bluuebunny/crossref_metadata_2025 huggingface dataset:
3′ Untranslated Region-Dependent Degradation of the
aceA
mRNA, Encoding the Glyoxylate Cycle Enzyme Isocitrate Lyase, by RNase E/G in Corynebacterium glutamicum
Authors: Maeda Tomoya , Wachi Masaaki
Published: October 2012

We previously reported that the
Corynebacterium glutamicum
RNase E/G encoded by the
rneG
gene (NCgl2281) is required for the 5′ maturation of 5S rRNA. In the search for the intracellular target RNAs of RNase E/G other than the 5S rRNA precursor, we detected that the amount of isocitrate lyase, an enzyme of the glyoxylate cycle, increased in
rneG
knockout mutant cells grown on sodium acetate as the sole carbon source. Rifampin chase experiments showed that the half-life of the
aceA
mRNA was about 4 times longer in the
rneG
knockout mutant than in the wild type. Quantitative real-time PCR analysis also confirmed that the level of
aceA
mRNA was approximately 3-fold higher in the
rneG
knockout mutant strain than in the wild type. Such differences were not observed in other mRNAs encoding enzymes involved in acetate metabolism. Analysis by 3′ rapid amplification of cDNA ends suggested that RNase E/G cleaves the
aceA
mRNA at a single-stranded AU-rich region in the 3′ untranslated region (3′-UTR). The
lacZ
fusion assay showed that the 3′-UTR rendered
lacZ
mRNA RNase E/G dependent. These findings indicate that RNase E/G is a novel regulator of the glyoxylate cycle in
C. glutamicum
.
Population Genetic Structure of 4,12:a:− Salmonella enterica Strains from Harbor Porpoises
Authors: Haase Jana K. , Brown Derek J. , Weill François-Xavier , Mather Henry , Foster Geoffrey , Brisse Sylvain , Wain John , Achtman Mark
Published: October 2012

According to pulsed-field gel electrophoresis (PFGE) typing, 4,12:a:−
Salmonella enterica
isolates from harbor porpoises are highly diverse. However, porpoise isolates belong to only two multilocus sequence types within the eBurst group 18 (eBG18) genetic cluster, which also includes
S. enterica
serovars Bispebjerg and Abortusequi. Isolates of other, serologically similar serovars belong to unrelated eBGs. These assignments to eBGs were supported by eBG-specific sequences of the flagellar gene
fliC
.
Glucosylation and Other Biotransformations of T-2 Toxin by Yeasts of the Trichomonascus Clade
Authors: McCormick Susan P. , Price Neil P. J. , Kurtzman Cletus P.
Published: October 2012

Trichothecenes are sesquiterpenoid toxins produced by
Fusarium
species. Since these mycotoxins are very stable, there is interest in microbial transformations that can remove toxins from contaminated grain or cereal products. Twenty-three yeast species assigned to the
Trichomonascus
clade (Saccharomycotina, Ascomycota), including four
Trichomonascus
species and 19 anamorphic species presently classified in
Blastobotrys
, were tested for their ability to convert the trichothecene T-2 toxin to less-toxic products. These species gave three types of biotransformations: acetylation to 3-acetyl T-2 toxin, glycosylation to T-2 toxin 3-glucoside, and removal of the isovaleryl group to form neosolaniol. Some species gave more than one type of biotransformation. Three
Blastobotrys
species converted T-2 toxin into T-2 toxin 3-glucoside, a compound that has been identified as a masked mycotoxin in
Fusarium
-infected grain. This is the first report of a microbial whole-cell method for producing trichothecene glycosides, and the potential large-scale availability of T-2 toxin 3-glucoside will facilitate toxicity testing and development of methods for detection of this compound in agricultural and other products.
Staphylococcus aureus CC398 Clade Associated with Human-to-Human Transmission
Authors: McCarthy Alex J. , van Wamel Willem , Vandendriessche Stien , Larsen Jesper , Denis Olivier , Garcia-Graells Cristina , Uhlemann Ann-Catrin , Lowy Franklin D. , Skov Robert , Lindsay Jodi A.
Published: October 2012

Staphylococcus aureus
clonal complex 398 (CC398) isolates colonize livestock and can spread to human contacts. Genetic analysis of isolates epidemiologically associated with human-to-human, but not livestock, transmission in multiple countries and continents identified a common clade that was negative for
tet
(M) and positive for bacteriophage ϕ3. Another group of human-to-human-transmitted isolates belonged to the common livestock-associated clade but had acquired a unique ϕ7 bacteriophage.
Biosynthesis and Functions of a Melanoid Pigment Produced by Species of the Sporothrix Complex in the Presence of
l
-Tyrosine
Authors: Almeida-Paes Rodrigo , Frases Susana , Araújo Glauber de Sousa , de Oliveira Manoel Marques Evangelista , Gerfen Gary J. , Nosanchuk Joshua D. , Zancopé-Oliveira Rosely Maria
Published: October 2012

