bluuebunny/crossref_metadata_2025 huggingface dataset:
Promoting a healthy childcare environment: A focus group study of childcare providers' perspectives
Authors: Park So Hyun , Min Haeyoung
Published: December 2019

Establishing healthy habits in the childcare setting for children aged 2–5 is pivotal for their healthy development and obesity prevention. The purpose of this qualitative study was to explore the perceptions and practices that childcare providers demonstrate when they promote healthy eating and physical activity among preschoolers. Four focus group interviews with 16 childcare providers were conducted in northwest Florida. A semistructured focus group guide was used. Discussions were audiorecorded and transcribed verbatim. Content analysis was performed to analyze the data with the use of ATLAS.ti software. This study followed the COREQ guidelines. Three major themes emerged from the data: (i) efforts to encourage children to eat healthfully and engage in physical activity; (ii) challenges in promoting healthy behaviors among children; and (iii) the need for more childcare provider training and parental involvement to encourage healthy eating. Regular childcare provider training and parental involvement are needed to establish a healthy environment for preschoolers.
Illuminating the gendered nature of health‐promoting activities among nursing staff in forensic psychiatric care
Authors: Kumpula Esa , Gustafsson Lena‐Karin , Ekstrand Per
Published: December 2019

When people in Sweden are sentenced and handed over to forensic psychiatric care (FPC), the authorities have overall responsibility for their health recovery. How nursing staff construct gender through their relations in this context affects their understanding of health promotion activities. The aim of this study was to illuminate, using a gender perspective, the understanding of nursing staff with respect to health promotion activities for patients. Four focus group interviews were conducted with nursing staff in two FPC clinics in Sweden. The study has a qualitative inductive design with an ethnographic approach. This study sheds new light on FPC in which its dual goals of protecting society and providing care are viewed from a gender perspective. When relationships within the nursing staff group and the nurse–patient relationship are justified by the goal of protecting society, gender becomes invisible. This might cause patients' individual conditions and needs for certain types of activities to go unnoticed. One of the implications of ignoring gender relations in nursing staff health promotion activities is that it risks contributing to gender stereotypes which impact on the nurse–patient relationship and the quality of care.
Forecasting semi‐arid biome shifts in the Anthropocene
Authors: Kulmatiski Andrew , Yu Kailiang , Mackay D. Scott , Holdrege Martin C. , Staver Ann Carla , Parolari Anthony J. , Liu Yanlan , Majumder Sabiha , Trugman Anna T.
Published: December 2019

Shrub encroachment, forest decline and wildfires have caused large‐scale changes in semi‐arid vegetation over the past 50 years. Climate is a primary determinant of plant growth in semi‐arid ecosystems, yet it remains difficult to forecast large‐scale vegetation shifts (i.e. biome shifts) in response to climate change. We highlight recent advances from four conceptual perspectives that are improving forecasts of semi‐arid biome shifts. Moving from small to large scales, first, tree‐level models that simulate the carbon costs of drought‐induced plant hydraulic failure are improving predictions of delayed‐mortality responses to drought. Second, tracer‐informed water flow models are improving predictions of species coexistence as a function of climate. Third, new applications of ecohydrological models are beginning to simulate small‐scale water movement processes at large scales. Fourth, remotely‐sensed measurements of plant traits such as relative canopy moisture are providing early‐warning signals that predict forest mortality more than a year in advance. We suggest that a community of researchers using modeling approaches (e.g. machine learning) that can integrate these perspectives will rapidly improve forecasts of semi‐arid biome shifts. Better forecasts can be expected to help prevent catastrophic changes in vegetation states by identifying improved monitoring approaches and by prioritizing high‐risk areas for management.
Staging International Environmental Justice: The International Monsanto Tribunal
Authors: Prete Giovanni , Cournil Christel
Published: December 2019

Opinions tribunals dealing with environmental issues have multiplied over the last several years as a consequence of the rise of international environmental law and its promotion by international networks. Drawing on an ethnographic investigation of one of those tribunals—the International Monsanto Tribunal— this article reflects on the many objectives they often pursue: strengthening political positions, publicizing environmental and health social struggles, and promoting legal theories. In our case, we show that articulating those objectives involved intense work to stage the tribunal's legitimacy. We analyze this work and how it was put to the test during and after the sessions of the tribunal. Our article broadly suggests that environmental opinion tribunals are political arenas where rights and identities are not only asserted but also negotiated and legitimized.
Representing Religious Discrimination at the Margins: Temporalities and “Appropriate” Identities of the State in Turkey
Authors: Özkan Nazlı
Published: December 2019

In Turkey during Ramadan in the 2000s, a group of Sunni Muslims attacked a Kurdish Alevi family that was not fasting. The Alevis, Turkey's second‐largest religious group, do not fast during Ramadan, creating tensions in mixed neighborhoods in which Sunni Islam is the reigning religion. This article analyzes how an Alevi‐run television station covered this event, from its location in a provincial town to related protests in Istanbul, to show how minority media may partake in the reproduction of the dominant norms that perpetuate such discriminatory acts. They do so by producing a “presentist” temporal perspective on discrimination that prioritizes more contemporary problems of the minoritized community at the expense of a longer history of structural violence. In this case, this temporalization portrayed Alevis as exclusively Turks, neglecting, therefore, Kurdish Alevis, and it disguised the state's long‐term involvement in Alevis’ marginalization. This article presents an alternative perspective on minority media in anthropology, which often interprets state alignments as strategic leverage to defend community rights. The article argues that minority media producers can also be strategic in their alignments with their communities and may use this alignment as a façade when securing their ties with states.
Characterization of two novel HLA‐DQA1*05:05:01 variants, identified in Brazilian individuals
Authors: Vianna Romulo , Secco Danielle , Santos Angela , Porto Luís Cristóvão
Published: December 2019

