bluuebunny/crossref_metadata_2025_split huggingface dataset:
Pyrosequencing of Tagged 16S rRNA Gene Amplicons for Rapid Deciphering of the Microbiomes of Fermented Foods Such as Pearl Millet Slurries
Authors: Humblot Christèle , Guyot Jean-Pierre 
Published: May 2009

ABSTRACT

Pearl millet slurries, mixed with groundnuts or not, were chosen as a model to investigate the feasibility of obtaining a rapid overview of community structure and population dynamics of fermented foods using pyrosequencing of tagged 16S rRNA gene amplicons. From 14 fermented samples collected either in a traditional small-scale processing unit in Burkina Faso or at laboratory scale, 137,469 sequences of bacterial 16S rRNA gene amplicons were characterized. Except for a few
Proteobacteria
, almost all the bacterial sequences were attributed to cultivable bacteria. This approach enabled 80.7% of the sequences to be attributed to a family and 70% to a genus but did not enable identification to the species level. The bacterial sequences were assigned to four phyla, with
Firmicutes
representing the highest diversity, followed by
Proteobacteria
,
Actinobacteria
, and
Bacteroidetes
, which were found only in the slurries prepared in traditional production units. Most of the
Firmicutes
were lactic acid bacteria, mainly represented by members of the
Lactobacillus
,
Pediococcus
,
Leuconostoc
, and
Weissella
genera, whose ratio varied from the onset to the end of the fermentation. The other bacteria present at the beginning of fermentation were generally no longer detected at the end, which is consistent with already-known patterns in the microbial ecology of fermented foods. In conclusion, this method seems very promising for rapid and preliminary microbial characterization in many samples of an unknown food sample, by determining numerous nucleic sequences simultaneously without the need for cloning and cultivation-dependent methods.
Versatile Solidified Nanofibrous Cellulose-Containing Media for Growth of Extremophiles
Authors: Tsudome Mikiko , Deguchi Shigeru , Tsujii Kaoru , Ito Susumu , Horikoshi Koki 
Published: May 2009

ABSTRACT

Solidified media that employ a porous matrix of nanofibrous cellulose are described. The physicochemical stability of the porous structure allows the development of solidified media that can support the growth of extremophiles, such as acidophilic
Acidiphilium
, alkaliphilic
Bacillus
, thermophilic
Geobacillus
and
Thermus
, alkalithermophilic
Bacillus
, and acidothermophilic
Sulfolobus
microbes. The cellulose-supported media have several advantages over agar- and gellan gum-derived media, including versatility and stability.
Screening of Thermotolerant
Gluconobacter
Strains for Production of 5-Keto-
d
-Gluconic Acid and Disruption of Flavin Adenine Dinucleotide-Containing
d
-Gluconate Dehydrogenase
Authors: Saichana Ittipon , Moonmangmee Duangtip , Adachi Osao , Matsushita Kazunobu , Toyama Hirohide 
Published: May 2009

ABSTRACT

We isolated thermotolerant
Gluconobacter
strains that are able to produce 5-keto-
d
-gluconic acid (5KGA) at 37°C, a temperature at which regular mesophilic 5KGA-producing strains showed much less growth and 5KGA production. The thermotolerant strains produced 2KGA as the major product at both 30 and 37°C. The amount of ketogluconates produced at 37°C was slightly less than the amount produced at 30°C. To improve the yield of 5KGA in these strains, we disrupted flavin adenine dinucleotide-gluconate dehydrogenase (FAD-GADH), which is responsible for 2KGA production. Genes for FAD-GADH were cloned by using inverse PCR and an in vitro cloning strategy. The sequences obtained for three thermotolerant strains were identical and showed high levels of identity to the FAD-GADH sequence reported for the genome of
Gluconobacter oxydans
621 H. A kanamycin resistance gene cassette was used to disrupt the FAD-GADH genes in the thermotolerant strains. The mutant strains produced 5KGA exclusively, and the final yields were over 90% at 30°C and 50% at 37°C. We found that the activity of pyrroloquinoline quinone (PQQ)-dependent glycerol dehydrogenase, which is responsible for 5KGA production, increased in response to addition of PQQ and CaCl
2
in vitro when cells were grown at 37°C. Addition of 5 mM CaCl
2
to the culture medium of the mutant strains increased 5KGA production to the point where over 90% of the initial substrate was converted. The thermotolerant
Gluconobacter
strains that we isolated in this study provide a promising new option for industrial 5KGA production.
Phylogeny and Virulence of Naturally Occurring Type III Secretion System-Deficient
Pectobacterium
Strains
Authors: Kim Hye-Sook , Ma Bing , Perna Nicole T. , Charkowski Amy O. 
Published: May 2009

