bluuebunny/crossref_metadata_2025_split huggingface dataset:
Gliding Mutants of
Myxococcus xanthus
with High Reversal Frequencies and Small Displacements
Authors: Spormann Alfred M. , Kaiser Dale 
Published: December 2019

ABSTRACT

Myxococcus xanthus
cells move on a solid surface by gliding motility. Several genes required for gliding motility have been identified, including those of the A- and S-motility systems as well as the
mgl
and
frz
genes. However, the cellular defects in gliding movement in many of these mutants were unknown. We conducted quantitative, high-resolution single-cell motility assays and found that mutants defective in
mglAB
or in
cglB
, an A-motility gene, reversed the direction of gliding at frequencies which were more than 1 order of magnitude higher than that of wild type cells (2.9 min
−1
for Δ
mglAB
mutants and 2.7 min
−1
for
cglB
mutants, compared to 0.17 min
−1
for wild-type cells). The average gliding speed of Δ
mglAB
mutant cells was 40% of that of wild-type cells (on average 1.9 μm/min for Δ
mglAB
mutants, compared to 4.4 μm/min for wild-type cells). The
mglA
-dependent reversals and gliding speeds were dependent on the level of intracellular MglA protein:
mglB
mutant cells, which contain only 15 to 20% of the wild-type level of MglA protein, glided with an average reversal frequency of about 1.8 min
−1
and an average speed of 2.6 μm/min. These values range between those exhibited by wild-type cells and by Δ
mglAB
mutant cells. Epistasis analysis of
frz
mutants, which are defective in aggregation and in single-cell reversals, showed that a
frzD
mutation, but not a
frzE
mutation, partially suppressed the
mglA
phenotype. In contrast to
mgl
mutants,
cglB
mutant cells were able to move with wild-type speeds only when in close proximity to each other. However, under those conditions, these mutant cells were found to glide less often with those speeds. By analyzing double mutants, the high reversing movements and gliding speeds of
cglB
cells were found to be strictly dependent on type IV pili, encoded by S-motility genes, whereas the high-reversal pattern of
mglAB
cells was only partially reduced by a
pilR
mutation. These results suggest that the MglA protein is required for both control of reversal frequency and gliding speed and that in the absence of A motility, type IV pilus-dependent cell movement includes reversals at high frequency. Furthermore,
mglAB
mutants behave as if they were severely defective in A motility but only partially defective in S motility.
Localization of
Escherichia coli rpoC
Mutations That Affect RNA Polymerase Assembly and Activity at High Temperature
Authors: Nedea Eduard C. , Markov Dmitriy , Naryshkina Tatyana , Severinov Konstantin 
Published: December 2019

ABSTRACT

We localized five
rpoC
(β′) mutations affecting
Escherichia coli
RNA polymerase assembly. The Ts4, XH56, and R120 mutations changed β′ residues conserved throughout eubacteria; the JE10092 mutation occurred in the hypervariable region;
rpoC1
(TsX) changed a universally conserved residue and corresponds to yeast
rpb1-1
. Thus, distinct, predominantly conserved β′ residues participate in interactions holding RNA polymerase together.
The
Escherichia coli
Homologue of Yeast Rer2, a Key Enzyme of Dolichol Synthesis, Is Essential for Carrier Lipid Formation in Bacterial Cell Wall Synthesis
Authors: Kato Jun-ichi , Fujisaki Shingo , Nakajima Ken-ichi , Nishimura Yukinobu , Sato Miyuki , Nakano Akihiko 
Published: December 2019

ABSTRACT

We found in the
Escherichia coli
genome sequence a homologue of
RER2
, a
Saccharomyces cerevisiae
gene required for proper localization of an endoplasmic reticulum protein, and designated it
rth
(
RER2
homologue). The disruption of this gene was lethal for
E. coli
. To reveal its biological function, we isolated temperature-sensitive mutants of the
rth
gene. The mutant cells became swollen and burst at the nonpermissive temperature, indicating that their cell wall integrity was defective. Further analysis showed that the mutant cells were deficient in the activity of
cis
-prenyltransferase, namely, undecaprenyl diphosphate synthase, a key enzyme of the carrier lipid formation of peptidoglycan synthesis. The cellular level of undecaprenyl phosphate was in fact markedly decreased in the mutants. These results are consistent with the fact that the Rer2 homologue of
Micrococcus luteus
shows undecaprenyl diphosphate synthase activity (N. Shimizu, T. Koyama, and K. Ogura, J. Biol. Chem. 273:19476–19481, 1998) and demonstrate that
E. coli
Rth is indeed responsible for the maintenance of cell wall rigidity. Our work on the yeast
rer2
mutants shows that they are defective in the activity of
cis
-prenyltransferase, namely, dehydrodolichyl diphosphate synthase, a key enzyme of dolichol synthesis. Taking these data together, we conclude that the
RER2
gene family encodes
cis
-prenyltransferase, which plays an essential role in cell wall biosynthesis in bacteria and in dolichol synthesis in eukaryotic cells and has been well conserved during evolution.
Role of
CIS
in Replication of an IncB Plasmid
Authors: Praszkier J. , Pittard A. J. 
Published: December 2019