Characterization of two novel HLA‐DQA1*05:05:01 variants, HLA‐DQA1*05:05:01:24 and ‐DQA1*05:05:01:25.
Prevalence and impact of cholecystitis on outcome in dogs with gallbladder mucocele
Authors: Rogers Erin , Jaffey Jared A. , Graham Amber , Hostnik Eric T. , Jacobs Casandra , Fox‐Alvarez William , Van Eerde Eric , Arango John , Williams Fred , DeClue Amy E.
Published: December 2019

Background

Gallbladder mucocele is a potentially life‐threatening extrahepatic biliary disease in dogs. The primary aims of this study were to evaluate the prevalence of cholecystitis in dogs with gross and histopathologically confirmed gallbladder mucocele and to investigate if there is an association between cholecystitis, including its subtypes (eg, acute, acute on chronic, with necrosis, chronic), and survival. Our secondary objective was to evaluate if there is an association between cholecystitis and intraoperative bacteriological culture positivity.

Key Findings

Two hundred nineteen dogs with gallbladder mucocele were included in this multi‐institutional retrospective study, of which 63 (28.8%) dogs had histopathological evidence of cholecystitis. The most common forms of cholecystitis were acute on chronic (n = 22/63, 34.9%) and with necrosis (n = 20, 31.7%). Thirty‐one (14.1%) dogs had growth of at least 1 bacterial isolate; however, 88.7% had antimicrobials administered within the 48 hours before surgery or intraoperatively. There was not an association between cholecystitis or its subtypes and survival. Furthermore, there was not an association between cholecystitis and intraoperative bacteriological culture positivity. A total of 38 (17.4%) dogs either died or were euthanized during hospitalization.

Significance

Cholecystitis is a common comorbidity in dogs with gallbladder mucocele but was not associated with decreased survival.
Isolation Frequency Switchable MIMO Antenna for PCS, WIMAX and WLAN Application
Authors: Jabire Adamu Halilu , Abdu Anas , Saminu Sani , Sadiq Abubakar Muhammad , Adamu Mohammed Jajere
Published: December 2019

In this study, a lumped component based frequency reconfigurable multiple-input-multiple-output (MIMO) receiving wire design is presented. The proposed antenna is composed of a planar structure in form of F-shaped together with a slotted and defected ground structure for bandwidth and isolation enhancement. The MIMO antenna operates in six frequencies upon the state of the four lumped element switches. The proposed receiving wire design exhibits a multiband frequency reconfigurable characteristics from 1-7GHz with isolation of more than 14dB for the whole band, with efficiency of about 75%. The MIMO antenna’s behavior in terms of ratio of square root of the sum of power reflected wave to the incident wave (TARC), ECC and CCL are all within the acceptable limits. The design is suitable for personal communication system (PCS), WIMAX and WLAN wireless applications.
Modal Analysis of a Circular Slot Monopole Antenna for UWB application
Authors: Jabire Adamu Halilu , Abdu Anas , Sani Salisu , Saminu Sani , Uba Taura Aliu , Olatunji Obalowu Mohammed
Published: December 2019

In this study, a wideband double slot antenna is introduced. The proposed monopole antenna is designed using the theory of characteristic mode for slot monopoles. A circular shape is used as the initial design stage, to enhance the bandwidth, two circular slots are employed. Four modes have been excited to gain a physical insight and to find out which mode is dominant. The design and analysis were completed utilizing both the time domain and multilayer solver in CST 2017, without considering the feeding port. An antenna model in UWB frequency is constructed. Experimented and simulated results shows that the proposed planar structure has a wide impedance width with good radiation characteristics.
Adults Want to Play Too: Feasibility of an Adult Physical Activity Program Designed to Maximize Enjoyment
Authors: Upham Kent , Auer Brandon J. , Sciamanna Christopher N. , Mowen Andrew J. , Smyth Joshua M. , Conroy David E. , Silvis Matthew , Kraschnewski Jennifer L. , Rovniak Liza S. , Lehman Erik , Kearcher Kalen , Vizzini Maggie , Cesarone Louis
Published: December 2019

Background: Few adults in the United States obtain sufficient physical activity (PA) despite knowledge of the associated health benefits. The current feasibility study examined the feasibility of a novel modified sports intervention designed to promote enjoyment and sustained PA in sedentary adults. Methods: The US adults (N = 22, mean age 39.2 y, male/female percentage 54.5/45.5) in Central Pennsylvania participated in the PlayFit sports program for 60-minute sessions, 2 to 3 times per week, over the course of 10 weeks and 24 game sessions; completing 198 person sessions collectively. Primary outcomes were PA (accelerometry) and intervention satisfaction. Results: Percentage of time in moderate to vigorous activity ranged from 35.0% (volleyball) to 91.2% (ultimate frisbee). Percentage of time spent in vigorous activity ranged from 0.0% (volleyball) to 29.5% (team handball). Satisfaction, based on a 10-point scale with 10 being the most satisfied, ranged from 7.7 (kickball) to 8.7 (floor hockey and soccer). On average, all sports were rated highly, with the majority rated >8.5 and one rated <8.0. Percentage of time spent in the moderate to vigorous range was lower in men than in women (73.2% vs 80.0%, P = .01), but did not differ by age or body mass index. Conclusions: PlayFit is a promising first step in exploring the potential of modified sports programs to enhance population PA levels.
Feeling the α-synuclein strain
Authors: Haigh Cathryn
Published: December 2019