ABSTRACT

Pectobacterium
species are enterobacterial plant-pathogenic bacteria that cause soft rot disease in diverse plant species. Previous epidemiological studies of
Pectobacterium
species have suffered from an inability to identify most isolates to the species or subspecies level. We used three previously described DNA-based methods, 16S-23S intergenic transcribed spacer PCR-restriction fragment length polymorphism analysis, multilocus sequence analysis (MLSA), and pulsed-field gel electrophoresis, to examine isolates from diseased stems and tubers and found that MLSA provided the most reliable classification of isolates. We found that strains belonging to at least two
Pectobacterium
clades were present in each field examined, although representatives of only three of five
Pectobacterium
clades were isolated. Hypersensitive response and DNA hybridization assays revealed that strains of both
Pectobacterium carotovorum
and
Pectobacterium wasabiae
lack a type III secretion system (T3SS). Two of the T3SS-deficient strains assayed lack genes adjacent to the T3SS gene cluster, suggesting that multiple deletions occurred in
Pectobacterium
strains in this locus, and all strains appear to have only six rRNA operons instead of the seven operons typically found in
Pectobacterium
strains. The virulence of most of the T3SS-deficient strains was similar to that of T3SS-encoding strains in stems and tubers.
In Situ Production of Crenarchaeol in Two California Hot Springs
Authors: Pitcher Angela , Schouten Stefan , Sinninghe Damsté Jaap S. 
Published: May 2009

ABSTRACT

Crenarchaeol, a membrane-spanning glycerol dialkyl glycerol tetraether (GDGT) containing a cyclohexane moiety in addition to four cyclopentane moieties, was originally hypothesized to be synthesized exclusively by the mesophilic
Crenarchaeota
. Recent studies reporting the occurrence of crenarchaeol in hot springs and as a membrane constituent of the recently isolated thermophilic crenarchaeote “
Candidatus
Nitrosocaldus yellowstonii,” however, have raised questions regarding its taxonomic distribution and function. To determine whether crenarchaeol in hot springs is indeed synthesized by members of the
Archaea
in situ or is of allochthonous origin, we quantified crenarchaeol present in the form of both intact polar lipids (IPLs) and core lipids in sediments of two California hot springs and in nearby soils. IPL-derived crenarchaeol (IPL-crenarchaeol) was found in both hot springs and soils, suggesting in situ production of this GDGT over a wide temperature range (12°C to 89°C). Quantification of archaeal
amoA
gene abundance by quantitative PCR showed a good correspondence with IPL-crenarchaeol, suggesting that it was indeed derived from living cells and that crenarchaeol-synthesizing members of the
Archaea
in our samples may also be ammonia oxidizers.
Response of
Listeria monocytogenes
to Disinfection Stress at the Single-Cell and Population Levels as Monitored by Intracellular pH Measurements and Viable-Cell Counts
Authors: Kastbjerg Vicky G. , Nielsen Dennis S. , Arneborg Nils , Gram Lone 
Published: May 2009

