bluuebunny/crossref_metadata_2025 huggingface dataset:
Evaluation of
Cryptosporidium parvum
Genotyping Techniques
Authors: Sulaiman Irshad M. , Xiao Lihua , Lal Altaf A.
Published: December 2019

We evaluated the specificity and sensitivity of 11 previously described species differentiation and genotyping PCR protocols for detection of
Cryptosporidium
parasites. Genomic DNA from three species of
Cryptosporidium
parasites (genotype 1 and genotype 2 of
C. parvum
,
C. muris
, and
C. serpentis
), two
Eimeria
species (
E. neischulzi
and
E. papillata
), and
Giardia duodenalis
were used to evaluate the specificity of primers. Furthermore, the sensitivity of the genotyping primers was tested by using genomic DNA isolated from known numbers of oocysts obtained from a genotype 2
C. parvum
isolate. PCR amplification was repeated at least three times with all of the primer pairs. Of the 11 protocols studied, 10 amplified
C. parvum
genotypes 1 and 2, and the expected fragment sizes were obtained. Our results indicate that two species-differentiating protocols are not
Cryptosporidium
specific, as the primers used in these protocols also amplified the DNA of
Eimeria
species. The sensitivity studies revealed that two nested PCR-restriction fragment length polymorphism (RFLP) protocols based on the small-subunit rRNA and dihydrofolate reductase genes are more sensitive than single-round PCR or PCR-RFLP protocols.
Distribution of Bifidobacterial Species in Human Intestinal Microflora Examined with 16S rRNA-Gene-Targeted Species-Specific Primers
Authors: Matsuki Takahiro , Watanabe Koichi , Tanaka Ryuichiro , Fukuda Masafumi , Oyaizu Hiroshi
Published: December 2019

In order to clarify the distribution of bifidobacterial species in the human intestinal tract, a 16S rRNA-gene-targeted species-specific PCR technique was developed and used with DNAs extracted from fecal samples obtained from 48 healthy adults and 27 breast-fed infants. To cover all of the bifidobacterial species that have been isolated from and identified in the human intestinal tract, species-specific primers for
Bifidobacterium longum
,
B. infantis
,
B. dentium
, and
B. gallicum
were developed and used with primers for
B. adolescentis
,
B. angulatum
,
B. bifidum
,
B. breve
, and the
B. catenulatum
group (
B. catenulatum
and
B. pseudocatenulatum
) that were developed in a previous study (T. Matsuki, K. Watanabe, R. Tanaka, and H. Oyaizu, FEMS Microbiol. Lett. 167:113–121, 1998). The specificity of the nine primers was confirmed by PCR, and the species-specific PCR method was found to be a useful means for identifying
Bifidobacterium
strains isolated from human feces. The results of an examination of bifidobacterial species distribution showed that the
B. catenulatum
group was the most commonly found taxon (detected in 44 of 48 samples [92%]), followed by
B. longum
and
B. adolescentis
, in the adult intestinal bifidobacterial flora and that
B. breve
,
B. infantis
, and
B. longum
were frequently found in the intestinal tracts of infants. The present study demonstrated that qualitative detection of the bifidobacterial species present in human feces can be accomplished rapidly and accurately.
Utilization of Trihalogenated Propanes by
Agrobacterium radiobacter
AD1 through Heterologous Expression of the Haloalkane Dehalogenase from
Rhodococcus
sp. Strain m15-3
Authors: Bosma Tjibbe , Kruizinga Edwin , de Bruin Erik J. , Poelarends Gerrit J. , Janssen Dick B.
Published: December 2019

Trihalogenated propanes are toxic and recalcitrant organic compounds. Attempts to obtain pure bacterial cultures able to use these compounds as sole carbon and energy sources were unsuccessful. Both the haloalkane dehalogenase from
Xanthobacter autotrophicus
GJ10 (DhlA) and that from
Rhodococcus
sp. strain m15-3 (DhaA) were found to dehalogenate trihalopropanes to 2,3-dihalogenated propanols, but the kinetic properties of the latter enzyme are much better. Broad-host-range dehalogenase expression plasmids, based on RSF1010 derivatives, were constructed with the haloalkane dehalogenase from
Rhodococcus
sp. strain m15-3 under the control of the heterologous promoters

P
lac

,

P
dhlA

, and

P
trc

. The resulting plasmids yielded functional expression in several gram-negative bacteria. A catabolic pathway for trihalopropanes was designed by introducing these broad-host-range dehalogenase expression plasmids into
Agrobacterium radiobacter
AD1, which has the ability to utilize dihalogenated propanols for growth. The recombinant strain AD1(pTB3), expressing the haloalkane dehalogenase gene under the control of the
dhlA
promoter, was able to utilize both 1,2,3-tribromopropane and 1,2-dibromo-3-chloropropane as sole carbon sources. Moreover, increased expression of the haloalkane dehalogenase resulted in elevated resistance to trihalopropanes.
Effect of Organic Complex Compounds on
Bacillus thermoamylovorans
Growth and Glucose Fermentation
Authors: Combet-Blanc Y. , Dieng M. C. , Kergoat P. Y.
Published: December 2019

