bluuebunny/crossref_metadata_2025_split huggingface dataset:
Effects of Interleukin-1 Receptor Antagonist Overexpression on Infection byListeria monocytogenes
Authors: Irikura Vera M. , Hirsch Emmet , Hirsh David 
Published: March 2020

ABSTRACTInterleukin-1 receptor antagonist (IL-1ra) is a naturally occurring cytokine whose only known function is the inhibition of interleukin-1 (IL-1). Using a reverse genetic approach in mice, we previously showed that increasing IL-1ra gene dosage leads to reduced survival of a primary listerial infection. In this study, we characterize further the role of endogenously produced IL-1ra and, by inference, IL-1 in murine listeriosis. IL-1ra overexpression inhibits, but does not eliminate, primary immune responses, reducing survival and increasing bacterial loads in the target organs. We demonstrate that IL-1ra functions in the innate immune response to regulate the peak leukocyte levels in the blood, the accumulation of leukocytes at sites of infection, and the activation of macrophages during a primary infection. Reduced macrophage class II major histocompatibility complex expression was observed despite increased gamma interferon (IFN-γ) levels, suggesting that IL-1 activity is essential along with IFN-γ for macrophage activation in vivo. We also show that IL-1ra plays a more limited role during secondary listeriosis, blunting the strength of the delayed-type hypersensitivity response to listerial antigen while not significantly altering cellular immunity to a second infectious challenge. When these results are compared to those for other mutant mice, IL-1ra appears to be unique among the cytokines studied to date in its regulation of leukocyte migration during primary listeriosis.
Characterization of the Gene Encoding a 26-Kilodalton Protein (OMP26) from Nontypeable
Haemophilus influenzae
and Immune Responses to the Recombinant Protein
Authors: El-Adhami Wafa , Kyd Jennelle M. , Bastin David A. , Cripps Allan W. 
Published: March 2020

ABSTRACT

A 26-kDa protein (OMP26) isolated and purified from nontypeable
Haemophilus influenzae
(NTHI) strain 289 has been shown to enhance clearance of infection following pulmonary challenge with NTHI in rats. DNA sequence analysis revealed that it was 99% identical to a gene encoding a cell envelope protein of the
H. influenzae
Rd strain (TIGR accession no. HI0916). The deduced amino acid sequence revealed a hydrophilic polypeptide rich in basic amino acids. Restriction fragment length polymorphism analysis suggested that the OMP26 gene was relatively conserved among isolates of NTHI. Analysis of the deduced amino acid sequence of the OMP26 gene from 20 different isolates showed that similarity with NTHI-289 ranged from 96.5% (1 isolate) to 99.5% (14 isolates). Two recombinant forms of OMP26, a full length 28-kDa protein (equivalent to preprotein) and a 26-kDa protein lacking a 23-amino-acid leader peptide (equivalent to processed protein), were assessed in immunization studies for the ability to induce an immune response that would be as effective as the native protein in enhancing the clearance of NTHI following pulmonary challenge in rats. Immunization with the recombinant protein that included the leader peptide was more effective in enhancing pulmonary clearance, and it induced a better cell-mediated response and higher titers of systemic and mucosal antibody. This study has characterized a 26-kDa protein from NTHI that shows significant potential as a vaccine candidate.
Vaccination with Cathepsin L Proteinases and with Leucine Aminopeptidase Induces High Levels of Protection against Fascioliasis in Sheep
Authors: Piacenza Lucía , Acosta Daniel , Basmadjian Isabel , Dalton John P. , Carmona Carlos 
Published: March 2020

ABSTRACTThe potential of different parasite proteinases for use as vaccine candidates against fascioliasis in sheep was studied by vaccinating animals with the cathepsin L proteinases CL1 and CL2 and with leucine aminopeptidase (LAP) purified from adult flukes. In the first trial, sheep were immunized with CL1 or CL2 and the mean protection levels obtained were 33 and 34%, respectively. Furthermore, a significant reduction in egg output was observed in sheep vaccinated either with CL1 (71%) or with CL2 (81%). The second trial was performed to determine the protective potential of the two cathepsin L proteinases assayed together, as well as in combination with LAP, and of LAP alone. The combination of CL1 and CL2 induced higher levels of protection (60%) than those produced when these enzymes were administered separately. Those sheep that received the cocktail vaccine including CL1, CL2, and LAP were significantly protected (78%) against metacercarial challenge, but vaccination with LAP alone elicited the highest level of protection (89%). All vaccine preparations induced high immunoglobulin G titers which were boosted after the challenge infection, but no correlations between antibody titers and worm burdens were found. However, the sera of those animals vaccinated with LAP contained LAP-neutralizing antibodies. Reduced liver damage, as assessed by the level of the liver enzyme gamma-glutamyl transferase, was observed in the groups vaccinated with CL1, CL2, and LAP or with LAP alone.
Genetic Control of Experimental Lyme Arthritis in the Absence of Specific Immunity
Authors: Brown Charles R. , Reiner Steven L. 
Published: March 2020