Sporothrix schenckii
is the etiological agent of sporotrichosis, the main subcutaneous mycosis in Latin America. Melanin is an important virulence factor of
S. schenckii
, which produces dihydroxynaphthalene melanin (DHN-melanin) in conidia and yeast cells. Additionally,
l
-dihydroxyphenylalanine (
l
-DOPA) can be used to enhance melanin production on these structures as well as on hyphae. Some fungi are able to synthesize another type of melanoid pigment, called pyomelanin, as a result of tyrosine catabolism. Since there is no information about tyrosine catabolism in
Sporothrix
spp., we cultured 73 strains, including representatives of newly described
Sporothrix
species of medical interest, such as
S. brasiliensis
,
S. schenckii
, and
S. globosa
, in minimal medium with tyrosine. All strains but one were able to produce a melanoid pigment with a negative charge in this culture medium after 9 days of incubation. An
S. schenckii
DHN-melanin mutant strain also produced pigment in the presence of tyrosine. Further analysis showed that pigment production occurs in both the filamentous and yeast phases, and pigment accumulates in supernatants during stationary-phase growth. Notably, sulcotrione inhibits pigment production. Melanin ghosts of wild-type and DHN mutant strains obtained when the fungus was cultured with tyrosine were similar to melanin ghosts yielded in the absence of the precursor, indicating that this melanin does not polymerize on the fungal cell wall. However, pyomelanin-producing fungal cells were more resistant to nitrogen-derived oxidants and to UV light. In conclusion, at least three species of the
Sporothrix
complex are able to produce pyomelanin in the presence of tyrosine, and this pigment might be involved in virulence.
Genetic Diversity among Clostridium botulinum Strains Harboring
bont/A2
and
bont/A3
Genes
Authors: Lúquez Carolina , Raphael Brian H. , Joseph Lavin A. , Meno Sarah R. , Fernández Rafael A. , Maslanka Susan E.
Published: October 2012

Clostridium botulinum
type A strains are known to be genetically diverse and widespread throughout the world. Genetic diversity studies have focused mainly on strains harboring one type A botulinum toxin gene,
bont/A1
, although all reported
bont/A
gene variants have been associated with botulism cases. Our study provides insight into the genetic diversity of
C. botulinum
type A strains, which contain
bont/A2
(
n
= 42) and
bont/A3
(
n
= 4) genes, isolated from diverse samples and geographic origins. Genetic diversity was assessed by using
bont
nucleotide sequencing, content analysis of the
bont
gene clusters, multilocus sequence typing (MLST), and pulsed-field gel electrophoresis (PFGE). Sequences of
bont
genes obtained in this study showed 99.9 to 100% identity with other
bont/A2
or
bont/A3
gene sequences available in public databases. The neurotoxin gene clusters of the subtype A2 and A3 strains analyzed in this study were similar in gene content.
C. botulinum
strains harboring
bont/A2
and
bont/A3
genes were divided into six and two MLST profiles, respectively. Four groups of strains shared a similarity of at least 95% by PFGE; the largest group included 21 out of 46 strains. The strains analyzed in this study showed relatively limited genetic diversity using either MLST or PFGE.
Some Like It Hot: Evolution and Ecology of Novel Endosymbionts in Bat Flies of Cave-Roosting Bats (Hippoboscoidea, Nycterophiliinae)
Authors: Morse Solon F. , Dick Carl W. , Patterson Bruce D. , Dittmar Katharina
Published: October 2012

We investigated previously unknown associations between bacterial endosymbionts and bat flies of the subfamily Nycterophiliinae (Diptera, Streblidae). Molecular analyses revealed a novel clade ofGammaproteobacteriainNycterophiliabat flies. This clade was not closely related toArsenophonus-like microbes found in its sister genusPhalconomusand other bat flies. High population infection rates inNycterophiliaacross a wide geographic area, the presence of the symbionts in pupae, the general codivergence between hosts and symbionts, and high AT composition bias in symbiont genes together suggest that this host-symbiont association is obligate in nature and ancient in origin. SomeNycterophiliasamples (14.8%) also containedWolbachiasupergroup F (Alphaproteobacteria), suggesting a facultative symbiosis. Likelihood-based ancestral character mapping revealed that, initially, obligate symbionts exhibited association with host-specificNycterophiliabat flies that use a broad temperature range of cave environments for pupal development. As this mutualism evolved, the temperature range of bat flies narrowed to an exclusive use of hot caves, which was followed by a secondary broadening of the bat flies' host associations. These results suggest that the symbiosis has influenced the environmental tolerance of parasite life history stages. Furthermore, the contingent change to an expanded host range ofNycterophiliabat flies upon narrowing the ecological niche of their developmental stages suggests that altered environmental tolerance across life history stages may be a crucial factor in shaping parasite-host relationships.
Lipase and Protease Double-Deletion Mutant of Pseudomonas fluorescens Suitable for Extracellular Protein Production
Authors: Son Myunghan , Moon Yuseok , Oh Mi Jin , Han Sang Bin , Park Ki Hyun , Kim Jung-Gon , Ahn Jung Hoon
Published: October 2012

Pseudomonas fluorescens
, a widespread Gram-negative bacterium, is an ideal protein manufacturing factory (PMF) because of its safety, robust growth, and high protein production.
P. fluorescens
possesses a type I secretion system (T1SS), which mediates secretion of a thermostable lipase (TliA) and a protease (PrtA) through its ATP-binding cassette (ABC) transporter. Recombinant proteins in
P. fluorescens
are attached to the C-terminal signal region of TliA for transport as fusion proteins to the extracellular medium. However, intrinsic TliA from the
P. fluorescens
genome interferes with detection of the recombinant protein and the secreted recombinant protein is hydrolyzed, due to intrinsic PrtA, resulting in decreased efficiency of the PMF. In this research, the lipase and protease genes of
P. fluorescens
SIK W1 were deleted using the targeted gene knockout method. Deletion mutant
P. fluorescens
Δ
tliA
Δ
prtA
secreted fusion proteins without TliA or protein degradation. Using wild-type
P. fluorescens
as an expression host, degradation of the recombinant protein varied depending on the type of culture media and aeration; however, degradation did not occur with the
P. fluorescens
Δ
tliA
Δ
prtA
double mutant irrespective of growth conditions. By homologous expression of
tliA
and the ABC transporter in a plasmid, TliA secreted from
P. fluorescens
Δ
prtA
and
P. fluorescens
Δ
tliA
Δ
prtA
cells was found to be intact, whereas that secreted from the wild-type
P. fluorescens
and
P. fluorescens
Δ
tliA
cells was found to be hydrolyzed. Our results demonstrate that the
P. fluorescens
Δ
tliA
Δ
prtA
deletion mutant is a promising T1SS-mediated PMF that enhances production and detection of recombinant proteins in extracellular media.
Characterization of Two Virulent Phages of Lactobacillus plantarum
Authors: Briggiler Marcó Mariángeles , Garneau Josiane E. , Tremblay Denise , Quiberoni Andrea , Moineau Sylvain
Published: October 2012