The effect of the concentration of a mixture (1/1 [wt/wt]) of yeast extract and bioTrypcase (YE+bT) on the growth and physiology of a new species,
Bacillus thermoamylovorans
, a moderately thermophilic, non-spore-forming, lactic acid-producing bacterium isolated from palm wine, was studied. At an initial glucose concentration of 100 mM,
B. thermoamylovorans
growth was limited when the concentration of YE+bT was lower than 5.0 g liter
−1
; under these conditions, cellular yield reached a maximum value of 0.4 g of cells per g of YE+bT. Growth limitation due to deficiency in growth factors led to a significant shift in glucose metabolism towards lactate production. Lactate constituted 27.5 and 76% of the end products of glucose fermentation in media containing YE+bT at 20.0 and 1.0 g liter
−1
, respectively. This result markedly differed from published data for lactic bacteria, which indicated that fermentative metabolism remained homolactic regardless of the concentration of YE. Our results showed that the ratio between cellular synthesis and energy production increased with the concentration of YE+bT in the culture medium. They indicate that the industrial production of lactic acid through glucose fermentation by
B. thermoamylovorans
can be optimized by using a medium where glucose is present in excess and the organic additives are limiting.
Helicobacter
Species Colonizing Pig Stomach: Molecular Characterization and Determination of Prevalence
Authors: Cantet Franck , Magras Catherine , Marais Armelle , Federighi Michel , Mégraud Francis
Published: December 2019

The infection rate of 60 pigs (10 pigs from each of six farms) by
Helicobacter
species was studied by two techniques. Histological examination of the cardiac area of the stomach yielded a 58% positive result versus an 80% positive result by PCR with genus-specific primers. Eighty percent of the 16S rRNA gene was amplified, classified in four groups by PCR-restriction fragment length polymorphism, and sequenced. Isolates from all farms except one (farm C) were identified as
Helicobacter heilmannii
type 1, while those from farm C were identified as
H. heilmannii
type 2. Attempts to culture this organism in vitro failed.
Helicobacter pylori
was not found in these animals.
Viability and DNA Maintenance in Nonculturable Spiral
Campylobacter jejuni
Cells after Long-Term Exposure to Low Temperatures
Authors: Lázaro Beatriz , Cárcamo Jose , Audícana Ana , Perales Ildefonso , Fernández-Astorga Aurora
Published: December 2019

Survival of
Campylobacter jejuni
at 4 and 20°C was investigated by using cellular integrity, respiratory activity, two-dimensional (2D) protein profile, and intact DNA content as indicators of potential viability of nonculturable cells. Intact DNA content after 116 days, along with cellular integrity and respiring cells, was detected for up to 7 months at 4°C by pulsed-field gel electrophoresis. Most changes in 2D protein profiles involved up- or down-regulation.
Reduction of Selenite and Detoxification of Elemental Selenium by the Phototrophic Bacterium
Rhodospirillum rubrum
Authors: Kessi J. , Ramuz M. , Wehrli E. , Spycher M. , Bachofen R.
Published: December 2019

The effect of selenite on growth kinetics, the ability of cultures to reduce selenite, and the mechanism of detoxification of selenium were investigated by using
Rhodospirillum rubrum
. Anoxic photosynthetic cultures were able to completely reduce as much as 1.5 mM selenite, whereas in aerobic cultures a 0.5 mM selenite concentration was only reduced to about 0.375 mM. The presence of selenite in the culture medium strongly affected cell division. In the presence of a selenite concentration of 1.5 mM cultures reached final cell densities that were only about 15% of the control final cell density. The cell density remained nearly constant during the stationary phase for all of the selenite concentrations tested, showing that the cells were not severely damaged by the presence of selenite or elemental selenium. Particles containing elemental selenium were observed in the cytoplasm, which led to an increase in the buoyant density of the cells. Interestingly, the change in the buoyant density was reversed after selenite reduction was complete; the buoyant density of the cells returned to the buoyant density of the control cells. This demonstrated that
R. rubrum
expels elemental selenium across the plasma membrane and the cell wall. Accordingly, electron-dense particles were more numerous in the cells during the reduction phase than after the reduction phase.
Characterization of Cadmium Uptake in
Lactobacillus plantarum
and Isolation of Cadmium and Manganese Uptake Mutants
Authors: Hao Zhiqi , Reiske Heinz R. , Wilson David B.
Published: December 2019

Two different Cd
2+
uptake systems were identified in
Lactobacillus plantarum
. One is a high-affinity, high-velocity Mn
2+
uptake system which also takes up Cd
2+
and is induced by Mn
2+
starvation. The calculated

K
m

and
V
max
are 0.26 μM and 3.6 μmol g of dry cell
−1
min
−1
, respectively. Unlike Mn
2+
uptake, which is facilitated by citrate and related tricarboxylic acids, Cd
2+
uptake is weakly inhibited by citrate. Cd
2+
and Mn
2+
are competitive inhibitors of each other, and the affinity of the system for Cd
2+
is higher than that for Mn
2+
. The other Cd
2+
uptake system is expressed in Mn
2+
-sufficient cells, and no

K
m

can be calculated for it because uptake is nonsaturable. Mn
2+
does not compete for transport through this system, nor does any other tested cation, i.e., Zn
2+
, Cu
2+
, Co
2+
, Mg
2+
, Ca
2+
, Fe
2+
, or Ni
2+
. Both systems require energy, since uncouplers completely inhibit their activities. Two Mn
2+
-dependent
L. plantarum
mutants were isolated by chemical mutagenesis and ampicillin enrichment. They required more than 5,000 times as much Mn
2+
for growth as the parental strain. Mn
2+
starvation-induced Cd
2+
uptake in both mutants was less than 5% the wild-type rate. The low level of long-term Mn
2+
or Cd
2+
accumulation by the mutant strains also shows that the mutations eliminate the high-affinity Mn
2+
and Cd
2+
uptake system.
Prediction of Toxigenic Fungal Growth in Buildings by Using a Novel Modelling System
Authors: Rowan Neil J. , Johnstone Cameron M. , McLean R. Craig , Anderson John G. , Clarke Joe A.
Published: December 2019

