bluuebunny/crossref_metadata_2025_split huggingface dataset:
Sulfate-Reducing Bacteria and Their Activities in Cyanobacterial Mats of Solar Lake (Sinai, Egypt)
Authors: Teske Andreas , Ramsing Niels B. , Habicht Kirsten , Fukui Manabu , Küver Jan , Jørgensen Bo Barker , Cohen Yehuda 
Published: December 2019

ABSTRACT

The sulfate-reducing bacteria within the surface layer of the hypersaline cyanobacterial mat of Solar Lake (Sinai, Egypt) were investigated with combined microbiological, molecular, and biogeochemical approaches. The diurnally oxic surface layer contained between 10
6
and 10
7
cultivable sulfate-reducing bacteria ml
−1
and showed sulfate reduction rates between 1,000 and 2,200 nmol ml
−1
day
−1
, both in the same range as and sometimes higher than those in anaerobic deeper mat layers. In the oxic surface layer and in the mat layers below, filamentous sulfate-reducing
Desulfonema
bacteria were found in variable densities of 10
4
to 10
6
cells ml
−1
. A
Desulfonema
-related, diurnally migrating bacterium was detected with PCR and denaturing gradient gel electrophoresis within and below the oxic surface layer. Facultative aerobic respiration, filamentous morphology, motility, diurnal migration, and aggregate formation were the most conspicuous adaptations of Solar Lake sulfate-reducing bacteria to the mat matrix and to diurnal oxygen stress. A comparison of sulfate reduction rates within the mat and previously published photosynthesis rates showed that CO
2
from sulfate reduction in the upper 5 mm accounted for 7 to 8% of the total photosynthetic CO
2
demand of the mat.
Microbial Dechlorination of 2,3,5,6-Tetrachlorobiphenyl under Anaerobic Conditions in the Absence of Soil or Sediment
Authors: Cutter Leah , Sowers Kevin R. , May Harold D. 
Published: December 2019

ABSTRACT

Bacterial enrichment cultures developed with Baltimore Harbor (BH) sediments were found to reductively dechlorinate 2,3,5,6-tetrachlorobiphenyl (2,3,5,6-CB) when incubated in a minimal estuarine medium containing short-chain fatty acids under anaerobic conditions with and without the addition of sediment. Primary enrichment cultures formed both
meta
and
ortho
dechlorination products from 2,3,5,6-CB. The lag time preceding dechlorination decreased from 30 to less than 20 days as the cultures were sequentially transferred into estuarine medium containing dried, sterile BH sediment. In addition, only
ortho
dechlorination was observed following transfer of the cultures. Sequential transfer into medium without added sediment also resulted in the development of a strict
ortho
-dechlorinating culture following a lag of more than 100 days. Upon further transfer into the minimal medium without sediment, the lag time decreased to less than 50 days. At this stage all cultures, regardless of the presence of sediment, would produce 2,3,5-CB and 3,5-CB from 2,3,5,6-CB. The strict
ortho
-dechlorinating activity in the sediment-free cultures has remained stable for more than 1 year through several transfers. These results reveal that the classical microbial enrichment technique using a minimal medium with a single polychlorinated biphenyl (PCB) congener selected for
ortho
dechlorination of 2,3,5,6-CB. Furthermore, this is the first report of sustained anaerobic PCB dechlorination in the complete absence of soil or sediment.
Phage Display of a Biologically Active
Bacillus thuringiensis
Toxin
Authors: Kasman Laura M. , Lukowiak Andrew A. , Garczynski Stephen F. , McNall Rebecca J. , Youngman Phil , Adang Michael J. 
Published: December 2019

ABSTRACT

Activated forms of
Bacillus thuringiensis
insecticidal toxins have consistently been found to form insoluble and inactive precipitates when they are expressed in
Escherichia coli
. Genetic engineering of these proteins to improve their effectiveness as biological pesticides would be greatly facilitated by the ability to express them in
E. coli
, since the molecular biology tools available for
Bacillus
are limited. To this end, we show that activated
B. thuringiensis
toxin (Cry1Ac) can be expressed in
E. coli
as a translational fusion with the minor phage coat protein of filamentous phage. Phage particles displaying this fusion protein were viable, infectious, and as lethal as pure toxin on a molar basis when the phage particles were fed to insects susceptible to native Cry1Ac. Enzyme-linked immunosorbent assay and Western blot analysis showed the fusion protein to be antigenically equivalent to native toxin, and micropanning with anti-Cry1Ac antibody was positive for the toxin-expressing phage. Phage display of
B. thuringiensis
toxins has many advantages over previous expression systems for these proteins and should make it possible to construct large libraries of toxin variants for screening or biopanning.
Whole-Cell Immunolocalization of Nitrogenase in Marine Diazotrophic Cyanobacteria,
Trichodesmium
spp
Authors: Lin Senjie , Henze Sheri , Lundgren Pernilla , Bergman Birgitta , Carpenter Edward J. 
Published: December 2019

