Limits to the rate of adaptive substitution in sexual populations

In large populations, many beneficial mutations may be simultaneously available and may compete with one another, slowing adaptation. By finding the probability of fixation of a favorable allele in a simple model of a haploid sexual population, we find limits to the rate of adaptive substitution, [Formula: see text], that depend on simple parameter combinations. When variance in fitness is low and linkage is loose, the baseline rate of substitution is [Formula: see text], where [Formula: see text] is the population size, [Formula: see text] is the rate of beneficial mutations per genome, and [Formula: see text] is their mean selective advantage. Heritable variance [Formula: see text] in log fitness due to unlinked loci reduces [Formula: see text] by [Formula: see text] under polygamy and [Formula: see text] under monogamy. With a linear genetic map of length [Formula: see text] Morgans, interference is yet stronger. We use a scaling argument to show that the density of adaptive substitutions depends on [Formula: see text], [Formula: see text], [Formula: see text], and [Formula: see text] only through the baseline density: [Formula: see text]. Under the approximation that the interference due to different sweeps adds up, we show that [Formula: see text], implying that interference prevents the rate of adaptive substitution from exceeding one per centimorgan per 200 generations. Simulations and numerical calculations confirm the scaling argument and confirm the additive approximation for [Formula: see text]; for higher [Formula: see text], the rate of adaptation grows above [Formula: see text], but only very slowly. We also consider the effect of sweeps on neutral diversity and show that, while even occasional sweeps can greatly reduce neutral diversity, this effect saturates as sweeps become more common-diversity can be maintained even in populations experiencing very strong interference. Our results indicate that for some organisms the rate of adaptive substitution may be primarily recombination-limited, depending only weakly on the mutation supply and the strength of selection.     

Robustness and information propagation in attractors of random boolean networks

Attractors represent the long-term behaviors of Random Boolean Networks. We study how the amount of information propagated between the nodes when on an attractor, as quantified by the average pairwise mutual information ([Formula: see text]), relates to the robustness of the attractor to perturbations ([Formula: see text]). We find that the dynamical regime of the network affects the relationship between [Formula: see text] and [Formula: see text]. In the ordered and chaotic regimes, [Formula: see text] is anti-correlated with [Formula: see text], implying that attractors that are highly robust to perturbations have necessarily limited information propagation. Between order and chaos (for so-called "critical" networks) these quantities are uncorrelated. Finite size effects cause this behavior to be visible for a range of networks, from having a sensitivity of 1 to the point where [Formula: see text] is maximized. In this region, the two quantities are weakly correlated and attractors can be almost arbitrarily robust to perturbations without restricting the propagation of information in the network.     

Modelling transmission of vector-borne pathogens shows complex dynamics when vector feeding sites are limited

The relationship between species richness and the prevalence of vector-borne disease has been widely studied with a range of outcomes. Increasing the number of host species for a pathogen may decrease infection prevalence (dilution effect), increase it (amplification), or have no effect. We derive a general model, and a specific implementation, which show that when the number of vector feeding sites on each host is limiting, the effects on pathogen dynamics of host population size are more complex than previously thought. The model examines vector-borne disease in the presence of different host species that are either competent or incompetent (i.e. that cannot transmit the pathogen to vectors) as reservoirs for the pathogen. With a single host species present, the basic reproduction ratio R(0) is a non-monotonic function of the population size of host individuals (H), i.e. a value [Formula: see text] exists that maximises R(0). Surprisingly, if [Formula: see text] a reduction in host population size may actually increase R(0). Extending this model to a two-host species system, incompetent individuals from the second host species can alter the value of [Formula: see text] which may reverse the effect on pathogen prevalence of host population reduction. We argue that when vector-feeding sites on hosts are limiting, the net effect of increasing host diversity might not be correctly predicted using simple frequency-dependent epidemiological models.     