There is growing concern about the adverse effects of fungal bioaerosols on the occupants of damp dwellings. Based on an extensive analysis of previously published data and on experiments carried out within this study, critical limits for the growth of the indoor fungi
Eurotium herbariorum
,
Aspergillus versicolor
, and
Stachybotrys chartarum
were mathematically described in terms of growth limit curves (isopleths) which define the minimum combination of temperature (T) and relative humidity (RH) at which growth will occur. Each growth limit curve was generated from a series of data points on a T-RH plot and mathematically fitted by using a third-order polynomial equation of the form RH =
a
3
T
3
+
a
2
T
2
+
a
1
T +
a
0
. This fungal growth prediction model was incorporated within the ESP-
r
(Environmental Systems Performance [
r
stands for “research”]) computer-based program for transient simulation of the energy and environmental performance of buildings. For any specified location, the ESP-
r
system is able to predict the time series evolution of local surface temperature and relative humidity, taking explicit account of constructional moisture flow, moisture generation sources, and air movement. This allows the predicted local conditions to be superimposed directly onto fungal growth curves. The concentration of plotted points relative to the curves allows an assessment of the risk of fungal growth. The system’s predictive capability was tested via laboratory experiments and by comparison with monitored data from a fungus-contaminated house.
Expression of the
ggpS
Gene, Involved in Osmolyte Synthesis in the Marine Cyanobacterium
Synechococcus
sp. Strain PCC 7002, Revealed Regulatory Differences between This Strain and the Freshwater Strain
Synechocystis
sp. Strain PCC 6803
Authors: Engelbrecht Friederike , Marin Kay , Hagemann Martin
Published: December 2019

Synthesis of the osmolyte glucosylglycerol (GG) in the marine cyanobacterium
Synechococcus
sp. strain PCC 7002 was characterized. The
ggpS
gene, which encodes the key enzyme (GG-phosphate synthase [GgpS]) in GG biosynthesis, was cloned by using PCR. A 2,030-bp DNA sequence which contained one open reading frame (ORF) was obtained. The protein deduced from this ORF exhibited 85% similarity to the GgpS of the freshwater cyanobacterium
Synechocystis
sp. strain PCC 6803. The function of the protein was confirmed by generating a
ggpS
null mutant, which was not able to synthesize GG and thus exhibited a salt-sensitive phenotype. Expression of the
ggpS
gene was analyzed in salt-shocked cells by performing Northern blot and immunoblot experiments. While almost no expression was detected in cells grown in low-salt medium, immediately after a salt shock the amounts of
ggpS
mRNA and GgpS protein increased up to 100-fold. The finding that salt-induced expression occurred was confirmed by measuring enzyme activities, which were negligible in control cells but clearly higher in salt-treated
Synechococcus
sp. cells. The salt-induced increase in GgpS activity could be inhibited by adding chloramphenicol, while in protein extracts of the freshwater cyanobacterium
Synechocystis
sp. strain PCC 6803 a constitutive, high level of enzyme activity that was not affected by chloramphenicol was found. A comparison of GG accumulation in the two cyanobacteria revealed that in the marine strain osmolyte synthesis seemed to be regulated mainly by transcriptional control, whereas in the freshwater strain control seemed to be predominantly posttranslational.
Phylogenetic Differentiation of Two Closely Related
Nitrosomonas
spp. That Inhabit Different Sediment Environments in an Oligotrophic Freshwater Lake
Authors: Whitby Corinne B. , Saunders Jon R. , Rodriguez Juana , Pickup Roger W. , McCarthy Alan
Published: December 2019

The population of ammonia-oxidizing bacteria in a temperate oligotrophic freshwater lake was analyzed by recovering 16S ribosomal DNA (rDNA) from lakewater and sediment samples taken throughout a seasonal cycle.
Nitrosospira
and
Nitrosomonas
16S rRNA genes were amplified in a nested PCR, and the identity of the products was confirmed by oligonucleotide hybridization.
Nitrosospira
DNA was readily identified in all samples, and nitrosomonad DNA of the
Nitrosomonas europaea-Nitrosomonas eutropha
lineage was also directly detected, but during the summer months only. Phylogenetic delineation with partial (345 bp) 16S rRNA gene sequences of clones obtained from sediments confirmed the fidelity of the amplified nitrosomonad DNA and identified two sequence clusters closely related to either
N. europaea
or
N. eutropha
that were equated with the littoral and profundal sediment sites, respectively. Determination of 701-bp sequences for 16S rDNA clones representing each cluster confirmed this delineation. A PCR-restriction fragment length polymorphism (RFLP) system was developed that enabled identification of clones containing
N. europaea
and
N. eutropha
16S rDNA sequences, including subclasses therein. It proved possible to analyze 16S rDNA amplified directly from sediment samples to determine the relative abundance of each species compared with that of the other.
N. europaea
and
N. eutropha
are very closely related, and direct evidence for their presence in lake systems is limited. The correlation of each species with a distinct spatial location in sediment is an unusual example of niche adaptation by two genotypically similar bacteria. Their occurrence and relative distribution can now be routinely monitored in relation to environmental variation by the application of PCR-RFLP analysis.
Combined Effects of pH and Sugar on Growth Rate of
Zygosaccharomyces rouxii
, a Bakery Product Spoilage Yeast
Authors: Membré Jeanne-Marie , Kubaczka Martine , Chéné Christine
Published: December 2019