ABSTRACT

Replication of the IncB plasmid pMU720 requires the synthesis of the
cis
-acting RepA protein and the presence of two DNA elements,
ori
and
CIS. CIS
is the 166-bp sequence separating the RepA coding sequence from
ori
. To investigate how this organization of the pMU720 replicon contributes to the mechanism of initiation of replication, mutations in the sequence and/or the length of
CIS
were introduced into the
CIS
region and their effects on the efficiency of replication of the pMU720 replicon in vivo was determined. The
CIS
region was found to be composed of two domains. The
repA
-proximal domain, which showed strong transcription termination activity, could be replaced by equivalent sequences from I-complex and IncL/M plasmids, whose replicons are organized in the same fashion as pMU720. Replacement by a
trpA
transcription terminator afforded only partial replication activity. The
repA
-distal domain was shown to be a spacer whose role was to position sequence(s) within
ori
on the correct face of the DNA helix vis-à-vis the
repA
-proximal portion of
CIS
. A model for the loading of RepA protein onto
ori
is discussed.
Adenosylcobalamin-Mediated Methyl Transfer by Toluate
cis
-Dihydrodiol Dehydrogenase of the TOL Plasmid pWW0
Authors: Lee Jang-Young , Park Hee-Sung , Kim Hak-Sung 
Published: December 2019

ABSTRACT

We identified and characterized a methyl transfer activity of the toluate
cis
-dihydrodiol (4-methyl-3,5-cyclohexadiene-
cis
-1,2-diol-1-carboxylic acid) dehydrogenase of the TOL plasmid pWW0 towards toluene
cis
-dihydrodiol (3-methyl-4,5-cyclohexadiene-
cis
-1,2-diol). When the purified enzyme from the recombinant
Escherichia coli
containing the
xylL
gene was incubated with toluene
cis
-dihydrodiol in the presence of NAD
+
, the end products differed depending on the presence of adenosylcobalamin (coenzyme B
12
). The enzyme yielded catechol in the presence of adenosylcobalamin, while it gave 3-methylcatechol in the absence of the cofactor. Adenosylcobalamin was transformed to methylcobalamin as a result of the enzyme reaction, which indicates that the methyl group of the substrate was transferred to adenosylcobalamin. Other derivatives of the cobalamin such as aquo (hydroxy)- and cyanocobalamin did not mediate the methyl transfer reaction. The dehydrogenation and methyl transfer reactions were assumed to occur concomitantly, and the methyl transfer reaction seemed to depend on the dehydrogenation. To our knowledge, the enzyme is the first dehydrogenase that shows a methyl transfer activity as well.
Disulfide Bridges Are Not Involved in Penicillin-Binding Protein 1b Dimerization in
Escherichia coli
Authors: Chalut Christian , Remy Marie-Hélène , Masson Jean-Michel 
Published: December 2019

ABSTRACT

PBP1b can be found as a dimer in
Escherichia coli
. Previous results suggested that dimerization involved the cysteine(s) in an intermolecular disulfide bond. We show that either deletion mutants or a mutant without cysteines is fully active and still binds penicillin and that the latter can also form dimers.
Earlier Positivity of Central-Venous- versus Peripheral-Blood Cultures Is Highly Predictive of Catheter-Related Sepsis
Authors: Blot François , Schmidt Eric , Nitenberg Gérard , Tancrède Cyrille , Leclercq Bernard , Laplanche Agnès , Andremont Antoine 
Published: December 2019