α-synuclein strains may be responsible for the differing disease etiologies of the α-Synucleinopathies.
Performance and Accuracy of Four Open-Source Tools for
In Silico
Serotyping of
Salmonella
spp. Based on Whole-Genome Short-Read Sequencing Data
Authors: Uelze Laura , Borowiak Maria , Deneke Carlus , Szabó István , Fischer Jennie , Tausch Simon H. , Malorny Burkhard
Published: December 2019

Salmonella
spp. are important foodborne pathogens. To reduce the number of infected patients, it is essential to understand which subtypes of the bacteria cause disease outbreaks. Traditionally, characterization of
Salmonella
requires serological testing, a laboratory method by which
Salmonella
isolates can be classified into over 2,600 distinct subtypes, called serovars. Due to recent advances in whole-genome sequencing, many tools have been developed to replace traditional testing methods with computational analysis of genome sequences. It is crucial to validate that these tools, many already in use for routine surveillance, deliver accurate and reliable serovar information. In this study, we set out to compare which of the currently available open-source command-line tools is most suitable to replace serological testing. A thorough evaluation of the differing computational approaches is highly important to ensure the backward compatibility of serotyping data and to maintain comparability between laboratories.
Genetic Determinants of Hydroxycinnamic Acid Metabolism in Heterofermentative Lactobacilli
Authors: Gaur Gautam , Oh Jee-Hwan , Filannino Pasquale , Gobbetti Marco , van Pijkeren Jan-Peter , Gänzle Michael G.
Published: December 2019

The metabolism of secondary plant metabolites, including phenolic compounds, by food-fermenting lactobacilli is a significant contributor to the safety, quality, and nutritional quality of fermented foods. The enzymes mediating hydrolysis, reduction, and decarboxylation of phenolic acid esters and phenolic acids in lactobacilli, however, are not fully characterized. The genomic analyses presented here provide evidence for three novel putative phenolic acid reductases. Matching comparative genomic analyses with phenotypic analysis and quantification of gene expression indicates that two of the three putative phenolic acid reductases, Par1 and HcrF, are involved in reduction of hydroxycinnamic acids to phenylpropionic acids; however, the activity of Par2 may be unrelated to phenolic acids and recognizes other secondary plant metabolites. These findings expand our knowledge on the metabolic potential of lactobacilli and facilitate future studies on activity and substrate specificity of enzymes involved in metabolism of phenolic compounds.
Elemental Sulfur and Thiosulfate Disproportionation by
Desulfocapsa sulfoexigens
sp. nov., a New Anaerobic Bacterium Isolated from Marine Surface Sediment
Authors: Finster Kai , Liesack Werner , Thamdrup Bo
Published: December 2019

A mesophilic, anaerobic, gram-negative bacterium, strain SB164P1, was enriched and isolated from oxidized marine surface sediment with elemental sulfur as the sole energy substrate in the presence of ferrihydrite. Elemental sulfur was disproportionated to hydrogen sulfide and sulfate. Growth was observed exclusively in the presence of a hydrogen sulfide scavenger, e.g., ferrihydrite. In the absence of a scavenger, sulfide and sulfate production were observed but no growth occurred. Strain SB164P1 grew also by disproportionation of thiosulfate and sulfite. With thiosulfate, the growth efficiency was higher in ferrihydrite-supplemented media than in media without ferrihydrite. Growth coupled to sulfate reduction was not observed. However, a slight sulfide production occurred in cultures incubated with formate and sulfate. Strain SB164P1 is the first bacterium described that grows chemolithoautotrophically exclusively by the disproportionation of inorganic sulfur compounds. Comparative 16S rDNA sequencing analysis placed strain SB164P1 into the delta subclass of the class
Proteobacteria
. Its closest relative is
Desulfocapsa thiozymogenes
, and slightly more distantly related are
Desulfofustis glycolicus
and
Desulforhopalus vacuolatus
. This phylogenetic cluster of organisms, together with members of the genus
Desulfobulbus
, forms one of the main lines of descent within the delta subclass of the
Proteobacteria
. Due to the common phenotypic characteristics and the phylogenetic relatedness to
Desulfocapsa thiozymogenes
, we propose that strain SB164P1 be designated the type strain of
Desulfocapsa sulfoexigens
sp. nov.
Effect of Inoculation Techniques and Relative Humidity on the Growth of Molds on the Surfaces of Yellow Layer Cakes
Authors: Fustier Patrick , Lafond Alain , Champagne Claude P. , Lamarche François
Published: December 2019

Four inoculation techniques were compared for initiation of growth on cake surfaces: spot, air cabinet, spray (atomizer), and talc addition methods. Molds were isolated from commercial cakes and were identified as
Aspergillus sydowii
,
Aspergillus ochraceus
,
Penicillium funiculosum
, and
Eurotium herbariorum
. Cake surfaces were inoculated with mold spores and incubated under three equilibrium relative humidity (ERH) levels: 97, 85, and 75%. Random contamination by spores in a ventilated air cabinet was the simplest method of inoculation, but standard deviations in the inoculation rates (20% on a relative scale) were almost twice those observed with the other methods. The spot method was the most reproducible. Cake samples inoculated in the air cabinet had colony counts 10 times lower than those obtained for potato dextrose agar plates at 97% ERH, which was not the case with the spray and talc methods. Growth of molds was much slower in the samples incubated in 75% relative humidity, with all methods. Colony counts were generally similar in systems adjusted at 85 to 97% ERH but were lower for samples incubated at 75% ERH. In comparisons of the shelf life estimates obtained by the various inoculation methods, a correlation coefficient (
r
2
) of 0.70 was obtained between the spot method and the other methods of inoculation, while talc, air cabinet, and spray shelf life data were correlated better (
r
2
≈ 0.97). The spot method appeared to be the method of choice in consideration of ease of use, precision, and the ability to enable the study of the effects of the environment on mold-free shelf life as well as on the rate of growth of molds on cakes.
The
TRI11
Gene of
Fusarium sporotrichioides
Encodes a Cytochrome P-450 Monooxygenase Required for C-15 Hydroxylation in Trichothecene Biosynthesis
Authors: Alexander Nancy J. , Hohn Thomas M. , McCormick Susan P.
Published: December 2019