ABSTRACT

The mechanism by which planktonic marine cyanobacteria of the genus
Trichodesmium
fix N
2
aerobically during photosynthesis without heterocysts is unknown. As an aid in understanding how these species protect nitrogenase, we have developed an immunofluorescence technique coupled to light microscopy (IF-LM) with which intact cyanobacteria can be immunolabeled and the distribution patterns of nitrogenase and other proteins can be described and semiquantified. Chilled ethanol was used to fix the cells, which were subsequently made permeable to antibodies by using dimethyl sulfoxide. Use of this technique demonstrated that about 3 to 20 cells (mean ± standard deviation, 9 ± 4) consecutively arranged in a
Trichodesmium
trichome were labeled with the nitrogenase antibody. The nitrogenase-containing cells were distributed more frequently around the center of the trichome and were rarely found at the ends. On average 15% of over 300 randomly encountered cells examined contained nitrogenase. The percentage of nitrogenase-containing cells (nitrogenase index [NI]) in an exponential culture was higher early in the light period than during the rest of the light-dark cycle, while that for a stationary culture was somewhat constant at a lower level throughout the light-dark cycle. The NI was not affected by treatment of the cultures with the photosynthetic inhibitor dichloro 1,3′-dimethyl urea or with low concentrations of ammonium (NH
4
Cl). However, incubation of cultures with 0.5 μM NH
4
Cl over 2 days reduced the NI. The IF technique combined with
14
C autoradiography showed that the CO
2
fixation rate was lower in nitrogenase-containing cells. The results of the present study suggest that (i) the IF-LM technique may be a useful tool for in situ protein localization in cyanobacteria, (ii) cell differentiation occurs in
Trichodesmium
and only a small fraction of cells in a colony have the potential to fix nitrogen, (iii) the photosynthetic activity (CO
2
uptake) is reduced if not absent in N
2
-fixing cells, and (iv) variation in the NI may be a modulator of nitrogen-fixing activity.
Minimally Invasive Detection of
Piscirickettsia salmonis
in Cultivated Salmonids via the PCR
Authors: Marshall Sergio , Heath Sekou , Henríquez Vitalia , Orrego Cristián 
Published: December 2019

ABSTRACT

The attributes of the PCR allowed implementation of an assay for specific detection of
Piscirickettsia salmonis
from a few microliters of fish serum. This opens the way to less invasive modes of sampling for this microbial pathogen in salmonids.
Failure of
Ixodes
Ticks To Inherit
Borrelia afzelii
Infection
Authors: Matuschka Franz-Rainer , Schinkel Thomas W. , Klug Birte , Spielman Andrew , Richter Dania 
Published: December 2019

ABSTRACT

To define conditions promoting inherited infection by Lyme disease spirochetes in
Ixodes
ticks, we variously infected ticks with
Borrelia afzelii
and examined their progenies by dark-field microscopy, immunofluorescence, PCR, and serial passage. No episode of inherited infection was evident, regardless of instar or gender infected or frequency of exposure. We suggest that these spirochetes rarely, if ever, are inherited by vector ticks.
Cloning and Expression of the
Listeria monocytogenes
Scott A
ptsH
and
ptsI
Genes, Coding for HPr and Enzyme I, Respectively, of the Phosphotransferase System
Authors: Christensen Douglas P. , Benson Andrew K. , Hutkins Robert W. 
Published: December 2019

ABSTRACT

The phosphoenolpyruvate (PEP)-dependent phosphotransferase system (PTS) utilizes high-energy phosphate present in PEP to drive the uptake of several different carbohydrates in bacteria. In order to examine the role of the PTS in the physiology of
Listeria monocytogenes
, we identified the
ptsH
and
ptsI
genes encoding the HPr and enzyme I proteins, respectively, of the PTS. Nucleotide sequence analysis indicated that the predicted proteins are nearly 70% similar to HPr and enzyme I proteins from other organisms. Purified
L. monocytogenes
HPr overexpressed in
Escherichia coli
was also capable of complementing an HPr defect in heterologous extracts of
Staphylococcus aureus ptsH
mutants. Additional studies of the transcriptional organization and control indicated that the
ptsH
and
ptsI
genes are organized into a transcription unit that is under the control of a consensus-like promoter and that expression of these genes is mediated by glucose availability and pH or by by-products of glucose metabolism.
Modeling of the Competitive Growth of
Listeria monocytogenes
and
Lactococcus lactis
in Vegetable Broth
Authors: Breidt Frederick , Fleming Henry P. 
Published: December 2019

