The pUltra plasmid series: a robust and flexible tool for fluorescent labeling of Enterobacteria

Fluorescent labeling has been an invaluable tool for the study of living organisms and bacterial species are no exception to this. Here we present and characterize the pUltra plasmids which express constitutively a fluorescent protein gene (GFP, RFP, YFP or CFP) from a strong synthetic promoter and are suitable for the fluorescent labeling of a broad range of Enterobacteria. The amount of expressed fluorophore from these genetic constructs is such, that the contours of the cells can be delineated on the basis of the fluorescent signal only. In addition, labeling through the pUltra plasmids can be used successfully for fluorescence and confocal microscopy while unambiguous distinction of cells labeled with different colors can be carried out efficiently by microscopy or flow cytometry. We compare the labeling provided by the pUltra plasmids with that of another plasmid series encoding fluorescent proteins and we show that the pUltra constructs are vastly superior in signal intensity and discrimination power without having any detectable growth rate effects for the bacterial population. We also use the pUltra plasmids to produce mixtures of differentially labeled pathogenic Escherichia, Shigella and Salmonella species which we test during infection of mammalian cells. We find that even inside the host cell, different strains can be distinguished effortlessly based on their fluorescence. We, therefore, conclude that the pUltra plasmids are a powerful labeling tool especially useful for complex biological experiments such as the visualization of ecosystems of different bacterial species or of enteric pathogens in contact with their hosts

Costs and benefits of provocation in bacterial warfare.

Competition in animals involves a wide variety of aggressive behaviors. One of the most sophisticated strategies for a focal actor is to provoke a competitor into uncontrolled aggression toward other competitors. Like animals, bacteria rely on a broad spectrum of molecular weapons, some of which provoke potential rivals by triggering retaliation. While bacterial provocation is well documented, its potential adaptive value has received little attention. Here, we examine the costs and benefits of provocation using mathematical modeling and experiments with <i>Escherichia coli</i> strains encoding colicin toxins. We show that provocation is typically costly in one-to-one encounters because a provoking strain receives a strong reciprocal attack compared with nonprovoking strains. By contrast, provocation can be strongly beneficial in communities including more than two toxin-producing strains, especially when the provoker is shielded from, or resistant to, its opponents' toxins. In these scenarios, we demonstrate that the benefit of provocation derives from a "divide-and-conquer" effect by which aggression-provoking toxin producers force their competitors into increased reciprocal aggression, leading to their cross-elimination. Furthermore, we show that this effect can be mimicked by using antibiotics that promote warfare among strains in a bacterial community, highlighting the potential of provocation as an antimicrobial approach

Coordination of Dynamic Software Components with JavaBIP

JavaBIP allows the coordination of software components by clearly separating the functional and coordination aspects of the system behavior. JavaBIP implements the principles of the BIP component framework rooted in rigorous operational semantics. Recent work both on BIP and JavaBIP allows the coordination of static components defined prior to system deployment, i.e., the architecture of the coordinated system is fixed in terms of its component instances. Nevertheless, modern systems, often make use of components that can register and deregister dynamically during system execution. In this paper, we present an extension of JavaBIP that can handle this type of dynamicity. We use first-order interaction logic to define synchronization constraints based on component types. Additionally, we use directed graphs with edge coloring to model dependencies among components that determine the validity of an online system. We present the software architecture of our implementation, provide and discuss performance evaluation results.Comment: Technical report that accompanies the paper accepted at the 14th International Conference on Formal Aspects of Component Softwar

Acidithiobacillus thiooxidans secretome containing a newly described lipoprotein Licanantase enhances chalcopyrite bioleaching rate

The nature of the mineral–bacteria interphase where electron and mass transfer processes occur is a key element of the bioleaching processes of sulfide minerals. This interphase is composed of proteins, metabolites, and other compounds embedded in extracellular polymeric substances mainly consisting of sugars and lipids (Gehrke et al., Appl Environ Microbiol 64(7):2743–2747, 1998). On this respect, despite Acidithiobacilli—a ubiquitous bacterial genera in bioleaching processes (Rawlings, Microb Cell Fact 4(1):13, 2005)—has long been recognized as secreting bacteria (Jones and Starkey, J Bacteriol 82:788–789, 1961; Schaeffer and Umbreit, J Bacteriol 85:492–493, 1963), few studies have been carried out in order to clarify the nature and the role of the secreted protein component: the secretome. This work characterizes for the first time the sulfur (meta)secretome of Acidithiobacillus thiooxidans strain DSM 17318 in pure and mixed cultures with Acidithiobacillus ferrooxidans DSM 16786, identifying the major component of these secreted fractions as a single lipoprotein named here as Licanantase. Bioleaching assays with the addition of Licanantase-enriched concentrated secretome fractions show that this newly found lipoprotein as an active protein additive exerts an increasing effect on chalcopyrite bioleaching rate