ABSTRACT

To diagnose catheter-related sepsis without removing the catheter, we evaluated the differential positivity times of cultures of blood drawn simultaneously from central venous catheter and peripheral sites. In a 450-bed cancer reference center, simultaneous central- and peripheral-blood cultures were prospectively performed for patients with suspicion of catheter-related sepsis over an 18-month period. Data for 64 patients for whom the same microorganisms were found when central- and peripheral-blood samples were cultured were retrospectively reviewed by two independent physicians blinded to the differential positivity time values in order to establish or refute the diagnosis of catheter-related sepsis. The diagnosis was established in 28 cases, refuted in 14, and indeterminate in the remaining 22. The differential positivity time was significantly greater for patients with catheter-related sepsis (
P
< 10
−4
). A cutoff limit of +120 min had 100% specificity and 96.4% sensitivity for the diagnosis of catheter-related sepsis. These results strongly suggest that measurement of the differential positivity time might be a reliable tool facilitating the diagnosis of catheter-related sepsis in patients with an indwelling catheter.
Evaluation of a Rapid Immunochromatographic Test for Diagnosis of Dengue Virus Infection
Authors: Vaughn David W. , Nisalak Ananda , Kalayanarooj Siripen , Solomon Tom , Dung Nguyen Minh , Cuzzubbo Andrea , Devine Peter L. 
Published: December 2019

ABSTRACT

A rapid (<7-min) immunochromatographic test for immunoglobulin M (IgM) and IgG antibodies to dengue viruses was evaluated by using hospital admission and discharge sera from 124 patients. The reference laboratory diagnosis was based on the results of virus isolation, hemagglutination-inhibition assay (HAI), and enzyme immunoassay (EIA). By the standard assays, patients experienced primary dengue virus infection (
n
= 30), secondary dengue virus infection (
n
= 48), Japanese encephalitis (JE) virus infection (
n
= 20), or no flavivirus infection (
n
= 26). The rapid test demonstrated 100% sensitivity in the diagnosis of dengue virus infection and was able to distinguish between primary and secondary dengue virus infections through the separate determinations of IgM and IgG. For all patients with primary dengue virus infection a positive test for IgM to dengue virus and a negative test for IgG to dengue virus were obtained, whereas for 46 of 48 patients (96%) with secondary dengue virus infection, a positive test for IgG to dengue virus with or without a positive test for IgM to dengue virus was obtained. The remaining two patients with secondary dengue virus infection had positive IgM test results and negative IgG test results. Furthermore, the rapid test was positive for patients confirmed to be infected with different dengue virus serotypes (12 infected with dengue virus serotype 1, 4 infected with dengue virus serotype 2, 3 infected with dengue virus serotype 3, and 2 infected with dengue virus serotype 4). The specificity of the test for nonflavivirus infections was 88% (3 of 26 positive), while for JE virus infections the specificity of the test was only 50% (10 of 20). However, most patients with secondary dengue virus infection were positive for both IgM and IgG antibodies to dengue virus, while no patients with JE virus infection had this profile, so cross-reactivity was only a concern for a small proportion of patients with secondary dengue infections. The rapid test demonstrated a good correlation with the reference EIA and HAI and should be useful for the rapid diagnosis of dengue virus infections.
Quantitation of Human Immunodeficiency Virus Type 1 RNA in Plasma by Using Blood Dried on Filter Paper
Authors: Fiscus Susan A. , Brambilla Donald , Grosso Lynell , Schock Jody , Cronin Michael 
Published: December 2019

ABSTRACT
We compared whole blood dried on filter paper to the standard assay with frozen cell-free plasma for use in the quantitation of the human immunodeficiency virus RNA load in blood. RNA values from filter paper, corrected for the hematocrit, gave results comparable to those of the standard assay in terms of sensitivity and reproducibility.
Inhibition of PCR by Agar from Bacteriological Transport Media
Authors: Gibb Alan Patrick , Wong Sallene 
Published: December 2019

ABSTRACT
We observed inhibition of PCR in throat swabs submitted in routine bacteriological transport media. Experimental studies showed that agar, which was extracted with DNA by DNAzol (Gibco BRL, Gaithersburg, Md.), was the inhibitory agent. No inhibitory effect was observed with a transport medium which did not contain agar.
Pulsed-Field Gel Electrophoresis of Genomic Digests Demonstrates Linkages among Food, Food Handlers, and Patrons in a Food-Borne
Salmonella javiana
Outbreak in Massachusetts
Authors: Lee Roger , Peppe Joseph , George Harvey 
Published: December 2019