ABSTRACTHost genetics play an important role in determining resistance or susceptibility to experimental Lyme arthritis. While specific immunity appears to regulate disease resolution, innate immunity appears to regulate disease severity. Intradermal infection withBorrelia burgdorferiyields severe arthritis in C3H/He (C3H) mice but only minimal arthritis in BALB/c mice. Intradermal infection of immunodeficient C3H SCID mice also results in severe arthritis, but arthritis of only moderate severity in BALB/c SCID mice. In the present study, we examined immunodeficient recombinase-activating gene-knockout (RAG-1−/−) (RAG−) mice from resistant C57BL/6 (B6) and DBA/2 (DBA) mouse strains.B. burgdorferi-infected B6 RAG−and DBA RAG−mice had little or no ankle swelling, a low occurrence of inflammatory infiltrates in tibiotarsal joints, and low arthritis severity scores in comparison to RAG+and RAG−BALB/c or C3H mice. Few differences in spirochete DNA levels in ankles of resistant and susceptible RAG−mice were seen. These data suggest that resistance to arthritis development followingB. burgdorferiinfection is not necessarily dependent on an acquired immune response and can occur despite the presence of high spirochete burden. Thus, genes expressed outside the specific immune response can be central regulators of experimental arthritis.
Salmonella typhimuriumEncodes a Putative Iron Transport System within the Centisome 63 Pathogenicity Island
Authors: Zhou Daoguo , Hardt Wolf-Dietrich , Galán Jorge E. 
Published: March 2020

ABSTRACTUpon entry into the host,Salmonella entericastrains are presumed to encounter an iron-restricted environment. Consequently, these bacteria have evolved a variety of often-redundant high-affinity acquisition systems to obtain iron in this restricted environment. We have identified an iron transport system that is encoded within the centisome 63 pathogenicity island ofSalmonella typhimurium. The nucleotide composition of this locus is significantly different from that of the rest of this pathogenicity island, suggesting a different ancestry and a mosaic structure for this region of theS. typhimuriumchromosome. This locus, designatedsit, consists of four open reading frames which encode polypeptides with extensive homology to theyfeABC iron transport system ofYersinia pestis, as well as other ABC transporters. ThesitAgene encodes a putative periplasmic binding protein,sitBencodes an ATP-binding protein, andsitCandsitDencode two putative permeases (integral membrane proteins). This operon is capable of complementing the growth defect of the enterobactin-deficientEscherichia colistrain SAB11 in iron-restricted minimal medium. Transcription of thesitoperon is repressed under iron-rich growth conditions in afur-dependent manner. Introduction of asitBCDdeletion into wild-typeS. typhimuriumresulted in no apparent growth defect in either nutrient-rich or minimal medium and no measurable virulence phenotype. These results further support the existence of redundant iron uptake systems inS. enterica.
Segmented Filamentous Bacteria Are Potent Stimuli of a Physiologically Normal State of the Murine Gut Mucosal Immune System
Authors: Talham Gwen L. , Jiang Han-Qing , Bos Nicolaas A. , Cebra John J. 
Published: March 2020