We characterized two
Lactobacillus plantarum
virulent siphophages, ATCC 8014-B1 (B1) and ATCC 8014-B2 (B2), previously isolated from corn silage and anaerobic sewage sludge, respectively. Phage B2 infected two of the eight
L. plantarum
strains tested, while phage B1 infected three. Phage adsorption was highly variable depending on the strain used. Phage defense systems were found in at least two
L. plantarum
strains, LMG9211 and WCSF1. The linear double-stranded DNA genome of the
pac
-type phage B1 had 38,002 bp, a G+C content of 47.6%, and 60 open reading frames (ORFs). Surprisingly, the phage B1 genome has 97% identity with that of
Pediococcus damnosus
phage clP1 and 77% identity with that of
L. plantarum
phage JL-1; these phages were isolated from sewage and cucumber fermentation, respectively. The double-stranded DNA (dsDNA) genome of the
cos
-type phage B2 had 80,618 bp, a G+C content of 36.9%, and 127 ORFs with similarities to those of
Bacillus
and
Lactobacillus
strains as well as phages. Some phage B2 genes were similar to ORFs from
L. plantarum
phage LP65 of the
Myoviridae
family. Additionally, 6 tRNAs were found in the phage B2 genome. Protein analysis revealed 13 (phage B1) and 9 (phage B2) structural proteins. To our knowledge, this is the first report describing such high identity between phage genomes infecting different genera of lactic acid bacteria.
Distribution of the SELMA Translocon in Secondary Plastids of Red Algal Origin and Predicted Uncoupling of Ubiquitin-Dependent Translocation from Degradation
Authors: Stork Simone , Moog Daniel , Przyborski Jude M. , Wilhelmi Ilka , Zauner Stefan , Maier Uwe G.
Published: October 2012

Protein import into complex plastids of red algal origin is a multistep process including translocons of different evolutionary origins. The
s
ymbiont-derived
E
RAD-
l
ike
ma
chinery (SELMA), shown to be of red algal origin, is proposed to be the transport system for preprotein import across the periplastidal membrane of heterokontophytes, haptophytes, cryptophytes, and apicomplexans. In contrast to the canonical endoplasmic reticulum-associated degradation (ERAD) system, SELMA translocation is suggested to be uncoupled from proteasomal degradation. We investigated the distribution of known and newly identified SELMA components in organisms with complex plastids of red algal origin by intensive data mining, thereby defining a set of core components present in all examined organisms. These include putative pore-forming components, a ubiquitylation machinery, as well as a Cdc48 complex. Furthermore, the set of known 20S proteasomal components in the periplastidal compartment (PPC) of diatoms was expanded. These newly identified putative SELMA components, as well as proteasomal subunits, were
in vivo
localized as PPC proteins in the diatom
Phaeodactylum tricornutum
. The presented data allow us to speculate about the specific features of SELMA translocation in contrast to the canonical ERAD system, especially the uncoupling of translocation from degradation.
Iron-Inducible Nuclear Translocation of a Myb3 Transcription Factor in the Protozoan Parasite Trichomonas vaginalis
Authors: Hsu Hong-Ming , Lee Yu , Indra Dharmu , Wei Shu-Yi , Liu Hsing-Wei , Chang Lung-Chun , Chen Chinpan , Ong Shiou-Jeng , Tai Jung-Hsiang
Published: October 2012

In
Trichomonas vaginalis
, a novel nuclear localization signal spanning the folded R2R3 DNA-binding domain of a Myb2 protein was previously identified. To study whether a similar signal is used for nuclear translocation by other Myb proteins, nuclear translocation of Myb3 was examined in this report. When overexpressed, hemagglutinin-tagged Myb3 was localized to nuclei of transfected cells, with a cellular distribution similar to that of endogenous Myb3. Fusion to a bacterial tetracycline repressor, R2R3, of Myb3 that spans amino acids (aa) 48 to 156 was insufficient for nuclear translocation of the fusion protein, unless its C terminus was extended to aa 167. The conserved isoleucine in helix 2 of R2R3, which is important for Myb2's structural integrity in maintaining DNA-binding activity and nuclear translocation, was also vital for the former activity of Myb3, but less crucial for the latter. Sequential nuclear influx and efflux of Myb3, which require further extension of the nuclear localization signal to aa 180, were immediately induced after iron repletion. Sequence elements that regulate nuclear translocation with cytoplasmic retention, nuclear influx, and nuclear efflux were identified within the C-terminal tail. These results suggest that the R2R3 DNA-binding domain also serves as a common module for the nuclear translocation of both Myb2 and Myb3, but there are intrinsic differences between the two nuclear localization signals.
Deletion of the Fungal Gene
soft
Disrupts Mutualistic Symbiosis between the Grass Endophyte Epichloë festucae and the Host Plant
Authors: Charlton Nikki D. , Shoji Jun-Ya , Ghimire Sita R. , Nakashima Jin , Craven Kelly D.
Published: October 2012