Several genes in the trichothecene biosynthetic pathway of
Fusarium sporotrichioides
have been shown to reside in a gene cluster. Sequence analysis of a cloned DNA fragment located 3.8 kb downstream from
TRI5
has led to the identification of the
TRI11
gene. The nucleotide sequence of
TRI11
predicts a polypeptide of 492 residues (
M
r
= 55,579) with significant similarity to members of the cytochrome P-450 superfamily. TRI11 is most similar to several fungal cytochromes P-450 (23 to 27% identity) but is sufficiently distinct to define a new cytochrome P-450 gene family, designated
CYP65A1
. Disruption of
TRI11
results in an altered trichothecene production phenotype characterized by the accumulation of isotrichodermin, a trichothecene pathway intermediate. The evidence suggests that
TRI11
encodes a C-15 hydroxylase involved in trichothecene biosynthesis.
Fingerprinting of Cyanobacteria Based on PCR with Primers Derived from Short and Long Tandemly Repeated Repetitive Sequences
Authors: Rasmussen Ulla , Svenning Mette M.
Published: December 2019

The presence of repeated DNA (short tandemly repeated repetitive [STRR] and long tandemly repeated repetitive [LTRR]) sequences in the genome of cyanobacteria was used to generate a fingerprint method for symbiotic and free-living isolates. Primers corresponding to the STRR and LTRR sequences were used in the PCR, resulting in a method which generate specific fingerprints for individual isolates. The method was useful both with purified DNA and with intact cyanobacterial filaments or cells as templates for the PCR. Twenty-three
Nostoc
isolates from a total of 35 were symbiotic isolates from the angiosperm
Gunnera
species, including isolates from the same
Gunnera
species as well as from different species. The results show a genetic similarity among isolates from different
Gunnera
species as well as a genetic heterogeneity among isolates from the same
Gunnera
species. Isolates which have been postulated to be closely related or identical revealed similar results by the PCR method, indicating that the technique is useful for clustering of even closely related strains. The method was applied to nonheterocystus cyanobacteria from which a fingerprint pattern was obtained.
Detection of Toxin-Producing Cyanobacteria by Use of Paramagnetic Beads for Cell Concentration and DNA Purification
Authors: Rudi Knut , Larsen Frank , Jakobsen Kjetill S.
Published: December 2019

Early detection of water blooms caused by potential toxin-producing cyanobacteria is important in environmental monitoring. We present a new nucleic acid-based method for detection of cyanobacteria in water that utilizes the same paramagnetic solid phase (beads) for both bacterial cell concentration and subsequent DNA purification. In the cell concentration step, the beads were attracted to a magnet after cell adsorption (in an alcohol- and salt-containing solution), and the supernatant was removed. For DNA purification, a buffer containing guanidine thiocyanate and Sarkosyl lysed the concentrated cells. The addition of alcohol precipitated the released DNA onto the same solid phase as was used for the cell concentration. Finally, to remove PCR inhibitors, the DNA was washed twice in alcohol while bound to the beads. All of the bead-DNA complex was used in the subsequent PCR amplification. The detection limit, as measured by 16S rDNA PCR amplification, was 50 cells in a 0.5-ml water sample, which is considerably lower than the limit (500 cells/ml) of toxic cyanobacteria tolerated in drinking water (New South Wales Blue-Green Algae Task Force, 1992). Testing of water from natural habitats showed a detection limit in the same range as that for the defined samples. The detection limits and the simplicity of the method (paramagnetic beads can be handled in automated systems) suggest that our method is suitable for routine environmental monitoring.
Comparison of Energy and Growth Yields for
Desulfitobacterium dehalogenans
during Utilization of Chlorophenol and Various Traditional Electron Acceptors
Authors: Mackiewicz M. , Wiegel J.
Published: December 2019

Desulfitobacterium dehalogenans
grew with formate as the electron donor and 3-chloro-4-hydroxyphenylacetate (3-Cl-4-OHPA) as the electron acceptor, yielding
Y

X
/formate

,
Y

X
/2e
−

, and
Y

X
/ATP

ranging from 3.2 to 11.3 g of biomass (dry weight)/mol, thus indicating that energy was conserved through reductive dechlorination. Pyruvate was utilized as the electron donor and acceptor, yielding stoichiometric amounts of acetate and lactate, respectively, and a
Y

X
/reduced acceptor

of 13.0 g of biomass (dry weight)/mol. The supplementation of pyruvate-containing medium with additional electron acceptors, such as 3-Cl-4-OHPA, nitrate, fumarate, or sulfite, caused pyruvate to be replaced as the electron acceptor and nearly doubled the
Y