ABSTRACT

A total of 66 isolates of
Salmonella javiana
isolated from food, food handlers, and patrons that were epidemiologically linked to an outbreak of gastroenteritis were analyzed by pulsed-field gel electrophoresis. Analysis with restriction endonucleases
Xba
I and
Spe
I supported the epidemiologic association and suggested a pathway of transmission among food, food handlers, and patrons.
Use of a Gastric Juice-Based PCR Assay To Detect
Helicobacter pylori
Infection in Culture-Negative Patients
Authors: Yoshida Haruhiko , Hirota Katsutaro , Shiratori Yasushi , Nihei Takeshi , Amano Shin , Yoshida Akira , Kawamata Osamu , Omata Masao 
Published: December 2019

ABSTRACT

A gastric juice-based PCR assay was compared with culture, microscopy, and a rapid urease test with specimens from 114 subjects. The PCR and conventional tests were positive for 76 and 62% of the subjects, respectively. The prevalence of gastroduodenal disease and seropositivity for anti-
Helicobacter pylori
immunoglobulin G were similarly high among conventional-test-positive and PCR-only-positive subjects compared to all-negative ones. The PCR assay is recommended to confirm the
H. pylori
status of culture-negative peptic-ulcer patients.
Detection of
Helicobacter pylori
in Fecal Samples of Gnotobiotic Mice Infected with
H. pylori
by an Immunomagnetic-Bead Separation Technique
Authors: Osaki Takako , Taguchi Haruhiko , Yamaguchi Hiroyuki , Kamiya Shigeru 
Published: December 2019

ABSTRACT

By an immunomagnetic-bead (IMB) separation technique, isolation of
Helicobacter pylori
from gastrointestinal and fecal samples of gnotobiotic mice infected with the microorganism was tried. The isolation rate of
H. pylori
from stomach samples after IMB separation was not higher than that of direct culture of the samples. After IMB separation of feces,
H. pylori
was detectable by PCR, although
H. pylori
was not culturable.
Amplification of Ehrlichial DNA from Dogs 34 Months after Infection withEhrlichia canis
Authors: Harrus Shimon , Waner Trevor , Aizenberg Itzhak , Foley Janet E. , Poland Amy M. , Bark Hylton 
Published: December 2019

ABSTRACTIn order to determine whether dogs in the subclinical phase of canine monocytic ehrlichiosis (CME) are carriers ofEhrlichia canisand to determine the significance of persistent indirect immunofluorescent anti-E. canisantibody titers during this phase, PCR was performed with blood, bone marrow, and splenic aspirates collected 34 months postinoculation from six clinically healthy beagle dogs experimentally infected withE. canis. At least one of the three samples (spleen, bone marrow, and blood) from four of the six dogs was PCR positive. The spleens of all four of these dogs were PCR positive, and the bone marrow and blood of two of the four dogs were PCR positive. Indirect immunofluorescent-antibody titers increased progressively during the first 5 months postinfection, remained high for an additional period of more than 11 months, and declined thereafter, suggesting that the dogs were recovering from the disease. Five of the dogs remained seropositive 34 months postinfection. The data obtained in this study demonstrate for the first time that clinically healthy dogs in the subclinical phase of CME are carriers of the rickettsia. It was shown that dogs can harborE. canisfor years without developing the chronic clinical disease and that dogs can eliminate the parasite and recover from CME without medical treatment. Our findings suggest that the spleen is the organ most likely to harborE. canisparasites during the subclinical phase and the last organ to accommodate the parasite before elimination. It was concluded that PCR of DNA extracted from splenic aspirates is a reliable method for determining the carrier state of CME.
Development of Specific Immunoglobulins G, M, and A Following Primary
Toxoplasma gondii
Infection in Pregnant Women
Authors: Jenum Pål A. , Stray-Pedersen Babill 
Published: December 2019