ABSTRACT

Current mathematical models used by food microbiologists do not address the issue of competitive growth in mixed cultures of bacteria. We developed a mathematical model which consists of a system of nonlinear differential equations describing the growth of competing bacterial cell cultures. In this model, bacterial cell growth is limited by the accumulation of protonated lactic acid and decreasing pH. In our experimental system, pure and mixed cultures of
Lactococcus lactis
and
Listeria monocytogenes
were grown in a vegetable broth medium. Predictions of the model indicate that pH is the primary factor that limits the growth of
L. monocytogenes
in competition with a strain of
L. lactis
which does not produce the bacteriocin nisin. The model also predicts the values of parameters that affect the growth and death of the competing populations. Further development of this model will incorporate the effects of additional inhibitors, such as bacteriocins, and may aid in the selection of lactic acid bacterium cultures for use in competitive inhibition of pathogens in minimally processed foods.
Analysis ofEscherichia coliO157:H7 Survival in Ovine or Bovine Manure and Manure Slurry
Authors: Kudva Indira T. , Blanch Kathryn , Hovde Carolyn J. 
Published: December 2019

ABSTRACTFarm animal manure or manure slurry may disseminate, transmit, or propagateEscherichia coliO157:H7. In this study, the survival and growth ofE. coliO157:H7 in ovine or bovine feces under various experimental and environmental conditions were determined. A manure pile collected from experimentally inoculated sheep was incubated outside under fluctuating environmental conditions.E. coliO157:H7 survived in the manure for 21 months, and the concentrations of bacteria recovered ranged from <102to 106CFU/g at different times over the course of the experiment. The DNA fingerprints ofE. coliO157:H7 isolated at month 1 and month 12 were identical or very similar. A secondE. coliO157:H7-positive ovine manure pile, which was periodically aerated by mixing, remained culture positive for 4 months. AnE. coliO157:H7-positive bovine manure pile was culture positive for 47 days. In the laboratory,E. coliO157:H7 was inoculated into feces, untreated slurry, or treated slurry and incubated at −20, 4, 23, 37, 45, and 70°C.E. coliO157:H7 survived best in manure incubated without aeration at temperatures below 23°C, but it usually survived for shorter periods of time than it survived in manure held in the environment. The bacterium survived at least 100 days in bovine manure frozen at −20°C or in ovine manure incubated at 4 or 10°C for 100 days, but under all other conditions the length of time that it survived ranged from 24 h to 40 days. In addition, we found that the Shiga toxin type 1 and 2 genes inE. coliO157:H7 had little or no influence on bacterial survival in manure or manure slurry. The long-term survival ofE. coliO157:H7 in manure emphasizes the need for appropriate farm waste management to curtail environmental spread of this bacterium. This study also highlights the difficulties in extrapolating laboratory data to on-farm conditions.
Role of Endoproteolytic Dibasic Proprotein Processing in Maturation of Secretory Proteins in
Trichoderma reesei
Authors: Goller Sabine P. , Schoisswohl Doris , Baron Michel , Parriche Martine , Kubicek Christian P. 
Published: December 2019

