bluuebunny/crossref_metadata_2025_split huggingface dataset:
Living in an Extremely Polluted Environment: Clues from the Genome of Melanin-Producing Aeromonas salmonicida subsp. pectinolytica 34mel
T
Authors: Pavan María Elisa , Pavan Esteban E. , López Nancy I. , Levin Laura , Pettinari M. Julia 
Published: May 2015

ABSTRACT

Aeromonas salmonicida
subsp.
pectinolytica
34mel
T
can be considered an extremophile due to the characteristics of the heavily polluted river from which it was isolated. While four subspecies of
A. salmonicida
are known fish pathogens, 34mel
T
belongs to the only subspecies isolated solely from the environment. Genome analysis revealed a high metabolic versatility, the capability to cope with diverse stress agents, and the lack of several virulence factors found in pathogenic
Aeromonas
. The most relevant phenotypic characteristics of 34mel
T
are pectin degradation, a distinctive trait of
A. salmonicida
subsp.
pectinolytica
, and melanin production. Genes coding for three pectate lyases were detected in a cluster, unique to this microorganism, that contains all genes needed for pectin degradation. Melanin synthesis in 34mel
T
is hypothesized to occur through the homogentisate pathway, as no tyrosinases or laccases were detected and the homogentisate 1,2-dioxygenase gene is inactivated by a transposon insertion, leading to the accumulation of the melanin precursor homogentisate. Comparative genome analysis of other melanogenic
Aeromonas
strains revealed that this gene was inactivated by transposon insertions or point mutations, indicating that melanin biosynthesis in
Aeromonas
occurs through the homogentisate pathway. Horizontal gene transfer could have contributed to the adaptation of 34mel
T
to a highly polluted environment, as 13 genomic islands were identified in its genome, some of them containing genes coding for fitness-related traits. Heavy metal resistance genes were also found, along with others associated with oxidative and nitrosative stresses. These characteristics, together with melanin production and the ability to use different substrates, may explain the ability of this microorganism to live in an extremely polluted environment.
Comparison of Quantitative PCR and Droplet Digital PCR Multiplex Assays for Two Genera of Bloom-Forming Cyanobacteria, Cylindrospermopsis and Microcystis
Authors: Te Shu Harn , Chen Enid Yingru , Gin Karina Yew-Hoong 
Published: May 2015

ABSTRACT

The increasing occurrence of harmful cyanobacterial blooms, often linked to deteriorated water quality and adverse public health effects, has become a worldwide concern in recent decades. The use of molecular techniques such as real-time quantitative PCR (qPCR) has become increasingly popular in the detection and monitoring of harmful cyanobacterial species. Multiplex qPCR assays that quantify several toxigenic cyanobacterial species have been established previously; however, there is no molecular assay that detects several bloom-forming species simultaneously.
Microcystis
and
Cylindrospermopsis
are the two most commonly found genera and are known to be able to produce microcystin and cylindrospermopsin hepatotoxins. In this study, we designed primers and probes which enable quantification of these genera based on the RNA polymerase C1 gene for
Cylindrospermopsis
species and the c-phycocyanin beta subunit-like gene for
Microcystis
species. Duplex assays were developed for two molecular techniques—qPCR and droplet digital PCR (ddPCR). After optimization, both qPCR and ddPCR assays have high linearity and quantitative correlations for standards. Comparisons of the two techniques showed that qPCR has higher sensitivity, a wider linear dynamic range, and shorter analysis time and that it was more cost-effective, making it a suitable method for initial screening. However, the ddPCR approach has lower variability and was able to handle the PCR inhibition and competitive effects found in duplex assays, thus providing more precise and accurate analysis for bloom samples.
Gene Replacement for the Generation of Designed Novel Avermectin Derivatives with Enhanced Acaricidal and Nematicidal Activities
Authors: Huang Jun , Chen An-Liang , Zhang Hui , Yu Zhen , Li Mei-Hong , Li Na , Lin Jia-Tan , Bai Hua , Wang Ji-Dong , Zheng Yu-Guo 
Published: May 2015