ABSTRACT

Listeria monocytogenes
has a remarkable ability to survive and persist in food production environments. The purpose of the present study was to determine if cells in a population of
L. monocytogenes
differ in sensitivity to disinfection agents as this could be a factor explaining persistence of the bacterium. In situ analyses of
Listeria monocytogenes
single cells were performed during exposure to different concentrations of the disinfectant Incimaxx DES to study a possible population subdivision. Bacterial survival was quantified with plate counting and disinfection stress at the single-cell level by measuring intracellular pH (pH
i
) over time by fluorescence ratio imaging microscopy. pH
i
values were initially 7 to 7.5 and decreased in both attached and planktonic
L. monocytogenes
cells during exposure to sublethal and lethal concentrations of Incimaxx DES. The response of the bacterial population was homogenous; hence, subpopulations were not detected. However, pregrowth with NaCl protected the planktonic bacterial cells during disinfection with Incimaxx (0.0015%) since pH
i
was higher (6 to 6.5) for the bacterial population pregrown with NaCl than for cells grown without NaCl (pH
i
5 to 5.5) (
P
< 0.05). The protective effect of NaCl was reflected by viable-cell counts at a higher concentration of Incimaxx (0.0031%), where the salt-grown population survived better than the population grown without NaCl (
P
< 0.05). NaCl protected attached cells through drying but not during disinfection. This study indicates that a population of
L. monocytogenes
cells, whether planktonic or attached, is homogenous with respect to sensitivity to an acidic disinfectant studied on the single-cell level. Hence a major subpopulation more tolerant to disinfectants, and hence more persistent, does not appear to be present.
Isolation from Sewage Influent and Characterization of Novel
Staphylococcus aureus
Bacteriophages with Wide Host Ranges and Potent Lytic Capabilities
Authors: Synnott Aidan J. , Kuang Ying , Kurimoto Miki , Yamamichi Keiko , Iwano Hidetomo , Tanji Yasunori 
Published: May 2009

ABSTRACT

Staphylococcus aureus
is a gram-positive pathogen that causes a variety of diseases, including bovine mastitis, which has severe economic consequences. Standard antibiotic treatment results in selection of resistant strains, leading to a need for alternative treatments, such as bacteriophage therapy. Forty-nine
S. aureus
isolates were obtained from the milk of mastitic cows for use in screening of staphylococcal phages. Fifteen isolates which were positive for both coagulase and hemolysin were assayed by PCR for variation in the X region and the immunoglobulin G-binding region of the protein A gene (
spa
) and in the carboxy terminus of the coagulase gene (
coa
) and for the presence of enterotoxin C, G, H, and I genes. The host ranges of 52 phages isolated from sewage influent were determined by performing spot tests with the 15
S. aureus
isolates, and two phages were subsequently chosen for further analysis. ΦSA039 had the widest host range, producing clear plaques on 13 of the 15 isolates (87%), while ΦSA012 produced clear plaques on 8 isolates (53%) and was the only phage that produced a clear plaque on a nonmastitic
S. aureus
strain. Transmission electron microscopy revealed that the phages were similar sizes and belonged to the
Myoviridae
family. Measurement of optical densities during coculture with
S. aureus
isolates confirmed the breadth of the ΦSA039 host range and showed that ΦSA012 had potent lytic capability. ΦSA012-resistant bacteria did not appear for three of seven isolates tested (43%) after 65 h of incubation. These two phages are proposed as candidates for phage therapy of bovine mastitis.
Attenuation ofEdwardsiella tardaVirulence by Small Peptides That Interfere with LuxS/Autoinducer Type 2 Quorum Sensing
Authors: Zhang Min , Jiao Xu-dong , Hu Yong-hua , Sun Li 
Published: May 2009