The effects of citric acid-modified pH (pH 2.5, 2.75, 3, 3.5, 4, 4.5, 5, and 5.5) and a 30% glucose–70% sucrose mixture (300, 400, 500, 600, 700, 800, 875, and 900 g/liter) on an osmophilic yeast,
Zygosaccharomyces rouxii
, were determined by using synthetic medium. One hundred experiments were carried out; 50-ml culture flasks were inoculated with 10
3
CFU ml
−1
by using a collection strain and a wild-type strain cocktail. The biomass was measured by counting cell colonies, and growth curves were fitted by using a Baranyi equation. The growth rate decreased linearly with sugar concentration, while the effect of pH was nonlinear. Indeed, the optimal pH range was found to be pH 3.5 to 5, and pH 2.5 resulted in a 30% reduction in the growth rate. Finally, we evaluated the performance of two nonlinear predictive models developed previously to describe bacterial contamination. Equations derived from the Rosso and Ratkowsky models gave similar results; however, the model that included dimensionless terms based on the Ratkowsky equation was preferred because it contained fewer estimated parameters and also because biological interpretation of the results was easier.
Molecular Characterization of Laccase Genes from the Basidiomycete
Coprinus cinereus
and Heterologous Expression of the Laccase Lcc1
Authors: Yaver Debbie S. , Overjero Maria Del Carmen , Xu Feng , Nelson Beth A. , Brown Kim M. , Halkier Torben , Bernauer Sheryl , Brown Stephen H. , Kauppinen Sakari
Published: December 2019

A laccase from
Coprinus cinereus
is active at alkaline pH, an essential property for some potential applications. We cloned and sequenced three laccase genes (
lcc1
,
lcc2
, and
lcc3
) from the ink cap basidiomycete
C. cinereus
. The
lcc1
gene contained 7 introns, while both
lcc2
and
lcc3
contained 13 introns. The predicted mature proteins (Lcc1 to Lcc3) are 58 to 80% identical at the amino acid level. The predicted Lcc1 contains a 23-amino-acid C-terminal extension rich in arginine and lysine, suggesting that C-terminal processing may occur during its biosynthesis. We expressed the Lcc1 protein in
Aspergillus oryzae
and purified it. The Lcc1 protein as expressed in
A. oryzae
has an apparent molecular mass of 66 kDa on sodium dodecyl sulfate-polyacrylamide gel electrophoresis and absorption maxima at 278 and 614 nm. Based on the N-terminal protein sequence of the laccase, a 4-residue propeptide was processed during the maturation of the enzyme. The dioxygen specificity of the laccase showed an apparent

K
m

of 21 ± 2 μM and a catalytic constant of 200 ± 10 min
−1
for O
2
with 2,2′-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) as the reducing substrate at pH 5.5. Lcc1 from
A. oryzae
may be useful in industrial applications. This is the first report of a basidiomycete laccase whose biosynthesis involves both N-terminal and C-terminal processing.
Screening of Probiotic Activities of Forty-Seven Strains of
Lactobacillus
spp. by In Vitro Techniques and Evaluation of the Colonization Ability of Five Selected Strains in Humans
Authors: Jacobsen C. N. , Rosenfeldt Nielsen V. , Hayford A. E. , Møller P. L. , Michaelsen K. F. , Pærregaard A. , Sandström B. , Tvede M. , Jakobsen M.
Published: December 2019

The probiotic potential of 47 selected strains of
Lactobacillus
spp. was investigated. The strains were examined for resistance to pH 2.5 and 0.3% oxgall, adhesion to Caco-2 cells, and antimicrobial activities against enteric pathogenic bacteria in model systems. From the results obtained in vitro, five strains,
Lactobacillus rhamnosus
19070-2,
L. reuteri
DSM 12246,
L. rhamnosus
LGG,
L. delbrueckii
subsp.
lactis
CHCC 2329, and
L. casei
subsp.
alactus
CHCC 3137, were selected for in vivo studies. The daily consumption by 12 healthy volunteers of two doses of 10
10
freeze-dried bacteria of the selected strains for 18 days was followed by a washout period of 17 days. Fecal samples were taken at days 0 and 18 and during the washout period at days 5 and 11.
Lactobacillus
isolates were initially identified by API 50CHL and internal transcribed spacer PCR, and their identities were confirmed by restriction enzyme analysis in combination with pulsed-field gel electrophoresis. Among the tested strains,
L. rhamnosus
19070-2,
L. reuteri
DSM 12246, and
L. rhamnosus
LGG were identified most frequently in fecal samples; they were found in 10, 8, and 7 of the 12 samples tested during the intervention period, respectively, whereas reisolations were less frequent in the washout period. The bacteria were reisolated in concentrations from 10
5
to 10
8
cells/g of feces. Survival and reisolation of the bacteria in vivo appeared to be linked to pH tolerance, adhesion, and antimicrobial properties in vitro.
Bacterial Adhesion at Synthetic Surfaces
Authors: Cunliffe D. , Smart C. A. , Alexander C. , Vulfson E. N.
Published: December 2019

A systematic investigation into the effect of surface chemistry on bacterial adhesion was carried out. In particular, a number of physicochemical factors important in defining the surface at the molecular level were assessed for their effect on the adhesion of
Listeria monocytogenes
,
Salmonella typhimurium
,
Staphylococcus aureus
, and
Escherichia coli
. The primary experiments involved the grafting of groups varying in hydrophilicity, hydrophobicity, chain length, and chemical functionality onto glass substrates such that the surfaces were homogeneous and densely packed with functional groups. All of the surfaces were found to be chemically well defined, and their measured surface energies varied from 15 to 41 mJ · m
−2
. Protein adsorption experiments were performed with
3
H-labelled bovine serum albumin and cytochrome
c
prior to bacterial attachment studies. Hydrophilic uncharged surfaces showed the greatest resistance to protein adsorption; however, our studies also showed that the effectiveness of poly(ethyleneoxide) (PEO) polymers was not simply a result of its hydrophilicity and molecular weight alone. The adsorption of the two proteins approximately correlated with short-term cell adhesion, and bacterial attachment for
L. monocytogenes
and
E. coli
also correlated with the chemistry of the underlying substrate. However, for
S. aureus
and
S. typhimurium
a different pattern of attachment occurred, suggesting a dissimilar mechanism of cell attachment, although high-molecular-weight PEO was still the least-cell-adsorbing surface. The implications of this for in vivo attachment of cells suggest that hydrophilic passivating groups may be the best method for preventing cell adsorption to synthetic substrates provided they can be grafted uniformly and in sufficient density at the surface.
Protein-Mediated Adhesion of the Dissimilatory Fe(III)-Reducing Bacterium
Shewanella alga
BrY to Hydrous Ferric Oxide
Authors: Caccavo Frank
Published: December 2019