Hyphal anastomosis, or vegetative hyphal fusion, establishes the interconnection of individual hyphal strands into an integrated network of a fungal mycelium. In contrast to recent advances in the understanding of the molecular basis for hyphal anastomosis, knowledge of the physiological role of hyphal anastomosis in the natural habitats of filamentous fungi is still very limited. To investigate the role of hyphal anastomosis in fungal endophyte-plant interactions, we generated mutant strains lacking the
Epichloë festucae soft
(
so
) gene, an ortholog of the hyphal anastomosis gene
so
in the endophytic fungus
E. festucae
. The
E. festucae
Δ
so
mutant strains grew similarly to the wild-type strain in culture but with reduced aerial hyphae and completely lacked hyphal anastomosis. The most striking phenotype of the
E. festucae
Δ
so
mutant strain was that it failed to establish a mutualistic symbiosis with the tall fescue plant host (
Lolium arundinaceum
); instead, it killed the host plant within 2 months after the initial infection. Microscopic examination revealed that the death of the tall fescue plant host was associated with the distortion and disorganization of plant cells. This study suggests that hyphal anastomosis may have an important role in the establishment/maintenance of fungal endophyte-host plant mutualistic symbiosis.
Roles of Hsl1p and Hsl7p in Swe1p Degradation: beyond Septin Tethering
Authors: King Kindra , Jin Michelle , Lew Daniel
Published: October 2012

The morphogenesis checkpoint in
Saccharomyces cerevisiae
couples bud formation to the cell division cycle by delaying nuclear division until cells have successfully constructed a bud. The cell cycle delay is due to the mitosis-inhibitory kinase Swe1p, which phosphorylates the cyclin-dependent kinase Cdc28p. In unperturbed cells, Swe1p is degraded via a mechanism thought to involve its tethering to a cortical scaffold of septin proteins at the mother-bud neck. In cells that experience stresses that delay bud formation, Swe1p is stabilized, accumulates, and promotes a G
2
delay. The tethering of Swe1p to the neck requires two regulators, called Hsl1p and Hsl7p. Hsl1p interacts with septins, and Hsl7p interacts with Swe1p; tethering occurs when Hsl1p interacts with Hsl7p. Here we created a version of Swe1p that is artificially tethered to the neck by fusion to a septin so that Swe1p no longer requires Hsl1p or Hsl7p for its localization to the neck. We show that the interaction between Hsl1p and Hsl7p, required for normal Swe1p degradation, is no longer needed for septin-Swe1p degradation, supporting the idea that the Hsl1p-Hsl7p interaction serves mainly to tether Swe1p to the neck. However, both Hsl1p and Hsl7p are still required for Swe1p degradation, implying that these proteins play additional roles beyond localizing Swe1p to the neck.
Pleiotropic Roles of the Msi1-Like Protein Msl1 in Cryptococcus neoformans
Authors: Yang Dong-Hoon , Maeng Shinae , Strain Anna K. , Floyd Anna , Nielsen Kirsten , Heitman Joseph , Bahn Yong-Sun
Published: October 2012

Msi1-like (MSIL) proteins contain WD40 motifs and have a pleiotropic cellular function as negative regulators of the Ras/cyclic AMP (cAMP) pathway and components of chromatin assembly factor 1 (CAF-1), yet they have not been studied in fungal pathogens. Here we identified and characterized an MSIL protein, Msl1, in
Cryptococcus neoformans
, which causes life-threatening meningoencephalitis in humans. Notably, Msl1 plays pleiotropic roles in
C. neoformans
in both cAMP-dependent and -independent manners largely independent of Ras. Msl1 negatively controls antioxidant melanin production and sexual differentiation, and this was repressed by the inhibition of the cAMP-signaling pathway. In contrast, Msl1 controls thermotolerance, diverse stress responses, and antifungal drug resistance in a Ras/cAMP-independent manner. Cac2, which is the second CAF-1 component, appears to play both redundant and distinct functions compared to the functions of Msl1. Msl1 is required for the full virulence of
C. neoformans
. Transcriptome analysis identified a group of Msl1-regulated genes, which include stress-related genes such as
HSP12
and
HSP78
. In conclusion, this study demonstrates pleiotropic roles of Msl1 in the human fungal pathogen
C. neoformans
, providing insight into a potential novel antifungal therapeutic target.
Rapid Triacylglycerol Turnover in Chlamydomonas reinhardtii Requires a Lipase with Broad Substrate Specificity
Authors: Li Xiaobo , Benning Christoph , Kuo Min-Hao
Published: October 2012

When deprived of nitrogen (N), the photosynthetic microalga
Chlamydomonas reinhardtii
accumulates large quantities of triacylglycerols (TAGs), making it a promising source of biofuel. Prominent transcriptional changes associated with the conditions leading to TAG accumulation have been found, suggesting that the key enzymes for TAG metabolism might be among those that fluctuate in their expression during TAG synthesis and breakdown. Using a
Saccharomyces cerevisiae
lipase null mutant strain for functional complementation, we identified the
CrLIP1
gene from
Chlamydomonas
based on its ability to suppress the lipase deficiency-related phenotypes of the yeast mutant. In
Chlamydomonas
, an inverse correlation was found between the
CrLIP1
transcript level and TAG abundance when
Chlamydomonas
cultures were reversibly deprived of N. The CrLIP1 protein expressed and purified from
Escherichia coli
exhibited lipolytic activity against diacylglycerol (DAG) and polar lipids. The lipase domain of CrLIP1 is most similar to two human DAG lipases, DAGLα and DAGLβ. The involvement of CrLIP1 in
Chlamydomonas
TAG hydrolysis was corroborated by reducing the abundance of the
CrLIP1
transcript with an artificial micro-RNA, which resulted in an apparent delay in TAG lipolysis when N was resupplied. Together, these data suggest that CrLIP1 facilitates TAG turnover in
Chlamydomonas
primarily by degrading the DAG presumably generated from TAG hydrolysis.
Roles of the Structural Symbiosis Polysaccharide (syp) Genes in Host Colonization, Biofilm Formation, and Polysaccharide Biosynthesis in Vibrio fischeri
Authors: Shibata Satoshi , Yip Emily S. , Quirke Kevin P. , Ondrey Jakob M. , Visick Karen L.
Published: October 2012

