactic responses towards attractants than the mutant strain exhibited. The wild-type strain exhibited better survival than the mutant strain in carrier materials used for soil inoculants, but no difference in the ability to promote plant growth was detected between the strains. In soil, the two strains colonized roots to the same extent. It appears that synthesis and utilization of PHB as a carbon and energy source byA. brasilenseunder stress conditions favor establishment of this bacterium and its survival in competitive environments. However, inA. brasilense, PHB production does not seem to provide an advantage in root colonization under the conditions tested.