ABSTRACT

Avermectin (AVM) and ivermectin (IVM) are potent pesticides and acaricides which have been widely used during the past 30 years. As insect resistance to AVM and IVM is greatly increasing, alternatives are urgently needed. Here, we report two novel AVM derivatives, tenvermectin A (TVM A) and TVM B, which are considered a potential new generation of agricultural and veterinary drugs. The molecules of the TVMs were designed based on structure and pharmacological property comparisons among AVM, IVM, and milbemycin (MBM). To produce TVMs, a genetically engineered strain, MHJ1011, was constructed from
Streptomyces avermitilis
G8-17, an AVM industrial strain. In MHJ1011, the native
aveA1
gene was seamlessly replaced with
milA1
from
Streptomyces hygroscopicus
. The total titer of the two TVMs produced by MHJ1011 reached 3,400 mg/liter. Insecticidal tests proved that TVM had enhanced activities against
Tetranychus cinnabarinus
and
Bursaphelenchus xylophilus
, as desired. This study provides a typical example of exploration for novel active compounds through a new method of polyketide synthase (PKS) reassembly for gene replacement. The results of the insecticidal tests may be of use in elucidating the structure-activity relationship of AVMs and MBMs.
Dual-Color Bioluminescence Imaging for Simultaneous Monitoring of the Intestinal Persistence of Lactobacillus plantarum and Lactococcus lactis in Living Mice
Authors: Daniel Catherine , Poiret Sabine , Dennin Véronique , Boutillier Denise , Lacorre Delphine Armelle , Foligné Benoit , Pot Bruno 
Published: May 2015

ABSTRACT

Lactic acid bacteria are found in the gastrointestinal tract of mammals and have received tremendous attention due to their health-promoting properties. We report the development of two dual-color luciferase-producing
Lactobacillus
(
Lb
.)
plantarum
and
Lactococcus
(
Lc
.)
lactis
strains for noninvasive simultaneous tracking in the mouse gastrointestinal tract. We previously described the functional expression of the red luciferase mutant (CBRluc) from
Pyrophorus plagiophthalamus
in
Lb. plantarum
NCIMB8826 and
Lc. lactis
MG1363 (C. Daniel, S. Poiret, V. Dennin, D. Boutillier, and B. Pot, Appl Environ Microbiol
79:
1086–1094, 2013,
http://dx.doi.org/10.1128/AEM.03221-12
). In this study, we determined that CBRluc is a better-performing luciferase for
in vivo
localization of both lactic acid bacteria after oral administration than the green click beetle luciferase mutant construct developed in this study. We further established the possibility to simultaneously detect red- and green-emitting lactic acid bacteria by dual-wavelength bioluminescence imaging in combination with spectral unmixing. The difference in spectra of light emission by the red and green click beetle luciferase mutants and dual bioluminescence detection allowed
in vitro
and
in vivo
quantification of the red and green emitted signals; thus, it allowed us to monitor the dynamics and fate of the two bacterial populations simultaneously. Persistence and viability of both strains simultaneously administered to mice in different ratios was studied
in vivo
in anesthetized mice and
ex vivo
in mouse feces. The application of dual-luciferase-labeled bacteria has considerable potential to simultaneously study the interactions and potential competitions of different targeted bacteria and their hosts.
Tulane Virus as a Potential Surrogate To Mimic Norovirus Behavior in Oysters
Authors: Drouaz Najoua , Schaeffer Julien , Farkas Tibor , Le Pendu Jacques , Le Guyader Françoise S. 
Published: May 2015

ABSTRACT
Oyster contamination by noroviruses is an important health and economic problem. The present study aimed to compare the behaviors of Norwalk virus (the prototype genogroup I norovirus) and two culturable viruses: Tulane virus and mengovirus. After bioaccumulation, tissue distributions were quite similar for Norwalk virus and Tulane virus, with the majority of viral particles detected in digestive tissues, while mengovirus was detected in large amounts in the gills and mantle as well as in digestive tissues. The levels of persistence of all three viruses over 8 days were comparable, but clear differences were observed over longer periods, with Norwalk and Tulane viruses displaying rather similar half-lives, unlike mengovirus, which was cleared more rapidly. These results indicate that Tulane virus may be a good surrogate for studying norovirus behavior in oysters, and they confirm the prolonged persistence of Norwalk virus in oyster tissues.
Genes Associated with Desiccation and Osmotic Stress in Listeria monocytogenes as Revealed by Insertional Mutagenesis
Authors: Hingston Patricia A. , Piercey Marta J. , Truelstrup Hansen Lisbeth 
Published: May 2015