ABSTRACT

The development of specific antibodies following primary
Toxoplasma gondii
infection during pregnancy was assessed by six different antibody assays: dye test, Platelia Toxo-IgG, Toxo-Screen DA IgG, Platelia Toxo-IgM, Toxo-ISAGA IgM, and Platelia Toxo-IgA. A total of 126 sera from 27 pregnant women, for whom the time of acquisition of infection could be estimated fairly accurately, were included. All tests showed great individual variation in the peak amounts of antibodies detected. The times elapsed after infection until the peak was reached also varied greatly from individual to individual: the ranges were 2 to 21 weeks for the dye test, 4 to 36 weeks for Platelia Toxo-IgG, 4 to 30 weeks for Toxo-Screen DA IgG, 2 to 18 weeks for Platelia Toxo-IgM, 1 to 6 weeks for Toxo-ISAGA IgM, and 2 to 21 weeks for Platelia Toxo-IgA. In the early phase of the infection the dye test and the specific-IgM tests were the most sensitive. Toxo-Screen DA IgG was more sensitive than Platelia Toxo-IgG in the acute phase, while Platelia Toxo-IgA was clearly the least sensitive assay. Of the sera collected 21 to 52 weeks after infection, all were positive by the dye test, all except one (which was negative by Platelia Toxo-IgG) were positive by the specific-IgG tests, approximately 80% were positive by the IgM tests, and 45% were positive by the IgA test. Due to the great individual variation it seems impossible to estimate when the infection occurred based on results obtained from a single serum, and it may even be difficult to assess when a titer increase in paired sera is detectable unless the first sample is only marginally positive. As a diagnostic criterion a dye test titer of ≥300 IU/ml has a low sensitivity for recent primary infection.
Molecular Epidemiology of Penicillin-Resistant
Streptococcus pneumoniae
Isolates Recovered in Italy from 1993 to 1996
Authors: Marchese Anna , Ramirez Mario , Schito Gian Carlo , Tomasz Alexander 
Published: December 2019

ABSTRACT

Thirty-nine penicillin-resistant
Streptococcus pneumoniae
isolates recovered among the approximately 700 pneumococcal strains collected from 1993 to 1996 in central and northern Italy were analyzed for several characteristics, including serotype, antibiotic susceptibility profile, chromosomal relatedness (by using pulsed-field gel electrophoresis [PFGE]), restriction fragment length polymorphism (RFLP) of the penicillin-binding protein (PBP) genes 1A, 2X, and 2B, and the presence of a variety of antibiotic resistance genes (determined by hybridization with appropriate DNA probes). The MICs of penicillin for most of the isolates (30 of 39) were high, in the range of 1 μg/ml or higher, and these 30 isolates carried additional resistance traits to two or more drugs (erythromycin, chloramphenicol, co-trimoxazole, and tetracycline) and expressed serotypes 9, 19, and 23 and three distinct PFGE patterns. More than half (22 of 30) of the isolates for which MICs were high were identified as representatives of two widespread international epidemic clones of
S. pneumoniae
. The first one of these clones (seven isolates) expressed serotype 23F and possessed all properties characteristic of the widespread Spanish/USA international clone. Seven additional strains with serotype 19 also had the same PFGE pattern, PBP gene, and RFLP polymorphisms, and other properties typical of the serotype 23 Spanish/USA clone, suggesting that these strains were the products of a capsular transformation event (from serotype 23F to serotype 19) in which the Spanish/USA clone was the recipient. The second international clone was represented by eight serotype 9 isolates which were resistant to penicillin and trimethoprim-sulfamethoxazole and had the molecular properties of the French/Spanish epidemic clone. The remaining eight isolates for which penicillin MICs were high appeared to represent a hitherto-undescribed “Italian” clone; they had a novel PFGE type, unique RFLPs for the PBP genes, and resistance to tetracycline, trimethoprim-sulfamethoxazole, and erythromycin, and the penicillin MICs for these isolates were 2 to 4 μg/ml.
Field Validation of Laboratory Tests for Clinical Diagnosis of Sheep-Associated Malignant Catarrhal Fever
Authors: Müller-Doblies U. U. , Li H. , Hauser B. , Adler H. , Ackermann M. 
Published: December 2019

ABSTRACT

Until recently, sheep-associated malignant catarrhal fever (SA-MCF) was diagnosed mainly on the basis of clinical presentation and histopathological changes. Using clinically diagnosed field cases, we have evaluated a seminested PCR and a competitive inhibition enzyme-linked immunosorbent assay (CI-ELISA) and compared these assays in the diagnosis of SA-MCF in cattle with histopathology as a provisional “gold standard.” Samples from 44 cattle with clinical signs suggestive of SA-MCF were examined by histopathology, PCR, and CI-ELISA. In addition, samples from healthy cattle were evaluated by PCR (
n
= 96) and CI-ELISA (
n
= 75). Based on histopathology, 38 of the 44 clinical cases were classified as SA-MCF positive, 3 were classified as inconclusive, and 3 were classified as SA-MCF negative. The sensitivity of PCR was 95 to 97%, whereas the specificity ranged between 94 and 100%. The CI-ELISA showed a sensitivity of 56 to 87% and a specificity between 91 and 100%. In the field, there is good correlation between the diagnoses of SA-MCF by histopathology, PCR, and CI-ELISA. These data also confirm the close association of ovine herpesvirus 2 with SA-MCF in Switzerland.