X
/ATP

(
Y

X
/acetate formed

). A comparison of the yields for 3-Cl-4-OHPA with those for other traditional electron acceptors indicates that the dehalogenation reaction led to the formation of similar amounts of energy equivalents. The various electron acceptors were used concomitantly with 3-Cl-4-OHPA in nonacclimated cultures, but the utilization rates and amounts utilized differed.
Purification and Properties of a Polyester Polyurethane-Degrading Enzyme from
Comamonas acidovorans
TB-35
Authors: Akutsu Yukie , Nakajima-Kambe Toshiaki , Nomura Nobuhiko , Nakahara Tadaatsu
Published: December 2019

A polyester polyurethane (PUR)-degrading enzyme, PUR esterase, derived from
Comamonas acidovorans
TB-35, a bacterium that utilizes polyester PUR as the sole carbon source, was purified until it showed a single band in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). This enzyme was bound to the cell surface and was extracted by addition of 0.2%
N
,
N
-bis(3-
d
-gluconamidopropyl)deoxycholamide (deoxy-BIGCHAP). The results of gel filtration and SDS-PAGE showed that the PUR esterase was a monomer with a molecular mass of about 62,000 Da. This enzyme, which is a kind of esterase, degraded solid polyester PUR, with diethylene glycol and adipic acid released as the degradation products. The optimum pH for this enzyme was 6.5, and the optimum temperature was 45°C. PUR degradation by the PUR esterase was strongly inhibited by the addition of 0.04% deoxy-BIGCHAP. On the other hand, deoxy-BIGCHAP did not inhibit the activity when
p
-nitrophenyl acetate, a water-soluble compound, was used as a substrate. These observations indicated that this enzyme degrades PUR in a two-step reaction: hydrophobic adsorption to the PUR surface and hydrolysis of the ester bond of PUR.
Construction and Characterization of a 1,3-Propanediol Operon
Authors: Skraly Frank A. , Lytle Betsy L. , Cameron Douglas C.
Published: December 2019

The genes for the production of 1,3-propanediol (1,3-PD) in
Klebsiella pneumoniae
,
dhaB
, which encodes glycerol dehydratase, and
dhaT
, which encodes 1,3-PD oxidoreductase, are naturally under the control of two different promoters and are transcribed in different directions. These genes were reconfigured into an operon containing
dhaB
followed by
dhaT
under the control of a single promoter. The operon contains unique restriction sites to facilitate replacement of the promoter and other modifications. In a fed-batch cofermentation of glycerol and glucose,
Escherichia coli
containing the operon consumed 9.3 g of glycerol per liter and produced 6.3 g of 1,3-PD per liter. The fermentation had two distinct phases. In the first phase, significant cell growth occurred and the products were mainly 1,3-PD and acetate. In the second phase, very little growth occurred and the main products were 1,3-PD and pyruvate. The first enzyme in the 1,3-PD pathway, glycerol dehydratase, requires coenzyme B
12
, which must be provided in
E. coli
fermentations. However, the amount of coenzyme B
12
needed was quite small, with 10 nM sufficient for good 1,3-PD production in batch cofermentations. 1,3-PD is a useful intermediate in the production of polyesters. The 1,3-PD operon was designed so that it can be readily modified for expression in other prokaryotic hosts; therefore, it is useful for metabolic engineering of 1,3-PD pathways from glycerol and other substrates such as glucose.
The Cytochrome
c
Maturation Operon Is Involved in Manganese Oxidation in
Pseudomonas putida
GB-1
Authors: de Vrind J. P. M. , Brouwers G. J. , Corstjens P. L. A. M. , den Dulk J. , de Vrind-de Jong E. W.
Published: December 2019

A
Pseudomonas putida
strain, strain GB-1, oxidizes Mn
2+
to Mn oxide in the early stationary growth phase. It also secretes a siderophore (identified as pyoverdine) when it is subjected to iron limitation. After transposon (Tn
5
) mutagenesis several classes of mutants with differences in Mn
2+
oxidation and/or secretion of the Mn
2+
-oxidizing activity were identified. Preliminary analysis of the Tn
5
insertion site in one of the nonoxidizing mutants suggested that a multicopper oxidase-related enzyme is involved in Mn
2+
oxidation. The insertion site in another mutant was preliminarily identified as a gene involved in the general protein secretion pathway. Two mutants defective in Mn
2+
-oxidizing activity also secreted porphyrins into the medium and appeared to be derepressed for pyoverdine production. These strains were chosen for detailed analysis. Both mutants were shown to contain Tn
5
insertions in the
ccmF
gene, which is part of the cytochrome
c
maturation operon. They were cytochrome oxidase negative and did not contain
c
-type cytochromes. Complementation with part of the
ccm
operon isolated from the wild type restored the phenotype of the parent strain. These results indicate that a functional
ccm
operon is required for Mn
2+
oxidation in
P. putida
GB-1. A possible relationship between porphyrin secretion resulting from the
ccm
mutation and stimulation of pyoverdine production is discussed.
Application of Molecular Biological Techniques to a Seasonal Study of Ammonia Oxidation in a Eutrophic Freshwater Lake
Authors: Hastings Richard C. , Saunders Jon R. , Hall Grahame H. , Pickup Roger W. , McCarthy Alan J.
Published: December 2019