The symbiosis polysaccharide locus,syp, is required forVibrio fischerito form a symbiotic association with the squidEuprymna scolopes. It is also required for biofilm formation induced by the unlinked regulator RscS. Thesyplocus includes 18 genes that can be classified into four groups based on putative function: 4 genes encode putative regulators, 6 encode glycosyltransferases, 2 encode export proteins, and the remaining 6 encode proteins with other functions, including polysaccharide modification. To understand the roles of each of the 14 structuralsypgenes in colonization and biofilm formation, we generated nonpolar in-frame deletions of each gene. All of the deletion mutants exhibited defects in their ability to colonize juvenile squid, although the impact of the loss of SypB or SypI was modest. Consistent with their requirement for colonization, most of the structural genes were also required for RscS-induced biofilm formation. In particular, the production of wrinkled colonies, pellicles, and the matrix on the colony surface was eliminated or severely decreased in all mutants except for thesypBandsypImutants; in contrast, only a subset of genes appeared to play a role in attachment to glass. Finally, immunoblotting data suggested that the structural Syp proteins are involved in polysaccharide production and/or export. These results provide important insights into the requirements for thesypgenes under different environmental conditions and thus lay the groundwork for a more complete understanding of the matrix produced byV. fischerito enhance cell-cell interactions and promote symbiotic colonization.
Multiple Envelope Stress Response Pathways Are Activated in an Escherichia coli Strain with Mutations in Two Members of the DedA Membrane Protein Family
Authors: Sikdar Rakesh , Simmons Angelica R. , Doerrler William T.
Published: October 2012

We have reported that simultaneous deletion of two
Escherichia coli
genes,
yqjA
and
yghB
, encoding related and conserved inner membrane proteins belonging to the DedA protein family results in a number of intriguing phenotypes, including temperature sensitivity at 42°C, altered membrane lipid composition, and cell division defects. We sought to characterize these and other phenotypes in an effort to establish a function for this protein family in
E. coli
. Here, using reporter assays, we show that the major envelope stress response pathways Cpx, Psp, Bae, and Rcs are activated in strain BC202 (W3110; Δ
yqjA
Δ
yghB
) at the permissive growth temperature of 30°C. We previously demonstrated that 10 mM Mg
2+
, 400 mM NaCl, and overexpression of
tatABC
are capable of restoring normal growth to BC202 at elevated growth temperatures. Deletion of the
cpxR
gene from BC202 results in the loss of the ability of these supplements to restore growth at 42°C. Additionally, we report that the membrane potential of BC202 is significantly reduced and that cell division and growth can be restored either by expression of the multidrug transporter MdfA from a multicopy plasmid or by growth at pH 6.0. Together, these results suggest that the DedA family proteins YqjA and YghB are required for general envelope maintenance and homeostasis of the proton motive force under a variety of growth conditions.
The Zinc-Responsive Regulon of Neisseria meningitidis Comprises 17 Genes under Control of a Zur Element
Authors: Pawlik Marie-Christin , Hubert Kerstin , Joseph Biju , Claus Heike , Schoen Christoph , Vogel Ulrich
Published: October 2012

Zinc is a bivalent cation essential for bacterial growth and metabolism. The human pathogen
Neisseria meningitidis
expresses a homologue of the Zinc uptake regulator Zur, which has been postulated to repress the putative zinc uptake protein ZnuD. In this study, we elucidated the transcriptome of meningococci in response to zinc by microarrays and quantitative real-time PCR (qRT-PCR). We identified 15 genes that were repressed and two genes that were activated upon zinc addition. All transcription units (genes and operons) harbored a putative Zur binding motif in their promoter regions. A meningococcal Zur binding consensus motif (Zur box) was deduced
in silico
, which harbors a conserved central palindrome consisting of hexameric inverted repeats separated by three nucleotides (TGTTAT
DNH
ATAACA).
In vitro
binding of recombinant meningococcal Zur to this Zur box was shown for the first time using electrophoretic mobility shift assays. Zur binding to DNA depended specifically on the presence of zinc and was sensitive to mutations in the palindromic sequence. The Zur regulon among genes of unknown function comprised genes involved in zinc uptake, tRNA modification, and ribosomal assembly. In summary, this is the first study of the transcriptional response to zinc in meningococci.
Conjugal Transfer of a Virulence Plasmid in the Opportunistic Intracellular Actinomycete Rhodococcus equi
Authors: Tripathi V. N. , Harding W. C. , Willingham-Lane J. M. , Hondalus M. K.
Published: October 2012

Rhodococcus equi
is a facultative intracellular, Gram-positive, soilborne actinomycete which can cause severe pyogranulomatous pneumonia with abscessation in young horses (foals) and in immunocompromised people, such as persons with AIDS. All strains of
R. equi
isolated from foals and approximately a third isolated from humans contain a large, ∼81-kb plasmid which is essential for the intramacrophage growth of the organism and for virulence in foals and murine
in vivo
model systems. We found that the entire virulence plasmid could be transferred from plasmid-containing strains of
R. equi
(donor) to plasmid-free
R. equi
strains (recipient) at a high frequency and that plasmid transmission reestablished the capacity for intracellular growth in macrophages. Plasmid transfer required living cells and cell-to-cell contact and was unaffected by the presence of DNase, factors pointing to conjugation as the major means of genetic transfer. Deletion of a putative relaxase-encoding gene,
traA
, located in the proposed conjugative region of the plasmid, abolished plasmid transfer. Reversion of the
traA
mutation restored plasmid transmissibility. Finally, plasmid transmission to other
Rhodococcus
species and some additional related organisms was demonstrated. This is the first study showing a virulence plasmid transfer in
R. equi
, and it establishes a mechanism by which the virulence plasmid can move among bacteria in the soil.
Reexamining Transcriptional Regulation of the
Bacillus subtilis htpX
Gene and the
ykrK
Gene, Encoding a Novel Type of Transcriptional Regulator, and Redefining the YkrK Operator
Authors: Lin Ta-Hui , Huang Shih-Chien , Shaw Gwo-Chyuan
Published: October 2012