Critical motor number for fractional steps of cytoskeletal filaments in gliding assays

In gliding assays, filaments are pulled by molecular motors that are immobilized on a solid surface. By varying the motor density on the surface, one can control the number [Formula: see text] of motors that pull simultaneously on a single filament. Here, such gliding assays are studied theoretically using Brownian (or Langevin) dynamics simulations and taking the local force balance between motors and filaments as well as the force-dependent velocity of the motors into account. We focus on the filament stepping dynamics and investigate how single motor properties such as stalk elasticity and step size determine the presence or absence of fractional steps of the filaments. We show that each gliding assay can be characterized by a critical motor number, [Formula: see text]. Because of thermal fluctuations, fractional filament steps are only detectable as long as [Formula: see text]. The corresponding fractional filament step size is [Formula: see text] where [Formula: see text] is the step size of a single motor. We first apply our computational approach to microtubules pulled by kinesin-1 motors. For elastic motor stalks that behave as linear springs with a zero rest length, the critical motor number is found to be [Formula: see text], and the corresponding distributions of the filament step sizes are in good agreement with the available experimental data. In general, the critical motor number [Formula: see text] depends on the elastic stalk properties and is reduced to [Formula: see text] for linear springs with a nonzero rest length. Furthermore, [Formula: see text] is shown to depend quadratically on the motor step size [Formula: see text]. Therefore, gliding assays consisting of actin filaments and myosin-V are predicted to exhibit fractional filament steps up to motor number [Formula: see text]. Finally, we show that fractional filament steps are also detectable for a fixed average motor number [Formula: see text] as determined by the surface density (or coverage) of the motors on the substrate surface.     

Unifying Time to Contact Estimation and Collision Avoidance across Species

The [Formula: see text]-function and the [Formula: see text]-function are phenomenological models that are widely used in the context of timing interceptive actions and collision avoidance, respectively. Both models were previously considered to be unrelated to each other: [Formula: see text] is a decreasing function that provides an estimation of time-to-contact (ttc) in the early phase of an object approach; in contrast, [Formula: see text] has a maximum before ttc. Furthermore, it is not clear how both functions could be implemented at the neuronal level in a biophysically plausible fashion. Here we propose a new framework - the corrected modified Tau function - capable of predicting both [Formula: see text]-type ("[Formula: see text]") and [Formula: see text]-type ("[Formula: see text]") responses. The outstanding property of our new framework is its resilience to noise. We show that [Formula: see text] can be derived from a firing rate equation, and, as [Formula: see text], serves to describe the response curves of collision sensitive neurons. Furthermore, we show that [Formula: see text] predicts the psychophysical performance of subjects determining ttc. Our new framework is thus validated successfully against published and novel experimental data. Within the framework, links between [Formula: see text]-type and [Formula: see text]-type neurons are established. Therefore, it could possibly serve as a model for explaining the co-occurrence of such neurons in the brain.     

Mapping of redox state of mitochondrial cytochromes in live cardiomyocytes using Raman microspectroscopy

This paper presents a nonivasive approach to study redox state of reduced cytochromes [Formula: see text], [Formula: see text] and [Formula: see text] of complexes II and III in mitochondria of live cardiomyocytes by means of Raman microspectroscopy. For the first time with the proposed approach we perform studies of rod- and round-shaped cardiomyocytes, representing different morphological and functional states. Raman mapping and cluster analysis reveal that these cardiomyocytes differ in the amounts of reduced cytochromes [Formula: see text], [Formula: see text] and [Formula: see text]. The rod-shaped cardiomyocytes possess uneven distribution of reduced cytochromes [Formula: see text], [Formula: see text] and [Formula: see text] in cell center and periphery. Moreover, by means of Raman spectroscopy we demonstrated the decrease in the relative amounts of reduced cytochromes [Formula: see text], [Formula: see text] and [Formula: see text] in the rod-shaped cardiomyocytes caused by H(2)O(2)-induced oxidative stress before any visible changes. Results of Raman mapping and time-dependent study of reduced cytochromes of complexes II and III and cytochrome [Formula: see text] in cardiomyocytes are in a good agreement with our fluorescence indicator studies and other published data.     