The autotrophic ammonia-oxidizing bacteria in a eutrophic freshwater lake were studied over a 12-month period. Numbers of ammonia oxidisers in the lakewater were small throughout the year, and tangential-flow concentration was required to obtain meaningful estimates of most probable numbers. Sediments from littoral and profundal sites supported comparatively large populations of these bacteria, and the nitrification potential was high, particularly in summer samples from the littoral sediment surface. In enrichment cultures, lakewater samples nitrified at low (0.67 mM) ammonium concentrations only whereas sediment samples exhibited nitrification at high (12.5 mM) ammonium concentrations also. Enrichments at low ammonium concentration did not nitrify when inoculated into high-ammonium medium, but the converse was not true. This suggests that the water column contains a population of ammonia oxidizers that is sensitive to high ammonium concentrations. The observation of nitrification at high ammonium concentration by isolates from some winter lakewater samples, identified as nitrosospiras by 16S rRNA probing, is consistent with the hypothesis that sediment ammonia oxidizers enter the water column at overturn. With only one exception, nested PCR amplification enabled the detection of
Nitrosospira
16S rDNA in all samples, but
Nitrosomonas
(
N. europaea-eutropha
lineage) 16S rDNA was never obtained. However, the latter were part of the sediment and water column communities, because their 16S rRNA could be detected by specific oligonucleotide probing of enrichment cultures. Furthermore, a specific PCR amplification regime for the
Nitrosomonas europaea
ammonia monooxygenase gene (
amoA
) yielded positive results when applied directly to sediment and lakewater samples. Patterns of
Nitrosospira
and
Nitrosomonas
detection by 16S rRNA oligonucleotide probing of sediment enrichment cultures were complex, but lakewater enrichments at low ammonium concentration were positive for nitrosomonads and not nitrosospiras. Analysis of enrichment cultures has therefore provided evidence for the existence of subpopulations within the lake ammonia-oxidizing community distinguishable on the basis of ammonium tolerance and possibly showing a seasonal distribution between the sediment and water column.
Enhanced Utilization of Phosphonate and Phosphite by
Klebsiella aerogenes
Authors: Imazu Kazuya , Tanaka Shotaro , Kuroda Akio , Anbe Yuki , Kato Junichi , Ohtake Hisao
Published: December 2019

Klebsiella aerogenes
ATCC 9621 was able to utilize phosphonates (P
n
), including aminoethylphosphonate, ethylphosphonate, methylphosphonate (MP
n
), and phosphonoacetate, and inorganic phosphite (P
t
) as sole sources of phosphorus (P). The products of the
phn
gene cluster were absolutely required for P
n
breakdown and P
t
oxidation to inorganic phosphate (P
i
) in this organism. To determine if
K. aerogenes
ATCC 9621 could be engineered to enhance the utilization of P
n
and P
t
, a multicopy plasmid, pBI05, which carried the entire
phn
gene cluster, was introduced into this strain. Despite the increased dosage of the
phn
genes,
K. aerogenes
ATCC 9621(pBI05) could utilize only up to 1.1-fold more P
n
and P
t
than did the control strain with the parent vector alone. These results suggested that P
i
, which was generated from P
n
and P
t
, might limit further utilization of these P compounds. Consequently, to convert the resulting P
i
to polyphosphate (polyP), the plasmid pKP28, which carried the
K. aerogenes ppk
gene (which encodes polyP kinase), was introduced into
K. aerogenes
ATCC 9621(pBI05). Overexpression of the
ppk
gene in
K. aerogenes
ATCC 9621(pBI05, pKP28) resulted in a 2.5-fold increase in P
t
utilization over that of the control strain. This recombinant strain also accumulated approximately sixfold more P than did the control strain when the cells were grown with MP
n
as a sole source of P.
Xanthan Lyase of
Bacillus
sp. Strain GL1 Liberates Pyruvylated Mannose from Xanthan Side Chains
Authors: Hashimoto Wataru , Miki Hikaru , Tsuchiya Noriaki , Nankai Hirokazu , Murata Kousaku
Published: December 2019

When the bacterium
Bacillus
sp. strain GL1 was grown in a medium containing xanthan as the carbon source, the viscosity of the medium decreased in association with growth, showing that the bacterium had xanthan-depolymerizing enzymes. One of the xanthan-depolymerizing enzymes (xanthan lyase) was present in the medium and was found to be induced by xanthan. The xanthan lyase purified from the culture fluid was a monomer with a molecular mass of 75 kDa, and was most active at pH 5.5 and 50°C. The enzyme was highly specific for xanthan and produced pyruvylated mannose. The result indicates that the enzyme cleaved the linkage between the terminal pyruvylated mannosyl and glucuronyl residues in the side chain of xanthan.
Genetic Characterization of
Pseudomonas syringae
pv. syringae Strains from Stone Fruits in California
Authors: Little E. L. , Bostock R. M. , Kirkpatrick B. C.
Published: December 2019

Strains of
Pseudomonas syringae
pv. syringae were isolated from healthy and diseased stone fruit tissues sampled from 43 orchard sites in California in 1995 and 1996. These strains, together with
P. syringae
strains from other hosts and pathovars, were tested for pathogenicity and the presence of the
syrB
and
syrC
genes and were genetically characterized by using enterobacterial repetitive intergenic consensus (ERIC) primers and PCR. All 89 strains of
P. syringae
pv. syringae tested were moderately to highly pathogenic on Lovell peach seedlings regardless of the host of origin, while strains of other pathovars exhibited low or no pathogenicity. The 19 strains of
P. syringae
pv. syringae examined by restriction fragment length polymorphism analysis contained the
syrB
and
syrC
genes, whereas no hybridization occurred with 4 strains of other
P. syringae
pathovars. The
P. syringae
pv. syringae strains from stone fruit, except for a strain from New Zealand, generated ERIC genomic fingerprints which shared four fragments of similar mobility. Of the
P. syringae
pv. syringae strains tested from other hosts, only strains from rose, kiwi, and pear generated genomic fingerprints that had the same four fragments as the stone fruit strains. Analysis of the ERIC fingerprints from
P. syringae
pv. syringae strains showed that the strains isolated from stone fruits formed a distinct cluster separate from most of the strains isolated from other hosts. These results provide evidence of host specialization within the diverse pathovar
P. syringae
pv. syringae.
Characterization of Genes Involved in Biosynthesis of a Novel Antibiotic from
Burkholderia cepacia
BC11 and Their Role in Biological Control of
Rhizoctonia solani
Authors: Kang Yaowei , Carlson Russell , Tharpe Wendy , Schell Mark A.
Published: December 2019