ABSTRACT

Listeria monocytogenes
is a foodborne pathogen whose survival in food processing environments may be associated with its tolerance to desiccation. To probe the molecular mechanisms used by this bacterium to adapt to desiccation stress, a transposon library of 11,700
L. monocytogenes
mutants was screened, using a microplate assay, for strains displaying increased or decreased desiccation survival (43% relative humidity, 15°C) in tryptic soy broth (TSB). The desiccation phenotypes of selected mutants were subsequently assessed on food-grade stainless steel (SS) coupons in TSB plus 1% glucose (TSB-glu). Single transposon insertions in mutants exhibiting a change in desiccation survival of >0.5 log CFU/cm
2
relative to that of the wild type were determined by sequencing arbitrary PCR products. Strain morphology, motility, and osmotic stress survival (in TSB-glu plus 20% NaCl) were also analyzed. The initial screen selected 129 desiccation-sensitive (DS) and 61 desiccation-tolerant (DT) mutants, out of which secondary screening on SS confirmed 15 DT and 15 DS mutants. Among the DT mutants, seven immotile and flagellum-less strains contained transposons in genes involved in flagellum biosynthesis (
fliP
,
flhB
,
flgD
,
flgL
) and motor control (
motB
,
fliM
,
fliY
), while others harbored transposons in genes involved in membrane lipid biosynthesis, energy production, potassium uptake, and virulence. The genes that were interrupted in the 15 DS mutants included those involved in energy production, membrane transport, protein metabolism, lipid biosynthesis, oxidative damage control, and putative virulence. Five DT and 14 DS mutants also demonstrated similar significantly (
P
< 0.05) different survival relative to that of the wild type when exposed to osmotic stress, demonstrating that some genes likely have similar roles in allowing the organism to survive the two water stresses.
Dynamics of the Streptococcus gordonii Transcriptome in Response to Medium, Salivary α-Amylase, and Starch
Authors: Haase Elaine M. , Feng Xianghui , Pan Jiachuan , Miecznikowski Jeffrey C. , Scannapieco Frank A. 
Published: May 2015

ABSTRACT

Streptococcus gordonii
, a primary colonizer of the tooth surface, interacts with salivary α-amylase via amylase-binding protein A (AbpA). This enzyme hydrolyzes starch to glucose, maltose, and maltodextrins that can be utilized by various oral bacteria for nutrition. Microarray studies demonstrated that AbpA modulates gene expression in response to amylase, suggesting that the amylase-streptococcal interaction may function in ways other than nutrition. The goal of this study was to explore the role of AbpA in gene regulation through comparative transcriptional profiling of wild-type KS1 and AbpA
−
mutant KS1Ω
abpA
under various environmental conditions. A portion of the total RNA isolated from mid-log-phase cells grown in 5% CO
2
in (i) complex medium with or without amylase, (ii) defined medium (DM) containing 0.8% glucose with/without amylase, and (iii) DM containing 0.2% glucose and amylase with or without starch was reverse transcribed to cDNA and the rest used for RNA sequencing. Changes in the expression of selected genes were validated by quantitative reverse transcription-PCR. Maltodextrin-associated genes, fatty acid synthesis genes and competence genes were differentially expressed in a medium-dependent manner. Genes in another cluster containing a putative histidine kinase/response regulator, peptide methionine sulfoxide reductase, thioredoxin protein, lipoprotein, and cytochrome
c
-type protein were downregulated in KS1Ω
abpA
under all of the environmental conditions tested. Thus, AbpA appears to modulate genes associated with maltodextrin utilization/transport and fatty acid synthesis. Importantly, in all growth conditions AbpA was associated with increased expression of a potential two-component signaling system associated with genes involved in reducing oxidative stress, suggesting a role in signal transduction and stress tolerance.
Inactivation of Escherichia coli Cells in Aqueous Solution by Atmospheric-Pressure N
2
, He, Air, and O
2
Microplasmas
Authors: Zhou Renwu , Zhang Xianhui , Bi Zhenhua , Zong Zichao , Niu Jinhai , Song Ying , Liu Dongping , Yang Size 
Published: May 2015

ABSTRACT

Atmospheric-pressure N
2
, He, air, and O
2
microplasma arrays have been used to inactivate
Escherichia coli
cells suspended in aqueous solution. Measurements show that the efficiency of inactivation of
E. coli
cells is strongly dependent on the feed gases used, the plasma treatment time, and the discharge power. Compared to atmospheric-pressure N
2
and He microplasma arrays, air and O
2
microplasma arrays may be utilized to more efficiently kill
E. coli
cells in aqueous solution. The efficiencies of inactivation of
E. coli
cells in water can be well described by using the chemical reaction rate model, where reactive oxygen species play a crucial role in the inactivation process. Analysis indicates that plasma-generated reactive species can react with
E. coli
cells in water by direct or indirect interactions.
Total Biosynthesis and Diverse Applications of the Nonribosomal Peptide-Polyketide Siderophore Yersiniabactin
Authors: Ahmadi Mahmoud Kamal , Fawaz Samar , Jones Charles H. , Zhang Guojian , Pfeifer Blaine A. 
Published: May 2015