ABSTRACT

Cell extracts of
Trichoderma reesei
exhibited dibasic endopeptidase activity toward the carboxylic side of KR, RR, and PR sequences. This activity was stimulated by the presence of Ca
2+
ions and localized in vesicles of low bouyant density; it therefore exhibited some similarity to yeast Kex2. Analytical chromatofocusing revealed a single peak of activity. The dibasic endopeptidase activity was strongly and irreversibly inhibited in vitro as well as in vivo by 1 mM
p
-amidinophenylmethylsulfonyl fluoride (pAPMSF) but not by PMSF at concentrations up to 5 mM. We therefore used pAPMSF to study the role of the dibasic endopeptidase in the secretion of protein by
T. reesei
. Secretion of xylanase I (proprotein processing sequence -R-R-↓-R-↓-A-) and xylanase II (-K-R-↓-Q-) was strongly inhibited by 1 mM pAPMSF, and a larger, unprocessed enzyme form was detected intracellularly under these conditions. Secretion of cellobiohydrolase II (CBH II; -E-R-↓-Q-) was only slightly inhibited by pAPMSF, and no accumulation of unprocessed precursors was detected. In contrast, secretion of CBH I (-R-A-↓-Q-) was stimulated by pAPMSF addition, and a simultaneous decrease in the concentration of intracellular CBH I was detected. Similar experiments were also carried out with a single heterologous protein, ShBLE, the phleomycin-binding protein from
Streptoalloteichus hindustanus
, fused to a series of model proprotein-processing sequences downstream of the expression signals of the
Aspergillus nidulans gpdA
promoter. Consistent with the results obtained with homologous proteins, pAPMSF inhibited the secretion of ShBLE with fusions containing dibasic (RK and KR) target sequences, but it even stimulated secretion in fusions to LR, NHA, and EHA target sequences. Addition of 5 mM PMSF, a nonspecific inhibitor of serine protease, nonspecifically inhibited the secretion of heterologous proteins from fusions bearing the NHA and LR targets. These data point to the existence of different endoproteolytic proprotein processing enzymes in
T. reesei
and demonstrate that dibasic processing is obligatory for the secretion of the proproteins containing this target.
Competitive Dominance among Strains of Luminous Bacteria Provides an Unusual Form of Evidence for Parallel Evolution in Sepiolid Squid-Vibrio Symbioses
Authors: Nishiguchi Michele K. , Ruby Edward G. , McFall-Ngai Margaret J. 
Published: December 2019

ABSTRACT
One of the principal assumptions in symbiosis research is that associated partners have evolved in parallel. We report here experimental evidence for parallel speciation patterns among several partners of the sepiolid squid-luminous bacterial symbioses. Molecular phylogenies for 14 species of host squids were derived from sequences of both the nuclear internal transcribed spacer region and the mitochondrial cytochrome oxidase subunit I; the glyceraldehyde phosphate dehydrogenase locus was sequenced for phylogenetic determinations of 7 strains of bacterial symbionts. Comparisons of trees constructed for each of the three loci revealed a parallel phylogeny between the sepiolids and their respective symbionts. Because both the squids and their bacterial partners can be easily cultured independently in the laboratory, we were able to couple these phylogenetic analyses with experiments to examine the ability of the different symbiont strains to compete with each other during the colonization of one of the host species. Our results not only indicate a pronounced dominance of native symbiont strains over nonnative strains, but also reveal a hierarchy of symbiont competency that reflects the phylogenetic relationships of the partners. For the first time, molecular systematics has been coupled with experimental colonization assays to provide evidence for the existence of parallel speciation among a set of animal-bacterial associations.
Comparative Studies of Class IIa Bacteriocins of Lactic Acid Bacteria
Authors: Eijsink Vincent G. H. , Skeie Marianne , Middelhoven P. Hans , Brurberg May Bente , Nes Ingolf F. 
Published: December 2019

ABSTRACT

Four class IIa bacteriocins (pediocin PA-1, enterocin A, sakacin P, and curvacin A) were purified to homogeneity and tested for activity toward a variety of indicator strains. Pediocin PA-1 and enterocin A inhibited more strains and had generally lower MICs than sakacin P and curvacin A. The antagonistic activity of pediocin-PA1 and enterocin A was much more sensitive to reduction of disulfide bonds than the antagonistic activity of sakacin P and curvacin A, suggesting that an extra disulfide bond that is present in the former two may contribute to their high levels of activity. The food pathogen
Listeria monocytogenes
was among the most sensitive indicator strains for all four bacteriocins. Enterocin A was most effective in inhibiting
Listeria
, having MICs in the range of 0.1 to 1 ng/ml. Sakacin P had the interesting property of being very active toward
Listeria
but not having concomitant high levels of activity toward lactic acid bacteria. Strains producing class IIa bacteriocins displayed various degrees of resistance toward noncognate class IIa bacteriocins; for the sakacin P producer, it was shown that this resistance is correlated with the expression of immunity genes. It is hypothesized that variation in the presence and/or expression of such immunity genes accounts in part for the remarkably large variation in bacteriocin sensitivity displayed by lactic acid bacteria.
Enzymatic Properties of a Novel Liquefying α-Amylase from an Alkaliphilic
Bacillus
Isolate and Entire Nucleotide and Amino Acid Sequences
Authors: Igarashi Kazuaki , Hatada Yuji , Hagihara Hiroshi , Saeki Katsuhisa , Takaiwa Mikio , Uemura Takaaki , Ara Katsutoshi , Ozaki Katsuya , Kawai Shuji , Kobayashi Tohru , Ito Susumu 
Published: December 2019