Genetic manipulation of fluorescent pseudomonads has provided major insight into their production of antifungal molecules and their role in biological control of plant disease.
Burkholderia cepacia
also produces antifungal activities, but its biological control activity is much less well characterized, in part due to difficulties in applying genetic tools. Here we report genetic and biochemical characterization of a soil isolate of
B. cepacia
relating to its production of an unusual antibiotic that is very active against a variety of soil fungi. Purification and preliminary structural analyses suggest that this antibiotic (called AFC-BC11) is a novel lipopeptide associated largely with the cell membrane. Analysis of conditions for optimal production of AFC-BC11 indicated stringent environmental regulation of its synthesis. Furthermore, we show that production of AFC-BC11 is largely responsible for the ability of
B. cepacia
BC11 to effectively control the damping-off of cotton caused by the fungal pathogen
Rhizoctonia solani
in a gnotobiotic system. Using Tn
5
mutagenesis, we identified, cloned, and characterized a region of the genome of strain BC11 that is required for production of this antifungal metabolite. DNA sequence analysis suggested that this region encodes proteins directly involved in the production of a nonribosomally synthesized lipopeptide.
Isolation and Identification of
Helicobacter
spp. from Canine and Feline Gastric Mucosa
Authors: Jalava Katri , On Stephen L. W. , Vandamme Peter A. R. , Happonen Irmeli , Sukura Antti , Hänninen Marja-Liisa
Published: December 2019

It is known that virtually all healthy adult dogs and cats harbor spiral helicobacters in their gastric mucosa. Three species,
Helicobacter felis
,
Helicobacter bizzozeronii
, and
Helicobacter salomonis
have been isolated in vitro from the gastric mucosa of these animals. The aims of this study were to evaluate the efficacy of an isolation method for canine and feline gastric helicobacters that has been developed at the University of Helsinki; to estimate the prevalence and distribution of these taxa in the samples examined; and to assess the efficacy and validity of an extensive set of standardized conventional phenotypic tests, whole-cell protein profiling, and ultrastructural analysis in identifying the different species isolated from canine and feline gastric mucosa. We cultured 95 and 22 gastric mucosal biopsies from dogs and cats, respectively. Twenty-one
H. bizzozeronii
strains, 8
H. felis
strains, 8
H. salomonis
strains, 3 mixed cultures, 2 “
Flexispira rappini
”-like organisms, and 3 as yet uncharacterized strains were isolated from the dogs, and 3
H. felis
strains were isolated from the cats. The methods used here yielded
Helicobacter
isolation rates of 51% from dogs and 13.6% from cats, which exceed those reported previously. The main difficulties were primary isolation, mixed cultures, and identification to the species level. In the species identification, a detailed morphological examination was found to yield important phenotypic characteristics. A large panel of biochemical and tolerance tests did not clearly differentiate the closely related species
H. bizzozeronii
,
H. felis
, and
H. salomonis
. Highly standardized whole-cell protein profiling was shown to be an excellent method for species identification. Improvements in culture conditions for these bacteria are still needed, especially for cats. A genetic identification method not requiring culture is needed for future studies of these very fastidious helicobacters, as the clinical significance and ecology of these species within the gastric mucosa of the domestic carnivores remain largely unknown.
Resistance to Tellurite as a Selection Marker for Genetic Manipulations of
Pseudomonas
Strains
Authors: Sanchez-Romero Juan M. , Diaz-Orejas Ramon , De Lorenzo Victor
Published: December 2019

Resistance to the toxic compound potassium tellurite (Tel
r
) has been employed as a selection marker built into a set of transposon vectors and broad-host-range plasmids tailored for genetic manipulations of
Pseudomonas
strains potentially destined for environmental release. In this study, the activated Tel
r
determinants encoded by the cryptic
telAB
genes of plasmid RK2 were produced, along with the associated
kilA
gene, as DNA cassettes compatible with cognate vectors. In one case, the Tel
r
determinants were assembled between the I and O ends of a suicide delivery vector for mini-Tn
5
transposons. In another case, the
kilA
and
telAB
genes were combined with a minimal replicon derived from a variant of
Pseudomonas
plasmid pPS10, which is able to replicate in a variety of gram-negative hosts and is endowed with a modular collection of cloning and expression assets. Either in the plasmid or in the transposon vector, the Tel
r
marker was combined with a 12-kb DNA segment of plasmid pWW0 of
Pseudomonas putida
mt-2 encoding the upper TOL pathway enzymes. This allowed construction of antibiotic resistance-free but selectable
P. putida
strains with the ability to grow on toluene as the sole carbon source through an
ortho
-cleavage catabolic pathway.
Synthesis of Glycine Betaine from Exogenous Choline in the Moderately Halophilic Bacterium
Halomonas elongata
Authors: Cánovas David , Vargas Carmen , Csonka Laszlo N. , Ventosa Antonio , Nieto Joaquín J.
Published: December 2019