ABSTRACTSegmented filamentous bacteria (SFB) are autochthonous bacteria inhabiting the intestinal tracts of many species, including humans. We studied the effect of SFB on the mucosal immune system by monoassociating formerly germfree C3H/HeN mice with SFB. At various time points during 190 days of colonization, fragment cultures of small intestine and Peyer’s patches (PP) were analyzed for total immunoglobulin A (IgA) and SFB-specific IgA production. Also, phenotypic changes indicating germinal center reactions (GCRs) and the activation of CD4+T cells in PP were determined by using fluorescence-activated cell sorter analyses. A second group of SFB-monoassociated mice was colonized with a gram-negative commensal,Morganella morganii, to determine if the mucosal immune system was again stimulated and to evaluate the effect of prior colonization with SFB on the ability ofM. morganiito translocate to the spleen and mesenteric lymph nodes. We found that SFB stimulated GCRs in PP from day 6 after monoassociation, that GCRs only gradually waned over the entire length of colonization, that natural IgA production was increased to levels 24 to 63% of that of conventionally reared mice, and that SFB-specific IgA was produced but accounted for less than 1.4% of total IgA. Also, the proportion of CD4+, CD45RBlowT cells, indicative of activated cells, gradually increased in the PP to the level found in conventionally reared mice. Secondary colonization withM. morganiiwas able to stimulate GCRs anew, leading to a specific IgA antibody response. Previous stimulation of mucosal immunity by SFB did not prevent the translocation ofM. morganiiin the double-colonized mice. Our findings generally indicate that SFB are one of the single most potent microbial stimuli of the gut mucosal immune system.
Transcriptional Analysis of the
Staphylococcus aureus
Penicillin Binding Protein 2 Gene
Authors: Pinho Mariana G. , de Lencastre Herminia , Tomasz Alexander 
Published: March 2020

ABSTRACT

Sequencing of the vicinity of the staphylococcal
pbp2
gene and transcriptional analysis by primer extension and promoter fusions were used to show that
pbp2
is part of an operon that also includes a gene with high homology to
prfA
of
Bacillus subtilis
. Two distinct promoters were identified directing transcription of
pbp2
either alone or together with
prfA
. It was recently reported that transposon inactivation of
pbp2
causes a reduction in methicillin resistance, but complementation experiments were not fully successful. We now show that introduction of the intact
pbp2
gene with its two newly identified promoters into the chromosome of the transposon mutant resulted in the full recovery of high-level methicillin resistance.
Expression of
lacZ
from the Promoter of the
Escherichia coli spc
Operon Cloned into Vectors Carrying the W205
trp-lac
Fusion
Authors: Liang Sung-Tzu , Dennis Patrick P. , Bremer Hans 
Published: March 2020

ABSTRACT

The expression of
lacZ
has been analyzed and compared in a series of promoter cloning vectors by measuring the amount of
lacZ
mRNA by hybridization and the amount of β-galactosidase by standard enzymatic assay. Expression was driven by the promoter, P

spc

, of the
spc
ribosomal protein operon. The vectors contained either the standard W205
trp-lac
fusion with the
trp
operon transcription terminator,
trpt
, located in the
lacZ
leader sequence, or a deletion derivative that functionally inactivates
trpt
. In the presence of
trpt
,
lacZ
expression was temperature dependent so that increasing the growth temperature reduced the accumulation of
lacZ
mRNA and β-galactosidase activity. The frequency of transcript termination at
trpt
was estimated to be near zero at 20°C and at about 45% at 37°C. The amount of P

spc

-derived
lacZ
mRNA and the amount of β-galactosidase produced per
lacZ
mRNA varied, depending on the mRNA 5′ leader sequence between P

spc

and
lacZ
. These results demonstrate that the quantitative assessment of promoter activities with promoter cloning vectors requires careful analysis and interpretation. One particular construct without
trpt
did not seem to contain fortuitous transcription or translation signals generated at the fusion junction. In this strain,
lacZ
expression from P

spc

was compared at the enzyme activity and mRNA levels with a previously constructed strain in which
lacZ
was linked to the tandem P1 and P2 promoters of the
rrnB
operon. At any given growth rate, the different activities of β-galactosidase in these two strains were found to reflect the same differences in their amounts of
lacZ
mRNA. Assuming that the promoter-
lacZ
fusions in these strains reflect the properties of the promoters in their normal chromosomal setting, it was possible to estimate the absolute transcription activity of P

spc

and the relative translation efficiency of P

spc

-
lacZ
mRNA at different growth rates. Transcription from the
spc
promoter was found to increase from about 10 transcripts per min at a growth rate of 1.0 doublings/h to a maximum plateau of about 23 transcripts per min at growth rates above 1.5 doublings/h. The translation frequency of
lacZ
mRNA expressed from P

spc

was unaffected by growth rates.
Genetic and Sequence Analysis of the pTiC58
trb
Locus, Encoding a Mating-Pair Formation System Related to Members of the Type IV Secretion Family
Authors: Li Pei-Li , Everhart Dawn M. , Farrand Stephen K. 
Published: March 2020