ABSTRACT

A novel liquefying α-amylase (LAMY) was found in cultures of an alkaliphilic
Bacillus
isolate, KSM-1378. The specific activity of purified LAMY was approximately 5,000 U mg of protein
−1
, a value two- to fivefold greater between pH 5 and 10 than that of an industrial, thermostable
Bacillus licheniformis
enzyme. The enzyme had a pH optimum of 8.0 to 8.5 and displayed maximum activity at 55°C. The molecular mass deduced from sodium dodecyl sulfate-polyacrylamide gel electrophoresis was approximately 53 kDa, and the apparent isoelectric point was around pH 9. This enzyme efficiently hydrolyzed various carbohydrates to yield maltotriose, maltopentaose, maltohexaose, and maltose as major end products after completion of the reaction. Maltooligosaccharides in the maltose-to-maltopentaose range were unhydrolyzable by the enzyme. The structural gene for LAMY contained a single open reading frame 1,548 bp in length, corresponding to 516 amino acids that included a signal peptide of 31 amino acids. The calculated molecular mass of the extracellular mature enzyme was 55,391 Da. LAMY exhibited relatively low amino acid identity to other liquefying amylases, such as the enzymes from
B. licheniformis
(68.9%),
Bacillus amyloliquefaciens
(66.7%), and
Bacillus stearothermophilus
(68.6%). The four conserved regions, designated I, II, III, and IV, and the putative catalytic triad were found in the deduced amino acid sequence of LAMY. Essentially, the sequence of LAMY was consistent with the tertiary structures of reported amylolytic enzymes, which are composed of domains A, B, and C and which include the well-known (α/β)
8
barrel motif in domain A.
Characterization of a Defined 2,3,5,6-Tetrachlorobiphenyl-
ortho
-Dechlorinating Microbial Community by Comparative Sequence Analysis of Genes Coding for 16S rRNA
Authors: Pulliam Holoman Tracey R. , Elberson Margaret A. , Cutter Leah A. , May Harold D. , Sowers Kevin R. 
Published: December 2019

ABSTRACT

Defined microbial communities were developed by combining selective enrichment with molecular monitoring of total community genes coding for 16S rRNAs (16S rDNAs) to identify potential polychlorinated biphenyl (PCB)-dechlorinating anaerobes that
ortho
dechlorinate 2,3,5,6-tetrachlorobiphenyl. In enrichment cultures that contained a defined estuarine medium, three fatty acids, and sterile sediment, a
Clostridium
sp. was predominant in the absence of added PCB, but undescribed species in the δ subgroup of the class
Proteobacteria
, the low-G+C gram-positive subgroup, the
Thermotogales
subgroup, and a single species with sequence similarity to the deeply branching species
Dehalococcoides ethenogenes
were more predominant during active dechlorination of the PCB. Species with high sequence similarities to
Methanomicrobiales
and
Methanosarcinales
archaeal subgroups were predominant in both dechlorinating and nondechlorinating enrichment cultures. Deletion of sediment from PCB-dechlorinating enrichment cultures reduced the rate of dechlorination and the diversity of the community. Substitution of sodium acetate for the mixture of three fatty acids increased the rate of dechlorination, further reduced the community diversity, and caused a shift in the predominant species that included restriction fragment length polymorphism patterns not previously detected. Although PCB-dechlorinating cultures were methanogenic, inhibition of methanogenesis and elimination of the archaeal community by addition of bromoethanesulfonic acid only slightly inhibited dechlorination, indicating that the archaea were not required for
ortho
dechlorination of the congener. Deletion of
Clostridium
spp. from the community profile by addition of vancomycin only slightly reduced dechlorination. However, addition of sodium molybdate, an inhibitor of sulfate reduction, inhibited dechlorination and deleted selected species from the community profiles of the class
Bacteria
. With the exception of one 16S rDNA sequence that had the highest sequence similarity to the obligate perchloroethylene-dechlorinating
Dehalococcoides
, the 16S rDNA sequences associated with PCB
ortho
dechlorination had high sequence similarities to the δ, low-G+C gram-positive, and
Thermotogales
subgroups, which all include sulfur-, sulfate-, and/or iron(III)-respiring bacterial species.