HtpX is an integral cytoplasmic membrane metalloprotease well conserved in numerous bacteria. A recent study showed that expression of the
Bacillus subtilis htpX
gene is under dual negative control by Rok and a novel type of transcriptional regulator, YkrK. Here we report that expression of the
B. subtilis htpX
gene is strongly heat inducible. Contrary to the previous prediction,
ykrK
expression has been found to be not subject to autoregulation. We have identified the
htpX
promoter and the authentic
ykrK
promoter, which is also distinct from the previously predicted one. We have redefined a conserved inverted repeat sequence to be the YkrK operator, which is somewhat different from the previously proposed one. We provide evidence that YkrK is not a substrate of HtpX and that heat induction of
htpX
is not YkrK mediated. We have also found that the absence of FtsH or HtpX alone did not impair
B. subtilis
cell viability on LB agar plates at high temperature, whereas the absence of both FtsH and HtpX caused a severe growth defect under heat stress. This finding supports the notion that FtsH and HtpX may have partially overlapping functions in heat resistance. Finally, we show that
htpX
expression is subject to transient negative control by
sigB
under heat stress in a Rok- and YkrK-independent manner. Triple negative control of
htpX
expression at high temperature by
rok
,
sigB
, and
ykrK
may help cells to prevent uncontrolled and detrimental oversynthesis of the HtpX protease.
New Substrates on the Block: Clinically Relevant Resistances for EmrE and Homologues
Authors: Nasie Iris , Steiner-Mordoch Sonia , Schuldiner Shimon
Published: October 2012

Transporters of the small multidrug resistance (SMR) family are small homo- or heterodimers that confer resistance to multiple toxic compounds by exchanging substrate with protons. Despite the wealth of biochemical information on EmrE, the most studied SMR member, a high-resolution three-dimensional structure is missing. To provide proteins that are more amenable to biophysical and structural studies, we identified and partially characterized SMR transporters from bacteria living under extreme conditions of temperature and radiation. Interestingly, these homologues as well as EmrE confer resistance to streptomycin and tobramycin, two aminoglycoside antibiotics widely used in clinics. These are hydrophilic and clinically important substrates of SMRs, and study of their mode of action should contribute to understanding the mechanism of transport and to combating the phenomenon of multidrug resistance. Furthermore, our study of one of the homologues, a putative heterodimer, supports the suggestion that in the SMR family, heterodimers can also function as homodimers.
Polymorphisms in Inc Proteins and Differential Expression ofincGenes among Chlamydia trachomatis Strains Correlate with Invasiveness and Tropism of Lymphogranuloma Venereum Isolates
Authors: Almeida Filipe , Borges Vítor , Ferreira Rita , Borrego Maria José , Gomes João Paulo , Mota Luís Jaime
Published: October 2012

Chlamydia trachomatisis a human bacterial pathogen that multiplies only within an intracellular membrane-bound vacuole, the inclusion.C. trachomatisincludes ocular and urogenital strains, usually causing infections restricted to epithelial cells of the conjunctiva and genital mucosa, respectively, and lymphogranuloma venereum (LGV) strains, which can infect macrophages and spread into lymph nodes. However,C. trachomatisgenomes display >98% identity at the DNA level. In this work, we studied whetherC. trachomatisInc proteins, which have a bilobed hydrophobic domain that may mediate their insertion in the inclusion membrane, could be a factor determining these different types of infection and tropisms. Analyses of polymorphisms and phylogeny of 48 Inc proteins from 51 strains encompassing the three disease groups showed significant amino acid differences that were mainly due to variations between Inc proteins from LGV and ocular or urogenital isolates. Studies of the evolutionary dynamics ofincgenes suggested that 10 of them are likely under positive selection and indicated that most nonsilent mutations are LGV specific. Additionally, real-time quantitative PCR analyses in prototype and clinical strains covering the three disease groups identified threeincgenes with LGV-specific expression. We determined the transcriptional start sites of these genes and found LGV-specific nucleotides within their promoters. Thus, subtle variations in the amino acids of a subset of Inc proteins and in the expression ofincgenes may contribute to the unique tropism and invasiveness ofC. trachomatisLGV strains.
Biophysical and Bioinformatic Analyses Implicate the Treponema pallidum Tp34 Lipoprotein (Tp0971) in Transition Metal Homeostasis
Authors: Brautigam Chad A. , Deka Ranjit K. , Ouyang Zhiming , Machius Mischa , Knutsen Gregory , Tomchick Diana R. , Norgard Michael V.
Published: October 2012

Metal ion homeostasis is a critical function of many integral and peripheral membrane proteins. The genome of the etiologic agent of syphilis,
Treponema pallidum
, is compact and devoid of many metabolic enzyme genes. Nevertheless, it harbors genes coding for homologs of several enzymes that typically require either iron or zinc. The product of the
tp0971
gene of
T. pallidum
, designated Tp34, is a periplasmic lipoprotein that is thought to be tethered to the inner membrane of this organism. Previous work on a water-soluble (nonacylated) recombinant version of Tp34 established that this protein binds to Zn
2+
, which, like other transition metal ions, stabilizes the dimeric form of the protein. In this study, we employed analytical ultracentrifugation to establish that four transition metal ions (Ni
2+
, Co
2+
, Cu
2+
, and Zn
2+
) readily induce the dimerization of Tp34; Cu
2+
(50% effective concentration [EC
50
] = 1.7 μM) and Zn
2+
(EC
50
= 6.2 μM) were the most efficacious of these ions. Mutations of the crystallographically identified metal-binding residues hindered the ability of Tp34 to dimerize. X-ray crystallography performed on crystals of Tp34 that had been incubated with metal ions indicated that the binding site could accommodate the metals examined. The findings presented herein, coupled with bioinformatic analyses of related proteins, point to Tp34's likely role in metal ion homeostasis in
T. pallidum
.
Genome Sequence of the Bacterium Streptomyces davawensis JCM 4913 and Heterologous Production of the Unique Antibiotic Roseoflavin
Authors: Jankowitsch Frank , Schwarz Julia , Rückert Christian , Gust Bertolt , Szczepanowski Rafael , Blom Jochen , Pelzer Stefan , Kalinowski Jörn , Mack Matthias
Published: October 2012