ABSTRACT

Conjugal transfer of pTiC58 requires two regions,
tra
which contains the
oriT
and several genes involved in DNA processing and a region of undefined size and function that is located at the 2-o’clock position of the plasmid. Using transposon mutagenesis with Tn
3
HoHo1 and a binary transfer system, we delimited this second region, called
trb
, to an 11-kb interval between the loci for vegetative replication and nopaline catabolism. DNA sequence analysis of this region identified 13 significant open reading frames (ORFs) spanning 11,003 bp. The first, encoding
traI
, already has been described and is responsible for the synthesis of
Agrobacterium
autoinducer (AAI) (I. Hwang, P.-L. Li, L. Zhang, K. R. Piper, D. M. Cook, M. E. Tate, and S. K. Farrand, Proc. Natl. Acad. Sci. USA 91:4639–4643, 1994). Translation products of the next 11 ORFs showed similarities to those of
trbB
,
-C
,
-D
,
-E
,
-J
,
-K
,
-L
,
-F
,
-G
,
-H
, and
-I
of the
trb
region of the octopine-type Ti plasmid pTi15955 and of the
tra2
core region of RP4. In RP4, these genes encode mating-pair formation functions and are essential for the conjugal transfer of the IncP plasmid. Each of the
trb
gene homologues is oriented counterclockwise on the Ti plasmid. Expression of these genes, as measured by using the
lacZ
fusions formed by Tn
3
HoHo1, required the
traI
promoter and the transcriptional activator TraR along with its coinducer, AAI. While related to that of RP4, the
trb
system of pTiC58 did not allow propagation of the
trb
-specific bacteriophages PRD1, PRR1, and Pf3. The products of several
trb
genes of the Ti plasmid are similar to those of other loci that encode DNA transfer or protein secretion systems, all of which are members of the type IV secretion family.
The
facC
Gene of
Aspergillus nidulans
Encodes an Acetate-Inducible Carnitine Acetyltransferase
Authors: Stemple Christopher J. , Davis Meryl A. , Hynes Michael J. 
Published: March 2020

ABSTRACT

Mutations in the
facC
gene of
Aspergillus nidulans
result in an inability to use acetate as a sole carbon source. This gene has been cloned by complementation. The proposed translation product of the
facC
gene has significant similarity to carnitine acetyltransferases (CAT) from other organisms. Total CAT activity was found to be inducible by acetate and fatty acids and repressed by glucose. Acetate-inducible activity was found to be absent in
facC
mutants, while fatty acid-inducible activity was absent in an
acuJ
mutant. Acetate induction of
facC
expression was dependent on the
facB
regulatory gene, and an expressed FacB fusion protein was demonstrated to bind to 5′
facC
sequences. Carbon catabolite repression of
facC
expression was affected by mutations in the
creA
gene and a CreA fusion protein bound to 5′
facC
sequences. Mutations in the
acuJ
gene led to increased acetate induction of
facC
expression and also of an
amdS-lacZ
reporter gene, and it is proposed that this results from accumulation of acetate, as well as increased expression of
facB
. A model is presented in which
facC
encodes a cytosolic CAT enzyme, while a different CAT enzyme, which is
acuJ
dependent, is present in peroxisomes and mitochondria, and these activities are required for the movement of acetyl groups between intracellular compartments.
Characterization of Mutations That Allow p -Aminobenzoyl-Glutamate Utilization by
Escherichia coli
Authors: Hussein Mouyassar J. , Green Jacalyn M. , Nichols Brian P. 
Published: March 2020

ABSTRACT

An
Escherichia coli
strain deficient in
p
-aminobenzoate synthesis was mutagenized, and derivatives were selected for growth on folic acid. Supplementation was shown to be due to
p
-aminobenzoyl-glutamate present as a breakdown product in commercial folic acid preparations. Two classes of mutations characterized by the minimum concentration of
p
-aminobenzoyl-glutamate that could support growth were obtained. Both classes of mutations were genetically and physically mapped to about 30 min on the
E. coli
chromosome. A cloned wild-type gene from this region,
abgT
(formerly
ydaH
) could confer a similar
p
-aminobenzoyl-glutamate utilization phenotype on the parental strain. Interruption of
abgT
on the plasmid or on the chromosome of the mutant strain resulted in a loss of the phenotype.
abgT
was the third gene in an apparent operon containing
abgA
,
abgB
,
abgT
, and possibly
ogt
and might be regulated by a divergently transcribed LysR-type regulator encoded by
abgR
. Two different single-base-pair mutations that gave rise to the
p
-aminobenzoyl-glutamate utilization phenotype lay in the
abgR-abgA
intercistronic region and appeared to allow the expression of
abgT
. The second class of mutation was due to a tandem duplication of
abgB
and
abgT
fused to
fnr
. The
abgA
and
abgB
gene products were homologous to one another and to a family of aminoacyl aminohydrolases.
p
-Aminobenzoyl-glutamate hydrolysis could be detected in extracts from several of the mutant strains, but intact
abgA
and
abgB
were not essential for
p
-aminobenzoyl-glutamate utilization when
abgT
was supplied in
trans
.
Sister Chromatid Exchange Frequencies in
Escherichia coli
Analyzed by Recombination at the
dif
Resolvase Site
Authors: Steiner Walter W. , Kuempel Peter L. 
Published: March 2020