The rate and extent of bacterial Fe(III) mineral reduction are governed by molecular-scale interactions between the bacterial cell surface and the mineral surface. These interactions are poorly understood. This study examined the role of surface proteins in the adhesion of
Shewanella alga
BrY to hydrous ferric oxide (HFO). Enzymatic degradation of cell surface polysaccharides had no effect on cell adhesion to HFO. The proteolytic enzymes
Streptomyces griseus
protease and chymotrypsin inhibited the adhesion of
S. alga
BrY cells to HFO through catalytic degradation of surface proteins. Trypsin inhibited
S. alga
BrY adhesion solely through surface-coating effects. Protease and chymotrypsin also mediated desorption of adhered
S. alga
BrY cells from HFO while trypsin did not mediate cell desorption. Protease removed a single peptide band that represented a protein with an apparent molecular mass of 50 kDa. Chymotrypsin removed two peptide bands that represented proteins with apparent molecular masses of 60 and 31 kDa. These proteins represent putative HFO adhesion molecules.
S. alga
BrY adhesion was inhibited by up to 46% when cells were cultured at sub-MICs of chloramphenicol, suggesting that protein synthesis is necessary for adhesion. Proteins extracted from the surface of
S. alga
BrY cells inhibited adhesion to HFO by up to 41%. A number of these proteins bound specifically to HFO, suggesting that a complex system of surface proteins mediates
S. alga
BrY adhesion to HFO.
Adhesion of Biodegradative Anaerobic Bacteria to Solid Surfaces
Authors: van Schie Paula M. , Fletcher Madilyn
Published: December 2019

In order to exploit the ability of anaerobic bacteria to degrade certain contaminants for bioremediation of polluted subsurface environments, we need to understand the mechanisms by which such bacteria partition between aqueous and solid phases, as well as the environmental conditions that influence partitioning. We studied four strictly anaerobic bacteria,
Desulfomonile tiedjei
,
Syntrophomonas wolfei
,
Syntrophobacter wolinii
, and
Desulfovibrio
sp. strain G11, which theoretically together can constitute a tetrachloroethylene- and trichloroethylene-dechlorinating consortium. Adhesion of these organisms was evaluated by microscopic determination of the numbers of cells that attached to glass coverslips exposed to cell suspensions under anaerobic conditions. We studied the effects of the growth phase of the organisms on adhesion, as well as the influence of electrostatic and hydrophobic properties of the substratum. Results indicate that
S. wolfei
adheres in considerably higher numbers to glass surfaces than the other three organisms. Starvation greatly decreases adhesion of
S. wolfei
and
Desulfovibrio
sp. strain G11 but seems to have less of an effect on the adhesion of the other bacteria. The presence of Fe
3+
on the substratum, which would be electropositive, significantly increased the adhesion of
S. wolfei
, whereas the presence of silicon hydrophobic groups decreased the numbers of attached cells of all species. Measurements of transport of cells through hydrophobic-interaction and electrostatic-interaction columns indicated that all four species had negatively charged cell surfaces and that
D. tiedjei
and
Desulfovibrio
sp. strain G11 possessed some hydrophobic cell surface properties. These findings are an early step toward understanding the dynamic attachment of anaerobic bacteria in anoxic environments.
Substrate Uptake by Uncultured Bacteria from the Genus
Achromatium
Determined by Microautoradiography
Authors: Gray N. D. , Howarth R. , Pickup R. W. , Jones J. Gwyn , Head I. M.
Published: December 2019

Microautoradiography was used to investigate substrate uptake by natural communities of uncultured bacteria from the genus
Achromatium
. Studies of the uptake of
14
C-labelled substrates demonstrated that
Achromatium
cells from freshwater sediments were able to assimilate
14
C from bicarbonate, acetate, and protein hydrolysate; however,
14
C-labelled glucose was not assimilated. The pattern of substrate uptake by
Achromatium
spp. was therefore similar to those of a number of other freshwater and marine sulfur-oxidizing bacteria. Different patterns of radiolabelled bicarbonate uptake were noted for
Achromatium
communities from different geographical locations and indicated that one community (Rydal Water) possessed autotrophic potential, while the other (Hell Kettles) did not. Furthermore, the patterns of organic substrate uptake within a single population suggested that physiological diversity existed in natural communities of
Achromatium
. These observations are consistent with and may relate to the phylogenetic diversity observed in
Achromatium
communities. Incubation of
Achromatium
-bearing sediment cores from Rydal Water with
35
S-labelled sulfate in the presence and absence of sodium molybdate demonstrated that this bacterial population was capable of oxidizing sulfide to intracellular elemental sulfur. This finding supported the role of
Achromatium
in the oxidative component of a tightly coupled sulfur cycle in Rydal Water sediment. The oxidation of sulfide to sulfur and ultimately to sulfate by
Achromatium
cells from Rydal Water sediment is consistent with an ability to conserve energy from sulfide oxidation.
Thermicanus aegyptius
gen. nov., sp. nov., Isolated from Oxic Soil, a Fermentative Microaerophile That Grows Commensally with the Thermophilic Acetogen
Moorella thermoacetica
Authors: Gößner Anita S. , Devereux Richard , Ohnemüller Nadja , Acker Georg , Stackebrandt Erko , Drake Harold L.
Published: December 2019