ABSTRACTYersiniabactin (Ybt) is a mixed nonribosomal peptide-polyketide natural product natively produced by the pathogenYersinia pestis. The compound enables iron scavenging capabilities upon host infection and is biosynthesized by a nonribosomal peptide synthetase featuring a polyketide synthase module. This pathway has been engineered for expression and biosynthesis usingEscherichia colias a heterologous host. In the current work, the biosynthetic process for Ybt formation was improved through the incorporation of a dedicated step to eliminate the need for exogenous salicylate provision. When this improvement was made, the compound was tested in parallel applications that highlight the metal-chelating nature of the compound. In the first application, Ybt was assessed as a rust remover, demonstrating a capacity of ∼40% compared to a commercial removal agent and ∼20% relative to total removal capacity. The second application tested Ybt in removing copper from a variety of nonbiological and biological solution mixtures. Success across a variety of media indicates potential utility in diverse scenarios that include environmental and biomedical settings.
Candida albicans Kinesin Kar3 Depends on a Cik1-Like Regulatory Partner Protein for Its Roles in Mating, Cell Morphogenesis, and Bipolar Spindle Formation
Authors: Frazer Corey , Joshi Monika , Delorme Caroline , Davis Darlene , Bennett Richard J. , Allingham John S. 
Published: May 2015

ABSTRACT

Candida albicans
is a major fungal pathogen whose virulence is associated with its ability to transition from a budding yeast form to invasive hyphal filaments. The kinesin-14 family member
Ca
Kar3 is required for transition between these morphological states, as well as for mitotic progression and karyogamy. While kinesin-14 proteins are ubiquitous,
Ca
Kar3 homologs in hemiascomycete fungi are unique because they form heterodimers with noncatalytic kinesin-like proteins. Thus,
Ca
Kar3-based motors may represent a novel antifungal drug target. We have identified and examined the roles of a kinesin-like regulator of
Ca
Kar3. We show that
orf19.306
(dubbed
CaCIK1
) encodes a protein that forms a heterodimer with
Ca
Kar3, localizes
Ca
Kar3 to spindle pole bodies, and can bind microtubules and influence
Ca
Kar3 mechanochemistry despite lacking an ATPase activity of its own. Similar to
Ca
Kar3 depletion, loss of
Ca
Cik1 results in cell cycle arrest, filamentation defects, and an inability to undergo karyogamy. Furthermore, an examination of the spindle structure in cells lacking either of these proteins shows that a large proportion have a monopolar spindle or two dissociated half-spindles, a phenotype unique to the
C. albicans
kinesin-14 homolog. These findings provide new insights into mitotic spindle structure and kinesin motor function in
C. albicans
and identify a potentially vulnerable target for antifungal drug development.
Lipid Biosynthetic Genes Affect Candida albicans Extracellular Vesicle Morphology, Cargo, and Immunostimulatory Properties
Authors: Wolf Julie M. , Espadas Javier , Luque-Garcia Jose , Reynolds Todd , Casadevall Arturo 
Published: May 2015

ABSTRACT

Microbial secretion is integral for regulating cell homeostasis as well as releasing virulence factors during infection. The genes encoding phosphatidylserine synthase (
CHO1
) and phosphatidylserine decarboxylase (
PSD1
and
PSD2
) are
Candida albicans
genes involved in phospholipid biosynthesis, and mutations in these genes affect mitochondrial function, cell wall thickness, and virulence in mice. We tested the roles of these genes in several agar-based secretion assays and observed that the
cho1
Δ/Δ and
psd1
Δ/Δ
psd2
Δ/Δ strains manifested less protease and phospholipase activity. Since extracellular vesicles (EVs) are surrounded by a lipid membrane, we investigated the effects of these mutations on EV structure, composition, and biological activity. The
cho1
Δ/Δ mutant releases EVs comparable in size to wild-type EVs, but EVs from the
psd1
Δ/Δ
psd2
Δ/Δ strain are much larger than those from the wild type, including a population of >100-nm EVs not observed in the EVs from the wild type. Proteomic analysis revealed that EVs from both mutants had a significantly different protein cargo than that of EVs from the wild type. EVs were tested for their ability to activate NF-κB in bone marrow-derived macrophage cells. While wild-type and
psd1
Δ/Δ
psd2
Δ/Δ mutant-derived EVs activated NF-κB, the
cho1
Δ/Δ mutant-derived EV did not. These studies indicate that the presence and absence of these
C. albicans
genes have qualitative and quantitative effects on EV size, composition, and immunostimulatory phenotypes that highlight a complex interplay between lipid metabolism and vesicle production.
Rgc2 Regulator of Glycerol Channel Fps1 Functions as a Homo- and Heterodimer with Rgc1
Authors: Lee Jongmin , Levin David E. 
Published: May 2015