ABSTRACTEdwardsiella tardais a gram-negative pathogen with a broad host range that includes humans, animals, and fish. Recent studies have shown that the LuxS/autoinducer type 2 (AI-2) quorum sensing system is involved in the virulence ofE. tarda. In the present study, it was found that theE. tardaLuxS mutants bearing deletions of the catalytic site (C site) and the tyrosine kinase phosphorylation site, respectively, are functionally inactive and that these dysfunctional mutants can interfere with the activity of the wild-type LuxS. Two small peptides, 5411 and 5906, which share sequence identities with the C site of LuxS, were identified. 5411 and 5906 proved to be inhibitors of AI-2 activity and could vitiate the infectivity of the pathogenicE. tardastrain TX1. The inhibitory effect of 5411 and 5906 on AI-2 activity is exerted on LuxS, with which these peptides specifically interact. The expression of 5411 and 5906 in TX1 has multiple effects (altering biofilm production and the expression of certain virulence-associated genes), which are similar to those caused by interruption ofluxSexpression. Further study found that it is very likely that 5411 and 5906 can be released from the strains expressing them and, should TX1 be in the vicinity, captured by TX1. Based on this observation, a constitutive 5411 producer (Pseudomonassp. strain FP3/pT5411) was constructed in the form of a fish commensal isolate that expresses 5411 from a plasmid source. The presence of FP3/pT5411 in fish attenuates the virulence of TX1. Finally, it was demonstrated that fish expressing 5411 directly from tissues exhibit enhanced resistance against TX1 infection.
Influence of Culture Conditions and Medium Composition on the Production of Cellulose by Shiga Toxin-Producing
Escherichia coli
Cells
Authors: Yoo Byong Kwon , Chen Jinru 
Published: May 2009

ABSTRACT

Culture conditions favoring cellulose production by Shiga toxin-producing
Escherichia coli
included a 28°C incubation temperature, an aerobic atmosphere, and the presence of 2% ethanol in Luria-Bertani no-salt agar with pH 6.0 and a water activity of 0.99. These findings will assist in formulating microbiological media useful for cellulose and biofilm research.
Propagation of Human Enteropathogens in Constructed Horizontal Wetlands Used for Tertiary Wastewater Treatment
Authors: Graczyk Thaddeus K. , Lucy Frances E. , Tamang Leena , Mashinski Yessika , Broaders Michael A. , Connolly Michelle , Cheng Hui-Wen A. 
Published: May 2009

ABSTRACT

Constructed subsurface flow (SSF) and free-surface flow (FSF) wetlands are being increasingly implemented worldwide into wastewater treatments in response to the growing need for microbiologically safe reclaimed waters, which is driven by an exponential increase in the human population and limited water resources. Wastewater samples from four SSF and FSF wetlands in northwestern Ireland were tested qualitatively and quantitatively for
Cryptosporidium
spp.,
Giardia duodenalis
, and human-pathogenic microsporidia, with assessment of their viability. Overall, seven species of human enteropathogens were detected in wetland influents, vegetated areas, and effluents:
Cryptosporidium parvum
,
C. hominis
,
C. meleagridis
,
C. muris
,
G. duodenalis
,
Encephalitozoon hellem
, and
Enterocytozoon bieneusi
. SSF wetland had the highest pathogen removal rate (i.e.,
Cryptosporidium
, 97.4%;
G. duodenalis
, 95.4%); however, most of these values for FSF were in the negative area (mean, −84.0%), meaning that more pathogens were discharged by FSF wetlands than were delivered to wetlands with incoming wastewater. We demonstrate here that (i) the composition of human enteropathogens in wastewater entering and leaving SSF and FSF wetlands is highly complex and dynamic, (ii) the removal and inactivation of human-pathogenic microorganisms were significantly higher at the SSF wetland, (iii) FSF wetlands may not always provide sufficient remediation for human enteropathogens, (iv) wildlife can contribute a substantial load of human zoonotic pathogens to wetlands, (v) most of the pathogens discharged by wetlands were viable, (vi) large volumes of wetland effluents can contribute to contamination of surface waters used for recreation and drinking water abstraction and therefore represent a serious public health threat, and (vii) even with the best pathogen removal rates achieved by SSF wetland, the reduction of pathogens was not enough for a safety reuse of the reclaimed water. To our knowledge, this is the first report of
C. meleagridis
from Ireland.
Mn(II) Oxidation Is Catalyzed by Heme Peroxidases in “
Aurantimonas manganoxydans
” Strain SI85-9A1 and
Erythrobacter
sp. Strain SD-21
Authors: Anderson C. R. , Johnson H. A. , Caputo N. , Davis R. E. , Torpey J. W. , Tebo B. M. 
Published: May 2009