A thermophilic, fermentative microaerophile (ET-5b) and a thermophilic acetogen (ET-5a) were coisolated from oxic soil obtained from Egypt. The 16S rRNA gene sequence of ET-5a was 99.8% similar to that of the classic acetogen
Moorella thermoacetica
. Further analyses confirmed that ET-5a was a new strain of
M. thermoacetica
. For ET-5b, the nearest 16S rRNA gene sequence similarity value to known genera was approximately 88%. ET-5b was found to be a motile rod with a genomic G+C content of 50.3 mol%. Cells were weakly gram positive and lacked spores. Growth was optimal at 55 to 60°C and pH 6.5 to 7.0. ET-5b grew under both oxic and anoxic conditions, but growth was erratic under atmospheric concentrations of O
2
. Utilizable substrates included oligosaccharides and monosaccharides. Acetate, formate, and succinate supported growth only under oxic conditions. Saccharides yielded succinate, lactate, ethanol, acetate, formate, and H
2
under anoxic conditions; fermentation products were also formed under oxic conditions. A new genus is proposed, the type strain being
Thermicanus aegyptius
ET-5b gen. nov., sp. nov. (DSMZ 12793).
M. thermoacetica
ET-5a (DSMZ 12797) grew commensally with
T. aegyptius
ET-5b on oligosaccharides via the interspecies transfer of H
2
formate, and lactate. In support of this interaction, uptake hydrogenase and formate dehydrogenase specific activities were fundamentally greater in
M. thermoacetica
ET-5a than in
T. aegyptius
ET-5b. These results demonstrate that (i) soils subject to high temperatures harbor uncharacterized thermophilic microaerophiles, (ii) the classic acetogen
M. thermoacetica
resides in such soils, and (iii) trophic links between such soil bacteria might contribute to their in situ activities.
Display of Polyhistidine Peptides on the
Escherichia coli
Cell Surface by Using Outer Membrane Protein C as an Anchoring Motif
Authors: Xu Zhaohui , Lee Sang Yup
Published: December 2019

A novel cell surface display system was developed by employing
Escherichia coli
outer membrane protein C (OmpC) as an anchoring motif. Polyhistidine peptides consisting of up to 162 amino acids could be successfully displayed on the seventh exposed loop of OmpC. Recombinant cells displaying polyhistidine could adsorb up to 32.0 μmol of Cd
2+
per g (dry weight) of cells.
Effect of Various Ions, pH, and Osmotic Pressure on Oxidation of Elemental Sulfur by
Thiobacillus thiooxidans
Authors: Suzuki Isamu , Lee Douglas , Mackay Byron , Harahuc Lesia , Oh Jae Key
Published: December 2019

The oxidation of elemental sulfur by
Thiobacillus thiooxidans
was studied at pH 2.3, 4.5, and 7.0 in the presence of different concentrations of various anions (sulfate, phosphate, chloride, nitrate, and fluoride) and cations (potassium, sodium, lithium, rubidium, and cesium). The results agree with the expected response of this acidophilic bacterium to charge neutralization of colloids by ions, pH-dependent membrane permeability of ions, and osmotic pressure.
Long-Term Survival of Shiga Toxin-Producing
Escherichia coli
O26, O111, and O157 in Bovine Feces
Authors: Fukushima Hiroshi , Hoshina Ken , Gomyoda Manabu
Published: December 2019

Cattle are an important reservoir of Shiga toxin-producing
Escherichia coli
(STEC) O26, O111, and O157. The fate of these pathogens in bovine feces at 5, 15, and 25°C was examined. The feces of a cow naturally infected with STEC O26:H11 and two STEC-free cows were studied. STEC O26, O111, and O157 were inoculated into bovine feces at 10
1
, 10
3
, and 10
5
CFU/g. All three pathogens survived at 5 and 25°C for 1 to 4 weeks and at 15°C for 1 to 8 weeks when inoculated at the low concentration. On samples inoculated with the middle and high concentrations, O26, O111, and O157 survived at 25°C for 3 to 12 weeks, at 15°C for 1 to 18 weeks, and at 5°C for 2 to 14 weeks, respectively. Therefore, these pathogens can survive in feces for a long time, especially at 15°C. The surprising long-term survival of STEC O26, O111, and O157 in bovine feces shows that such feces are a potential vehicle for transmitting not only O157 but also O26 and O111 to cattle, food, and the environment. Appropriate handling of bovine feces is emphasized.
Fungi from Geothermal Soils in Yellowstone National Park
Authors: Redman Regina S. , Litvintseva Anastassia , Sheehan Kathy B. , Henson Joan M. , Rodriguez Rusty J.
Published: December 2019

Geothermal soils near Amphitheater Springs in Yellowstone National Park were characterized by high temperatures (up to 70°C), high heavy metal content, low pH values (down to pH 2.7), sparse vegetation, and limited organic carbon. From these soils we cultured 16 fungal species. Two of these species were thermophilic, and six were thermotolerant. We cultured only three of these species from nearby cool (0 to 22°C) soils. Transect studies revealed that higher numbers of CFUs occurred in and below the root zone of the perennial plantDichanthelium lanuginosum(hot springs panic grass). The dynamics of fungal CFUs in geothermal soil and nearby nongeothermal soil were investigated for 12 months by examining soil cores and in situ mesocosms. For all of the fungal species studied, the temperature of the soil from which the organisms were cultured corresponded with their optimum axenic growth temperature.
Removal of Mercury from Chloralkali Electrolysis Wastewater by a Mercury-Resistant
Pseudomonas putida
Strain
Authors: von Canstein H. , Li Y. , Timmis K. N. , Deckwer W.-D. , Wagner-Döbler I.
Published: December 2019