The role of choline in osmoprotection in the moderate halophile
Halomonas elongata
has been examined. Transport and conversion of choline to betaine began immediately after addition of choline to the growth medium. Intracellular accumulation of betaine synthesized from choline was salt dependent up to 2.5 M NaCl. Oxidation of choline was enhanced at 2.0 M NaCl in the presence or absence of externally provided betaine. This indicates that the NaCl concentration in the growth medium has major effects on the choline-betaine pathway of
H. elongata
.
Automated Confocal Laser Scanning Microscopy and Semiautomated Image Processing for Analysis of Biofilms
Authors: Kuehn Martin , Hausner Martina , Bungartz Hans-Joachim , Wagner Michael , Wilderer Peter A. , Wuertz Stefan
Published: December 2019

The purpose of this study was to develop and apply a quantitative optical method suitable for routine measurements of biofilm structures under in situ conditions. A computer program was designed to perform automated investigations of biofilms by using image acquisition and image analysis techniques. To obtain a representative profile of a growing biofilm, a nondestructive procedure was created to study and quantify undisturbed microbial populations within the physical environment of a glass flow cell. Key components of the computer-controlled processing described in this paper are the on-line collection of confocal two-dimensional (2D) cross-sectional images from a preset 3D domain of interest followed by the off-line analysis of these 2D images. With the quantitative extraction of information contained in each image, a three-dimensional reconstruction of the principal biological events can be achieved. The program is convenient to handle and was generated to determine biovolumes and thus facilitate the examination of dynamic processes within biofilms. In the present study,
Pseudomonas fluorescens
or a green fluorescent protein-expressing
Escherichia coli
strain, EC12, was inoculated into glass flow cells and the respective monoculture biofilms were analyzed in three dimensions. In this paper we describe a method for the routine measurements of biofilms by using automated image acquisition and semiautomated image analysis.
Analysis of the Bacteriolytic Enzymes of the Autolytic
Lactococcus lactis
subsp.
cremoris
Strain AM2 by Renaturing Polyacrylamide Gel Electrophoresis: Identification of a Prophage-Encoded Enzyme
Authors: Lepeuple Anne-Sophie , Van Gemert Ellis , Chapot-Chartier Marie-Pierre
Published: December 2019

Lactococcus lactis
subsp.
cremoris
AM2 was previously shown to lyse early and extensively during cheese ripening (M.-P. Chapot-Chartier, C. Deniel, M. Rousseau, L. Vassal, and J.-C. Gripon, Int. Dairy J. 4:251–269, 1994). We analyzed the bacteriolytic activities of autolytic strain AM2 by using renaturing sodium dodecyl sulfate-polyacrylamide gel electrophoresis performed with two different substrates in the gel,
Micrococcus lysodeikticus
and
L. lactis
autoclaved cells. Several lytic activities were detected in
L. lactis
AM2; a major lytic activity, designated A2 (46 kDa), was found only with the
L. lactis
cell substrate. This activity appears to be different from major peptidoglycan hydrolase AcmA characterized previously (G. Buist, J. Kok, K. J. Leenhouts, M. Dabrowska, G. Venema, and A. J. Haandrickman, J. Bacteriol. 177:1554–1563, 1995), which has a similar molecular mass. The two enzymes differ in substrate specificity as well as in sensitivity to pH and different chemical compounds.
L. lactis
AM2 is lysogenic and mitomycin C inducible. Enzyme A2 was shown to be inducible by mitomycin C and to be prophage encoded. It was identified as an enzyme similar to the lysin encoded by lactococcal small isometric temperate bacteriophages. A prophage-cured derivative of
L. lactis
AM2 was obtained, and this isolate exhibited different autolytic properties than AM2. After prolonged incubation in the stationary phase after growth on M17 medium, the extent of lysis of an AM2 culture was 60%, whereas over the same period there was almost no lysis in a prophage-cured derivative strain culture. These results suggest that the prophage lytic system is involved in the strain AM2 lysis observed in liquid medium and that it could also be involved in the lysis observed during cheese ripening.
Ethylene Removal by a Biofilter with Immobilized Bacteria
Authors: Elsgaard Lars
Published: December 2019

A biofilter which eliminated ethylene (C2H4) from the high parts-per-million range to levels near the limit for plant hormonal activity (0.01 to 0.1 ppm) was developed. Isolated ethylene-oxidizing bacteria were immobilized on peat-soil in a biofilter (687 cm3) and subjected to an atmospheric gas flow (73.3 ml min−1) with 2 or 117 ppm of C2H4. Ethylene was eliminated to a minimum level of 0.017 ppm after operation with 2.05 ppm of C2H4for 16 days. Also, the inlet C2H4concentration of 117 ppm was reduced to <0.04 ppm. During operation with 2 and 117 ppm of C2H4, an increase in the C2H4removal rate was observed, which was attributed to proliferation of the immobilized bacteria, notably in the first 0- to 5-cm segment of the biofilter. The maximal C2H4elimination capacity of the biofilter was 21 g of C2H4m−3day−1during operation with 117 ppm of C2H4in the inlet gas. However, for the first 0- to 5-cm segment of the biofilter, an elimination capacity of 146 g of C2H4m−3day−1was calculated. Transition of the biofilter temperature from 21 to 10°C caused a 1.6-fold reduction in the C2H4removal rate, which was reversed during operation for 18 days. Batch experiments with inoculated peat-soil demonstrated that C2H4removal still occurred after storage at 2, 8, and 20°C for 2, 3, and 4 weeks. However, the C2H4removal rate decreased with increasing storage time and was reduced by ca. 50% after storage for 2 weeks at all three temperatures. The biofilter could be a suitable tool for C2H4removal in, e.g., horticultural storage facilities, since it (i) removed C2H4to 0.017 ppm, (ii) had a good operational stability, and (iii) operated efficiently at 10°C.