ABSTRACT

A new type of manganese-oxidizing enzyme has been identified in two alphaproteobacteria, “
Aurantimonas manganoxydans
” strain SI85-9A1 and
Erythrobacter
sp. strain SD-21. These proteins were identified by tandem mass spectrometry of manganese-oxidizing bands visualized by native polyacrylamide gel electrophoresis in-gel activity assays and fast protein liquid chromatography-purified proteins. Proteins of both alphaproteobacteria contain animal heme peroxidase and hemolysin-type calcium binding domains, with the 350-kDa active Mn-oxidizing protein of
A. manganoxydans
containing stainable heme. The addition of both Ca
2+
ions and H
2
O
2
to the enriched protein from
Aurantimonas
increased manganese oxidation activity 5.9-fold, and the highest activity recorded was 700 μM min
−1
mg
−1
. Mn(II) is oxidized to Mn(IV) via an Mn(III) intermediate, which is consistent with known manganese peroxidase activity in fungi. The Mn-oxidizing protein in
Erythrobacter
sp. strain SD-21 is 225 kDa and contains only one peroxidase domain with strong homology to the first 2,000 amino acids of the peroxidase protein from
A. manganoxydans
. The heme peroxidase has tentatively been named MopA (manganese-oxidizing peroxidase) and sheds new light on the molecular mechanism of Mn oxidation in prokaryotes.
Distinct Roles for VeA and LaeA in Development and Pathogenesis ofAspergillus flavus
Authors: Amaike Saori , Keller Nancy P. 
Published: May 2009

ABSTRACTAspergillus flavus, a mycotoxigenic filamentous fungus, colonizes several important agricultural crops, such as maize and peanuts. Two proteins, VeA and LaeA, known to form a nuclear complex inAspergillus nidulanshave been found to positively regulate developmental processes in severalAspergillusspecies. Here, an examination of near-isogenicA. flavusmutants differing in copy number ofveAandlaeAalleles (0, 1, or at least 2 each) revealed critical roles for VeA and LaeA inA. flavusdevelopment and seed colonization. In contrast to the wild type, both null mutants were unable to metabolize host cell lipid reserves and were inhibited by oleic acid in growth assays. The copy number of LaeA but not VeA appeared critical for a density-dependent sclerotial-to-conidial shift, since the multicopylaeA(MClaeA) strain produced relatively constant sclerotial numbers with increasing population size rather than showing the decrease in sclerotia seen in both the wild-type and MCveAstrains. The MCveA-laeAstrain yielded an intermediate phenotype. This study revealed unique roles of VeA and LaeA in seed pathogenesis and fungal biology, distinct from their cooperative regulatory functions in aflatoxin and sclerotial development.
Characterization of a REG/PA28 Proteasome Activator Homolog in
Dictyostelium discoideum
Indicates that the Ubiquitin- and ATP-Independent REGγ Proteasome Is an Ancient Nuclear Protease
Authors: Masson Patrick , Lundin Daniel , Söderbom Fredrik , Young Patrick 
Published: May 2009

ABSTRACT

The nuclear proteasome activator REGγ/PA28γ is an ATP- and ubiquitin-independent activator of the 20S proteasome and has been proposed to degrade and thereby regulate both a key human oncogene, encoding the coactivator SRC-3/AIB1, and the cyclin-dependent kinase inhibitor p21 (Waf/Cip1). We report the identification and characterization of a PA28/REG homolog in
Dictyostelium
. Association of a recombinant
Dictyostelium
REG with the purified
Dictyostelium
20S proteasome led to the preferential stimulation of the trypsin-like proteasome peptidase activity. Immunolocalization studies demonstrated that the proteasome activator is localized to the nucleus and is present in growing as well as starving
Dictyostelium
cells. Our results indicate that the
Dictyostelium
PA28/REG activator can stimulate both the trypsin-like and chymotrypsin-like activities of the 20S proteasome and supports the idea that the REGγ-20S proteasome represents an early unique nuclear degradation pathway for eukaryotic cells.
Gα and Gβ Proteins Regulate the Cyclic AMP Pathway That Is Required for Development and Pathogenicity of the Phytopathogen
Mycosphaerella graminicola
Authors: Mehrabi Rahim , M'Barek Sarrah Ben , van der Lee Theo A. J. , Waalwijk Cees , de Wit Pierre J. G. M. , Kema Gerrit H. J. 
Published: May 2009