A mercury-resistant bacterial strain which is able to reduce ionic mercury to metallic mercury was used to remediate in laboratory columns mercury-containing wastewater produced during electrolytic production of chlorine. Factory effluents from several chloralkali plants in Europe were analyzed, and these effluents contained total mercury concentrations between 1.6 and 7.6 mg/liter and high chloride concentrations (up to 25 g/liter) and had pH values which were either acidic (pH 2.4) or alkaline (pH 13.0). A mercury-resistant bacterial strain,
Pseudomonas putida
Spi3, was isolated from polluted river sediments. Biofilms of
P. putida
Spi3 were grown on porous carrier material in laboratory column bioreactors. The bioreactors were continuously fed with sterile synthetic model wastewater or nonsterile, neutralized, aerated chloralkali wastewater. We found that sodium chloride concentrations up to 24 g/liter did not inhibit microbial mercury retention and that mercury concentrations up to 7 mg/liter could be treated with the bacterial biofilm with no loss of activity. When wastewater samples from three different chloralkali plants in Europe were used, levels of mercury retention efficiency between 90 and 98% were obtained. Thus, microbial mercury removal is a potential biological treatment for chloralkali electrolysis wastewater.
Lignin-Modifying Enzymes of the White Rot Basidiomycete
Ganoderma lucidum
Authors: D’Souza Trevor M. , Merritt Carlos S. , Reddy C. Adinarayana
Published: December 2019

Ganoderma lucidum
, a white rot basidiomycete widely distributed worldwide, was studied for the production of the lignin-modifying enzymes laccase, manganese-dependent peroxidase (MnP), and lignin peroxidase (LiP). Laccase levels observed in high-nitrogen (HN; 24 mM N) shaken cultures were much greater than those seen in low-nitrogen (2.4 mM N), malt extract, or wood-grown cultures and those reported for most other white rot fungi to date. Laccase production was readily seen in cultures grown with pine or poplar (100-mesh-size ground wood) as the sole carbon and energy source. Cultures containing both pine and poplar showed 5- to 10-fold-higher levels of laccase than cultures containing pine or poplar alone. Since syringyl units are structural components important in poplar lignin and other hardwoods but much less so in pine lignin and other softwoods, pine cultures were supplemented with syringic acid, and this resulted in laccase levels comparable to those seen in pine-plus-poplar cultures. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of concentrated extracellular culture fluid from HN cultures showed two laccase activity bands (
M
r
of 40,000 and 66,000), whereas isoelectric focusing revealed five major laccase activity bands with estimated pIs of 3.0, 4.25, 4.5, 4.8, and 5.1. Low levels of MnP activity (∼100 U/liter) were detected in poplar-grown cultures but not in cultures grown with pine, with pine plus syringic acid, or in HN medium. No LiP activity was seen in any of the media tested; however, probing the genomic DNA with the LiP cDNA (
CLG4
) from the white rot fungus
Phanerochaete chrysosporium
showed distinct hybridization bands suggesting the presence of
lip
-like sequences in
G. lucidum
.
Listeria monocytogenes
Scott A: Cell Surface Charge, Hydrophobicity, and Electron Donor and Acceptor Characteristics under Different Environmental Growth Conditions
Authors: Briandet Romain , Meylheuc Thierry , Maher Catherine , Bellon-Fontaine Marie Noëlle
Published: December 2019

We determined the variations in the surface physicochemical properties of
Listeria monocytogenes
Scott A cells that occurred under various environmental conditions. The surface charges, the hydrophobicities, and the electron donor and acceptor characteristics of
L. monocytogenes
Scott A cells were compared after the organism was grown in different growth media and at different temperatures; to do this, we used microelectrophoresis and the microbial adhesion to solvents method. Supplementing the growth media with glucose or lactic acid affected the electrical, hydrophobic, and electron donor and acceptor properties of the cells, whereas the growth temperature (37, 20, 15, or 8°C) primarily affected the electrical and electron donor and acceptor properties. The nonlinear effects of the growth temperature on the physicochemical properties of the cells were similar for cells cultivated in two different growth media, but bacteria cultivated in Trypticase soy broth supplemented with 6 g of yeast extract per liter (TSYE) were slightly more hydrophobic than cells cultivated in brain heart infusion medium (
P
< 0.05). Adhesion experiments conducted with
L. monocytogenes
Scott A cells cultivated in TSYE at 37, 20, 15, and 8°C and then suspended in a sodium chloride solution (1.5 × 10
−1
or 1.5 × 10
−3
M NaCl) confirmed that the cell surface charge and the electron donor and acceptor properties of the cells had an influence on their attachment to stainless steel.
Agar Underlay Method for Recovery of Sublethally Heat-Injured Bacteria
Authors: Kang D. H. , Siragusa G. R.
Published: December 2019

A method of recovering sublethally heat-injured bacteria was developed. The procedure (termed the agar underlay method) uses a nonselective agar underlaid with a selective medium. In a two-chambered petri dish, the Lutri plate (LP), a nonselective agar is inoculated with a population of sublethally heat-injured bacteria. After a 2-h repair incubation period, selective agar is added to the bottom chamber of the LP and incubated. By diffusing through the nonselective top agar, selective agents from the underlay medium impart selectivity to the system. By the agar underlay method, recovery rates of the heat-injured food-borne pathogens
Escherichia coli
O157:H7 and
Salmonella typhimurium
were not different (
P
> 0.05) from recovery rates determined with nonselective media. Sublethally heat-injured cells (60°C for 1.5 min in buffer or 80°C for 30 s on meat surfaces) grew and produced a typical colony morphology and color reaction when the agar underlay procedure was used with the appropriate respective selective agars. Unlike agar overlay methods for injury repair, the agar underlay procedure allows the typical selective-medium colony morphology to develop and allows colonies to be more easily picked for further characterization. Higher recovery rates of heat-injured fecal enterococci from bovine fecal samples and total coliforms from animal waste lagoons were obtained by the agar underlay method with selective agars than by direct plating on the respective selective media.
The Presence of Salt and a Curing Agent Reduces Bacteriocin Production by
Lactobacillus sakei
CTC 494, a Potential Starter Culture for Sausage Fermentation
Authors: Leroy Frédéric , de Vuyst Luc
Published: December 2019