ABSTRACT

Sister chromatid exchange (SCE) in
Escherichia coli
results in the formation of circular dimer chromosomes, which are converted back to monomers by a compensating exchange at the
dif
resolvase site. Recombination at
dif
is site specific and can be monitored by utilizing a density label assay that we recently described. To characterize factors affecting SCE frequency, we analyzed dimer resolution at the
dif
site in a variety of genetic backgrounds and conditions. Recombination at
dif
was increased by known hyperrecombinogenic mutations such as
polA
,
dut
, and
uvrD
. It was also increased by a
fur
mutation, which increased oxidative DNA damage. Recombination at
dif
was eliminated by a
recA
mutation, reflecting the role of RecA in SCE and virtually all homologous recombination in
E. coli
. Interestingly, recombination at
dif
was reduced to approximately half of the wild-type levels by single mutations in either
recB
or
recF
, and it was virtually eliminated when both mutations were present. This result demonstrates the importance of both RecBCD and RecF to chromosomal recombination events in wild-type cells.
RecA-Independent Pathways of Lambdoid Prophage Induction in
Escherichia coli
Authors: Rozanov Dmitry V. , D’Ari Richard , Sineoky Sergey P. 
Published: March 2020

ABSTRACT

Two
Escherichia coli
genes, expressed from multicopy plasmids, are shown to cause partial induction of prophage λ in
recA
mutant lysogens. One is
rcsA
, which specifies a positive transcriptional regulator of the
cps
genes, which are involved in capsular polysaccharide synthesis. The other is
dsrA
, which specifies an 85-nucleotide RNA that relieves repression of the
rcsA
gene by histone-like protein H-NS. Genetic contexts known to increase Cps expression also cause RecA-independent λ induction: the
rcsC137
mutation, which causes constitutive Cps expression, and the
lon
and
rcsA3
mutations, which stabilize RcsA. Lambdoid phages 21, φ80, and 434 are also induced by RcsA and DsrA overexpression in
recA
lysogens. Excess λ cI repressor specifically blocks λ induction, suggesting that induction involves repressor inactivation rather than repressor bypass. RcsA-mediated induction requires RcsB, the known effector of the
cps
operon, whereas DsrA-mediated induction is RcsB independent in stationary phase, pointing to the existence of yet another RecA-independent pathway of prophage induction.
Regulation of an Osmoticum-Responsive Gene in
Anabaena
sp. Strain PCC 7120
Authors: Schwartz Steven H. , Black Todd A. , Jäger Karin , Panoff Jean-Michel , Wolk C. Peter 
Published: March 2020

ABSTRACT

Salt-induced genes in the cyanobacterium
Anabaena
sp. strain PCC 7120 were identified by use of a Tn
5
-based transposon bearing
luxAB
as a reporter. The genomic sequence adjacent to one site of insertion of the transposon was identical in part to the sequence of the
lti2
gene, which was previously identified in a differential screen for cold-induced transcripts in
Anabaena variabilis
. The
lti2
-like gene was induced by sucrose and other osmotica and by low temperature, in addition to salt. Regulatory components necessary for the induction of this gene by osmotica were sought by a further round of transposon mutagenesis. One mutant that displayed reduced transcriptional activity of the
lti2
-like gene in response to exposure to osmotica had an insertion in an open reading frame, which was denoted
orrA
, whose predicted product showed sequence similarity to response regulators from two-component regulatory systems. The corresponding mutation was reconstructed and was shown, like the second-site transposon mutation, to result in reduced response to osmotic stress. Induction of the
lux
reporter gene by osmotica was restored by complementation with a genomic fragment containing the entire open reading frame for the presumptive response regulator, whereas a fragment containing a truncated copy of the open reading frame for the response regulator did not complement the mutation.