ABSTRACT

We identified and functionally characterized genes encoding three Gα proteins and one Gβ protein in the dimorphic fungal wheat pathogen
Mycosphaerella graminicola
, which we designated
MgGpa1
,
MgGpa2
,
MgGpa3
, and
MgGpb1
, respectively. Sequence comparisons and phylogenetic analyses showed that MgGPA1 and MgGPA3 are most related to the mammalian Gα
i
and Gα
s
families, respectively, whereas MgGPA2 is not related to either of these families. On potato dextrose agar (PDA) and in yeast glucose broth (YGB),
MgGpa1
mutants produced significantly longer spores than those of the wild type (WT), and these developed into unique fluffy mycelia in the latter medium, indicating that this gene negatively controls filamentation.
MgGpa3
mutants showed more pronounced yeast-like growth accompanied with hampered filamentation and secreted a dark-brown pigment into YGB. Germ tubes emerging from spores of
MgGpb1
mutants were wavy on water agar and showed a nested type of growth on PDA that was due to hampered filamentation, numerous cell fusions, and increased anastomosis. Intracellular cyclic AMP (cAMP) levels of
MgGpb1
and
MgGpa3
mutants were decreased, indicating that both genes positively regulate the cAMP pathway, which was confirmed because the WT phenotype was restored by adding cAMP to these mutant cultures. The cAMP levels in
MgGpa1
mutants and the WT were not significantly different, suggesting that this gene might be dispensable for cAMP regulation. In planta assays showed that mutants of
MgGpa1
,
MgGpa3
, and
MgGpb1
are strongly reduced in pathogenicity. We concluded that the heterotrimeric G proteins encoded by
MgGpa3
and
MgGpb1
regulate the cAMP pathway that is required for development and pathogenicity in
M. graminicola
.
Increased [
PSI

+

] Appearance by Fusion of Rnq1 with the Prion Domain of Sup35 in
Saccharomyces cerevisiae
Authors: Choe Young-Jun , Ryu Yangkyun , Kim Hyun-Jin , Seok Yeong-Jae 
Published: May 2009

ABSTRACT

During propagation, yeast prions show a strict sequence preference that confers the specificity of prion assembly. Although propagations of [
PSI
+
] and [
RNQ
+
] are independent of each other, the appearance of [
PSI
+
] is facilitated by the presence of [
RNQ
+
]. To explain the [
RNQ
+
] effect on the appearance of [
PSI
+
], the cross-seeding model was suggested, in which Rnq1 aggregates act as imperfect templates for Sup35 aggregation. If cross-seeding events take place in the cytoplasm of yeast cells, the collision frequency between Rnq1 aggregates and Sup35 will affect the appearance of [
PSI
+
]. In this study, to address whether cross-seeding occurs in vivo, a new [
PSI
+
] induction method was developed that exploits a protein fusion between the prion domain of Sup35 (NM) and Rnq1. This fusion protein successfully joins preexisting Rnq1 aggregates, which should result in the localization of NM around the Rnq1 aggregates and hence in an increased collision frequency between NM and Rnq1 aggregates. The appearance of [
PSI
+
] could be induced very efficiently, even with a low expression level of the fusion protein. This study supports the occurrence of in vivo cross-seeding between Sup35 and Rnq1 and provides a new tool that can be used to dissect the mechanism of the de novo appearance of prions.