Droplet printing reveals the importance of micron-scale structure for bacterial ecology

Bacteria often live in diverse communities where the spatial arrangement of strains and species is considered critical for their ecology. However, a test of this hypothesis requires manipulation at the fine scales at which spatial structure naturally occurs. Here we develop a droplet-based printing method to arrange bacterial genotypes across a sub-millimetre array. We print strains of the gut bacterium Escherichia coli that naturally compete with one another using protein toxins. Our experiments reveal that toxin-producing strains largely eliminate susceptible non-producers when genotypes are well-mixed. However, printing strains side-by-side creates an ecological refuge where susceptible strains can persist in large numbers. Moving to competitions between toxin producers reveals that spatial structure can make the difference between one strain winning and mutual destruction. Finally, we print different potential barriers between competing strains to understand how ecological refuges form, which shows that cells closest to a toxin producer mop up the toxin and protect their clonemates. Our work provides a method to generate customised bacterial communities with defined spatial distributions, and reveals that micron-scale changes in these distributions can drive major shifts in ecology

Intron variants of the p53 gene are associated with increased risk for ovarian cancer but not in carriers of BRCA1 or BRCA2 germline mutations

Two biallelic polymorphisms in introns 3 and 6 of the p53 gene were analysed for a possible risk-modifying effect for ovarian cancer. Germline DNA was genotyped from 310 German Caucasian ovarian cancer patients and 364 healthy controls. We also typed 124 affected and 276 unaffected female carriers with known deleterious BRCA1 or BRCA2 germline mutation from high-risk breast-ovarian cancer families. Genotyping was based on PCR and high-resolution gel electrophoresis. German ovarian cancer patients who carried the rare allele of the MspI restriction fragment length polymorphism (RELP) in intron 6 were found to have an overall 1.93-fold increased risk (95% confidence internal (CI) 1.27–2.91) which further increased with the age at diagnosis of 41–60 years (odds ratio (OR) 2.71, 95% CI 1.10–6.71 for 41–50 and OR 2.44, 95% CI 1.12–5.28 for 51–60). The 16 bp duplication polymorphism in intron 3 was in a strong linkage to the MspI RFLP. In BRCA1 or BRCA2 mutation carriers, no difference in allele frequency was observed for carriers affected or unaffected with ovarian cancer. Our data suggest that intronic polymorphisms of the p53 gene modify the risk for ovarian cancer patients but not in carriers with BRCA1 or BRCA2 mutations. © 1999 Cancer Research Campaig

Mutation analysis of genes that control the G1/S cell cycle in melanoma: TP53, CDKN1A, CDKN2A, and CDKN2B

BACKGROUND: The role of genes involved in the control of progression from the G1 to the S phase of the cell cycle in melanoma tumors in not fully known. The aim of our study was to analyse mutations in TP53, CDKN1A, CDKN2A, and CDKN2B genes in melanoma tumors and melanoma cell lines METHODS: We analysed 39 primary and metastatic melanomas and 9 melanoma cell lines by single-stranded conformational polymorphism (SSCP). RESULTS: The single-stranded technique showed heterozygous defects in the TP53 gene in 8 of 39 (20.5%) melanoma tumors: three new single point mutations in intronic sequences (introns 1 and 2) and exon 10, and three new single nucleotide polymorphisms located in introns 1 and 2 (C to T transition at position 11701 in intron 1; C insertion at position 11818 in intron 2; and C insertion at position 11875 in intron 2). One melanoma tumor exhibited two heterozygous alterations in the CDKN2A exon 1 one of which was novel (stop codon, and missense mutation). No defects were found in the remaining genes. CONCLUSION: These results suggest that these genes are involved in melanoma tumorigenesis, although they may be not the major targets. Other suppressor genes that may be informative of the mechanism of tumorigenesis in skin melanomas should be studied

The Eurasian Modern Pollen Database (EMPD), version 2

The Eurasian (née European) Modern Pollen Database (EMPD) was established in 2013 to provide a public database of high-quality modern pollen surface samples to help support studies of past climate, land cover, and land use using fossil pollen. The EMPD is part of, and complementary to, the European Pollen Database (EPD) which contains data on fossil pollen found in Late Quaternary sedimentary archives throughout the Eurasian region. The EPD is in turn part of the rapidly growing Neotoma database, which is now the primary home for global palaeoecological data. This paper describes version 2 of the EMPD in which the number of samples held in the database has been increased by 60 % from 4826 to 8134. Much of the improvement in data coverage has come from northern Asia, and the database has consequently been renamed the Eurasian Modern Pollen Database to reflect this geographical enlargement. The EMPD can be viewed online using a dedicated map-based viewer at https://empd2.github.io and downloaded in a variety of file formats at https://doi.pangaea.de/10.1594/PANGAEA.909130 (Chevalier et al., 2019)Swiss National Science Foundation | Ref. 200021_16959