Streptomyces davawensis
JCM 4913 synthesizes the antibiotic roseoflavin, a structural riboflavin (vitamin B
2
) analog. Here, we report the 9,466,619-bp linear chromosome of
S. davawensis
JCM 4913 and a 89,331-bp linear plasmid. The sequence has an average G+C content of 70.58% and contains six rRNA operons (16S-23S-5S) and 69 tRNA genes. The 8,616 predicted protein-coding sequences include 32 clusters coding for secondary metabolites, several of which are unique to
S. davawensis
. The chromosome contains long terminal inverted repeats of 33,255 bp each and atypical telomeres. Sequence analysis with regard to riboflavin biosynthesis revealed three different patterns of gene organization in
Streptomyces
species. Heterologous expression of a set of genes present on a subgenomic fragment of
S. davawensis
resulted in the production of roseoflavin by the host
Streptomyces coelicolor
M1152. Phylogenetic analysis revealed that
S. davawensis
is a close relative of
Streptomyces cinnabarinus
, and much to our surprise, we found that the latter bacterium is a roseoflavin producer as well.
Bacteremic Pneumonia Caused by Extensively Drug-Resistant Streptococcus pneumoniae
Authors: Kang Cheol-In , Baek Jin Yang , Jeon Kyeongman , Kim So Hyun , Chung Doo Ryeon , Peck Kyong Ran , Lee Nam Yong , Song Jae-Hoon
Published: October 2012

The emergence of antimicrobial resistance threatens the successful treatment of pneumococcal infections. Here we report a case of bacteremic pneumonia caused by an extremely drug-resistant strain of
Streptococcus pneumoniae
, nonsusceptible to at least one agent in all classes but vancomycin and linezolid, posing an important new public health threat in our region.
Role of the SP2 Domain and Its Proteolytic Cleavage in HIV-1 Structural Maturation and Infectivity
Authors: de Marco Alex , Heuser Anke-Mareil , Glass Bärbel , Kräusslich Hans-Georg , Müller Barbara , Briggs John A. G.
Published: October 2012

HIV-1 buds as an immature, noninfectious virion. Proteolysis of its main structural component, Gag, is required for morphological maturation and infectivity and leads to release of four functional domains and the spacer peptides SP1 and SP2. The N-terminal cleavages of Gag and the separation of SP1 from CA are all essential for viral infectivity, while the roles of the two C-terminal cleavages and the role of SP2, separating the NC and p6 domains, are less well defined. We have analyzed HIV-1 variants with defective cleavage at either or both sites flanking SP2, or largely lacking SP2, regarding virus production, infectivity, and structural maturation. Neither the presence nor the proteolytic processing of SP2 was required for particle release. Viral infectivity was almost abolished when both cleavage sites were defective and severely reduced when the fast cleavage site between SP2 and p6 was defective. This correlated with an increased proportion of irregular core structures observed by cryo-electron tomography, although processing of CA was unaffected. Mutation of the slow cleavage site between NC and SP2 or deletion of most of SP2 had only a minor effect on infectivity and did not induce major alterations in mature core morphology. We speculate that not only separation of NC and p6 but also the processing kinetics in this region are essential for successful maturation, while SP2 itself is dispensable.
Complete Genome Sequence of a J Subgroup Avian Leukosis Virus Isolated from Local Commercial Broilers
Authors: Li Hongxin , Xue Chunyi , Ji Jun , Chang Shuang , Shang Huiqin , Zhang Lingjun , Ma Jingyun , Bi Yingzuo , Xie Qingmei
Published: October 2012

Subgroup J avian leukosis virus (ALV-J) isolate GDKP1202 was isolated from a 50-day-old local yellow commercial broiler in the Guangdong province of China in 2012. Here we report the complete genomic sequence of the GDKP1202 isolate, which caused high mortality, serious growth suppression, thymic atrophy, and liver enlargement in commercial broilers. A novel potential binding site (5′-GGCACCTCC-3′) for c-
myb
was identified in the GDKP1202 genome. These findings will provide additional insights into the molecular characteristics in the genomes and pathogenicity of ALV-J.
Complete Genome Sequence of an H9N2 Avian Influenza Virus Isolated from Egret in Lake Dongting Wetland
Authors: Wang Ba , Chen Quanjiao , Chen Ze
Published: October 2012

We isolated a recombinant H9N2 avian influenza virus (AIV) from fresh egret feces in the Ardeidae protection region of the Dongting Lake wetland area in China, and it was designated A/Egret/Hunan/1/2012(H9N2). This is the first report of isolating H9N2 AIV from wild birds in the Dongting Lake wetland. Its eight gene segments are generated by reassortment of gene segments of different AIV subtypes. These results are helpful for understanding the epidemiology and evolution of AIV in wild birds during migration.
Complete Genome Sequence Analysis of a Natural Reassortant Infectious Bursal Disease Virus in China
Authors: Chen Feng , Liu Jiajia , Yan Zhuanqiang , Liu Di , Ji Jun , Qin Jianping , Li Haiyan , Ma Jingyun , Bi Yingzuo , Xie Qingmei
Published: October 2012