ABSTRACT
The plasma membrane aquaglyceroporin Fps1 is responsible for glycerol transport in yeast in response to changes in extracellular osmolarity. Fps1 functions as a homotetramer, and control of its channel activity in response to hyperosmotic shock involves a redundant pair of fungus-specific regulators, Rgc1 and Rgc2 (regulators of the glycerol channel), and the mitogen-activatd protein kinase (MAPK) Hog1 (high-osmolarity glycerol response). Rgc1 and Rgc2 maintain Fps1 in an open-channel state by binding to its C-terminal cytoplasmic domain. Phosphorylation of Rgc1 and Rgc2 by Hog1 induces their eviction from Fps1 and consequent channel closure. In the absence of Fps1 channel function, cells experience chronic cell wall stress, which may be exploited for antifungal drug development. We show here that Rgc1 and Rgc2 form homodimers and heterodimers with each other and that dimer formation of Rgc2 is mediated by its N-terminal domain. Mutations that prevent Rgc2 dimerization block its ability to open Fps1. Therefore, the Rgc-Rgc dimer interface might be an attractive drug target.
Evolution of Chemical Diversity in Echinocandin Lipopeptide Antifungal Metabolites
Authors: Yue Qun , Chen Li , Zhang Xiaoling , Li Kuan , Sun Jingzu , Liu Xingzhong , An Zhiqiang , Bills Gerald F. 
Published: May 2015

ABSTRACT

The echinocandins are a class of antifungal drugs that includes caspofungin, micafungin, and anidulafungin. Gene clusters encoding most of the structural complexity of the echinocandins provided a framework for hypotheses about the evolutionary history and chemical logic of echinocandin biosynthesis. Gene orthologs among echinocandin-producing fungi were identified. Pathway genes, including the nonribosomal peptide synthetases (NRPSs), were analyzed phylogenetically to address the hypothesis that these pathways represent descent from a common ancestor. The clusters share cooperative gene contents and linkages among the different strains. Individual pathway genes analyzed in the context of similar genes formed unique echinocandin-exclusive phylogenetic lineages. The echinocandin NRPSs, along with the NRPS from the
inp
gene cluster in
Aspergillus nidulans
and its orthologs, comprise a novel lineage among fungal NRPSs. NRPS adenylation domains from different species exhibited a one-to-one correspondence between modules and amino acid specificity that is consistent with models of tandem duplication and subfunctionalization. Pathway gene trees and Ascomycota phylogenies are congruent and consistent with the hypothesis that the echinocandin gene clusters have a common origin. The disjunct Eurotiomycete-Leotiomycete distribution appears to be consistent with a scenario of vertical descent accompanied by incomplete lineage sorting and loss of the clusters from most lineages of the
Ascomycota
. We present evidence for a single evolutionary origin of the echinocandin family of gene clusters and a progression of structural diversification in two fungal classes that diverged approximately 290 to 390 million years ago. Lineage-specific gene cluster evolution driven by selection of new chemotypes contributed to diversification of the molecular functionalities.
An Eco-friendly, Scaled-down Gram Stain Protocol
Authors: Gyure Ruth A. 
Published: May 2015

Currently, flushing large volumes of Gram stain reagents into sanitary sewage systems is no longer acceptable. These chemical wastes are highly regulated and must be collected, labeled, and disposed of in a responsible manner, usually by paying a commercial service to remove them to an authorized off-site facility. Such services are costly and, as expected, costs are proportional to volume of collected waste. This “old” method of Gram staining, even if effluent is collected, generates a high volume of liquid waste which is unnecessarily diluted with additional large volumes of water from the rinsing steps.
The purpose of using this scaled-down and eco-friendly protocol is to dramatically reduce the amount of liquid waste produced without sacrificing quality of results. This protocol is flexible, practical, and easy to implement. It does not require students to work at a bench sink, reduces user cost, and lowers environmental impact overall.