The specific conditions in the batter of raw fermented sausages may reduce the efficiency of bacteriocin-producing starter cultures. In this work, using in vitro fermentation, we found that sodium chloride and sodium nitrite interfere with the growth of
Lactobacillus sakei
CTC 494, an organism which produces the antilisterial bacteriocin sakacin K. Because sakacin K production follows primary metabolite kinetics, a decrease in cell formation resulted in a decrease in sakacin K production as well. Sodium chloride dramatically influenced bacteriocin production by decreasing both biomass production and specific bacteriocin production. Sodium nitrite, however, had no effect on specific bacteriocin production and decreased bacteriocin production only because of its effect on cell growth. Moreover, sodium nitrite enhanced the toxic effect of lactic acid on bacterial growth.
Utilization of Heterologous Siderophores Enhances Levels of Iron Available to
Pseudomonas putida
in the Rhizosphere
Authors: Loper Joyce E. , Henkels Marcella D.
Published: December 2019

Pseudomonas
spp. have the capacity to utilize siderophores produced by diverse species of bacteria and fungi, and the present study was initiated to determine if siderophores produced by rhizosphere microorganisms enhance the levels of iron available to a strain of
Pseudomonas putida
in this natural habitat. We used a previously described transcriptional fusion (
pvd
-
inaZ
) between an iron-regulated promoter (
pvd
) and the ice nucleation reporter gene (
inaZ
) to detect alterations in iron availability to
P. putida
. Ice nucleation activity (INA) expressed from the
pvd
-
inaZ
fusion by
P. putida
N1R or N1R Pvd
−
, a derivative deficient in the production of a pyoverdine siderophore, was inversely related to the concentration of ferric citrate in a culture medium. In culture, INA expressed by N1R Pvd
−
(
pvd
-
inaZ
) was reduced in the presence of the ferric complex of pseudobactin-358, a pyoverdine siderophore produced by
P. putida
WCS358 that can be utilized as a source of iron by N1R Pvd
−
. In the rhizosphere of cucumbers grown in sterilized soil, N1R Pvd
−
(
pvd
-
inaZ
) expressed INA, indicating that iron availability was sufficiently low in that habitat to allow transcription of the iron-regulated
pvd
promoter. Coinoculation with WCS358 or N1R significantly decreased INA expressed by N1R Pvd
−
(
pvd
-
inaZ
) in the rhizosphere, whereas coinoculation with a pyoverdine-deficient mutant of WCS358 did not reduce INA expressed by N1R Pvd
−
(
pvd
-
inaZ
). These results indicate that iron availability to N1R Pvd
−
(
pvd
-
inaZ
) in the rhizosphere was enhanced by the presence of another strain of
P. putida
that produces a pyoverdine that N1R Pvd
−
(
pvd
-
inaZ
) was able to utilize as a source of iron. In culture, strain N1R Pvd
−
also utilized ferric complexes of the siderophores enterobactin and aerobactin as sources of iron. In the rhizosphere of cucumbers grown in sterilized soil, INA expressed by N1R Pvd
−
(
pvd
-
inaZ
) was reduced in the presence of strains of
Enterobacter cloacae
that produced enterobactin, aerobactin, or both siderophores, but INA expressed by N1R Pvd
−
(
pvd
-
inaZ
) was not altered in the presence of a mutant of
E. cloacae
deficient in both enterobactin and aerobactin production. Therefore, the iron status of
P. putida
was altered by siderophores produced by an unrelated bacterium coinhabiting the rhizosphere. Finally, we demonstrated that INA expressed by N1R containing
pvd
-
inaZ
in the rhizosphere differed between plants grown in sterilized versus nonsterilized field soil. The results of this study demonstrate that (i)
P. putida
expresses genes for pyoverdine production and uptake in the rhizosphere, but the level of gene expression is influenced by other bacteria that coexist with
P. putida
in this habitat, and (ii) diverse groups of microorganisms can alter the availability of chemical resources in microbial habitats on root surfaces.
Induction and Transcription Studies of the Dextransucrase Gene in
Leuconostoc mesenteroides
NRRL B-512F
Authors: Quirasco M. , López-Munguía A. , Remaud-Simeon M. , Monsan P. , Farrés A.
Published: December 2019

Dextransucrase production by
Leuconostoc mesenteroides
NRRL B-512F in media containing carbon sources other than sucrose is reported for the first time. Dextransucrases were analyzed by gel electrophoresis and by an in situ activity assay. Their polymers and acceptor reaction products were also compared by
13
C nuclear magnetic resonance and high-performance liquid chromatography techniques, respectively. From these analyses, it was found that, independently of the carbon source,
L. mesenteroides
NRRL B-512F produced dextransucrases of the same size and product specificity. The 5′ ends of dextransucrase mRNAs isolated from cells grown under different culture conditions were identical. Based on this evidence, we conclude that dextransucrases obtained from cells grown on the various carbon sources result from the transcription of the same gene. The control of expression occurs at this level. The low dextransucrase yields from cultures in
d
-glucose or
d
-fructose and the enhancement of dextransucrase gene transcription in the presence of sucrose suggest that an activating phenomenon may be involved in the expression mechanism. Dextransucrase mRNA has a size of approximately 4.8 kb, indicating that the gene is located in a monocistronic operon. The transcription start point was localized 34 bp upstream from the ATG start codon. The −10 and −35 sequences found, TATAAT and TTTACA, were highly homologous to the only glycosyltransferase promoter sequence reported for lactic acid bacteria.