A novel isolate of infectious bursal disease virus (IBDV) was designated GX-NN-L. The GX-NN-L IBDV was a very virulent infectious bursal disease virus (vvIBDV) isolated from broiler flocks in Guangxi province, China, in 2011. The GX-NN-L IBDV caused high mortality, immunosuppression, low weight gain, and bursal atrophy in commercial broilers. Here, we report the complete genome sequence of the GX-NN-L IBDV, a reassortment strain with segments A and B derived from very virulent strains and attenuated IBDV, respectively. These findings from this study provide additional insights into the genetic exchange between attenuated and very virulent strains of IBDV and continuous monitoring of the spread of the virus in chicken.
Complete Genome Sequences of Highly Divergent Torque Teno Virus Type I from Swine Herds
Authors: Wang Meng-meng , Zhou Yan-jun , Zhang Qing-zhan , Hou Jun-wei , Yu Hai , Tong Guang-zhi
Published: October 2012

Here, we report three complete genome sequences of porcine torque teno virus type I (TTV1) which were obtained from swine tissues and sera from southern China through routine and nested PCR amplification and characterized together with other genome sequences already deposited in GenBank. The results showed that the TTV1 sequences were highly divergent and could be divided into 1a and 1b subtypes.
Complete Genome Sequence of the Porcine Kobuvirus Variant CH/HNXX-4/2012
Authors: Cao Weijun , Zheng Haixue , Zhang Keshan , Jin Ye , Lv Lv , Yang Fan , Liu Xiangtao
Published: October 2012

Porcine kobuvirus, an emerging virus, may be the underlying etiological cause of a large-scale outbreak of diarrhea in suckling piglets in China that started in 2010. We report the complete genome sequence of the porcine kobuvirus variant CH/HNXX-4/2012 with a 30-amino-acid deletion in its 2B-coding region that was isolated in this outbreak. This will help the phenotypic variation and evolutionary characteristics of porcine kobuvirus to be understood.
Complete Genomic Sequence of a Novel Reassortant H11N3 Influenza Virus Isolated from Domestic Ducks in Jiangsu, China
Authors: Chen Chaoyang , Zhao Guo , Gu Xiaobing , Gu Min , Hu Jiao , Li Qunhui , Zhao Qingqing , Wang Xiaoquan , Liu Xiaowen , Liu Xiufan
Published: October 2012

For the first time we report the complete genomic sequence of an H11N3 influenza virus from domestic ducks in China. Phylogenetic analysis showed that the H11N3 virus was a novel reassortant with its genes from different subtypes of domestic duck-origin avian influenza viruses, which further underlined that domestic ducks play a key role in the genetic reassortment and evolution of influenza viruses in China.
Complete Genome Sequences of a Novel Reassortant H4N2 Avian Influenza Virus Isolated from a Live Poultry Market in Eastern China
Authors: Teng Qiaoyang , Ji Xiwen , Li Guoxin , Li Xuesong , Li Zejun
Published: October 2012

A/duck/Shanghai/28-1/2009(H4N2) (DK28) was isolated from a live poultry market in Shanghai, China. Using PCR and sequencing analysis, we obtained the complete genome sequences of the DK28 virus. The sequence analysis demonstrated that this H4N2 virus was a novel multiple-gene reassortant avian influenza virus (AIV) whose genes originated from H1N1, H1N3, H3N3, H4N2, and H4N6. Knowledge regarding the complete genome sequences of the DK28 virus will be useful for epidemiological surveillance.
EMPIRICAL IMPLICATIONS FOR PROMOTING STUDENTS' ENTREPRENEURIAL INTENTIONS
Authors: TEGTMEIER SILKE
Published: October 2012

Ajzen's Theory of Planned Behaviour (TPB) suggests empirical implications to promote students' entrepreneurial intentions. In the course of an empirical study with 208 German students, regression analyses confirm that attitude, social norm, and perceived behavioural control contribute substantially to the prediction of start-up intentions. The use of index based measurements shows that entrepreneurial intention is significantly influenced by all three constructs (R2= 0.446). As beliefs based on information and experiences are decisive in order to emerge an intention, a promoting approach should start here. Following the TPB, changes at singular points are not sufficient. A discussion of concrete beliefs provides implications for future research as well as for practical interventions.
Optimizing matrix transposes using a POWER7 cache model and explicit prefetching
Authors: Mateescu Gabriel , Bauer Gregory H. , Fiedler Robert A.
Published: October 2012

We consider the problem of efficiently computing matrix transposes on the POWER7 architecture. We develop a matrix transpose algorithm that uses cache blocking, cache prefetching and data alignment. We model the POWER7 data cache and memory concurrency and use the model to predict the memory throughput of the proposed matrix transpose algorithm. The performance of our matrix transpose algorithm is up to five times higher than that of the dgetmo routine of the Engineering and Scientific Subroutine Library and is 2.5 times higher than that of the code generated by compiler-inserted prefetching. Numerical experiments indicate a good agreement between the predicted and the measured memory throughput.
Benchmarking and modeling disk-based storage tiers for practical storage design
Authors: Lee DongJin , O'Sullivan Michael , Walker Cameron
Published: October 2012

This paper investigates benchmarking and modeling for a disk-based storage system in order to design and build a practical storage tier. As a practical case study, we focus on the design of an archival storage tier. The archival tiers play a critical role in data preservation as almost all current data will eventually be archived and the demands placed on archival tiers are growing because of large regularly-scheduled back-ups. Archival tiers usually consist of tape-based devices with a large storage capacity, but limited I/O performance for retrieving data, especially when multiple retrieval requests are made simultaneously. As the cost of disk-based devices continues to decrease while the capacity of individual disks increases, disk-based systems are becoming a more realistic option for both enterprise and commodity archival storage tiers.

We utilize archival workloads developed from an analysis of historical data in order to provide accurate and robust benchmarks of system performance as an archive. We then embed our practical measurements in a measurementdriven optimization approach to design an archival system. Our approach produces a low cost design for a commodity disk-based archival storage system. Using our measurementdriven model, ideal storage building blocks are identified for a real-world archival tier design.