Reduced set of virulence genes allows high accuracy prediction of bacterial pathogenicity in humans

Although there have been great advances in understanding bacterial pathogenesis, there is still a lack of integrative information about what makes a bacterium a human pathogen. The advent of high-throughput sequencing technologies has dramatically increased the amount of completed bacterial genomes, for both known human pathogenic and non-pathogenic strains; this information is now available to investigate genetic features that determine pathogenic phenotypes in bacteria. In this work we determined presence/absence patterns of [Formula: see text] different virulence-related genes among more than [Formula: see text] finished bacterial genomes from both human pathogenic and non-pathogenic strains, belonging to different taxonomic groups (i.e: Actinobacteria, Gammaproteobacteria, Firmicutes, etc.). An accuracy of 95% using a cross-fold validation scheme with in-fold feature selection is obtained when classifying human pathogens and non-pathogens. A reduced subset of highly informative genes ([Formula: see text]) is presented and applied to an external validation set. The statistical model was implemented in the BacFier v1.0 software (freely available at [Formula: see text]), that displays not only the prediction (pathogen/non-pathogen) and an associated probability for pathogenicity, but also the presence/absence vector for the analyzed genes, so it is possible to decipher the subset of virulence genes responsible for the classification on the analyzed genome. Furthermore, we discuss the biological relevance for bacterial pathogenesis of the core set of genes, corresponding to eight functional categories, all with evident and documented association with the phenotypes of interest. Also, we analyze which functional categories of virulence genes were more distinctive for pathogenicity in each taxonomic group, which seems to be a completely new kind of information and could lead to important evolutionary conclusions.     

(13)C NMR Reveals No Evidence of n-π* Interactions in Proteins

An [Formula: see text] interaction between neighboring carbonyl groups has been postulated to stabilize protein structures. Such an interaction would affect the [Formula: see text]C chemical shielding of the carbonyl groups, whose paramagnetic component is dominated by [Formula: see text] and [Formula: see text] excitations. Model compound calculations indicate that both the interaction energetics and the chemical shielding of the carbonyl group are instead dominated by a classical dipole-dipole interaction. A set of high-resolution protein structures with associated carbonyl [Formula: see text]C chemical shift assignments verifies this correlation and provides no evidence for an inter-carbonyl [Formula: see text] interaction.     

Comparative analysis of numerical integration schemes of density equation for a computational model of bone remodelling

In this study, a computational model of bone remodelling problem as proposed by Weinans et al. (1992) is described and solved by other temporal integration techniques different from the Euler scheme. This model considers three types of numerical integration schemes of the evolution of the material density during the remodelling: Euler, Heun and Runge-Kutta methods. Also the strain and the density field are obtained inside each element, at Gauss points or at the nodes of the mesh. A square plate with 1.00?m of side subjected to non-uniform pressure is simulated with two meshes of quadrilateral element with size [Formula: see text] and [Formula: see text]?m. Two increments time size: [Formula: see text] and [Formula: see text] days are used. The results show that Euler, Heun and Runge-Kutta's methods correctly approached the problem of bone remodelling and that there were no appreciable differences in the patterns obtained by the mesh and time step used. In contrast, using an element-based approach and node-based approach, substantial differences were produced in bone remodelling density pattern. 'Chess board' type discontinuities were found in the element approach near the applied pressure area, as were well-defined columns away from this. The node-based approach showed continuity in density distribution. These patterns were well represented by the methods for resolving the density equation. This study concluded that any method of time integration could be used for these meshes and time steps size.     

Specificity of Hpa II and Hae III DNA methylases

The methylases M.HaeIII and M.HpaII recognize the tetranucleotide sequences [Formula: see text] and [Formula: see text] respectively, in DNA, and transfer a methyl group from S-adenosylmethionine to the 5-position of cytosine on each strand as indicated by the asterisks. Restriction endonuclease R.HaeIII does not cleave the methylated sequence [Formula: see text] but can cleave [Formula: see text] in which methylation is introduced on the unnatural external cytosine positions. Similarly, R.HpaII does not cleave [Formula: see text] but can cleave [Formula: see text].Images     

A Mathematical Model of Rift Valley Fever with Human Host

Rift Valley Fever is a vector-borne disease mainly transmitted by mosquito. To gain some quantitative insights into its dynamics, a deterministic model with mosquito, livestock, and human host is formulated as a system of nonlinear ordinary differential equations and analyzed. The disease threshold [Formula: see text] is computed and used to investigate the local stability of the equilibria. A sensitivity analysis is performed and the most sensitive model parameters to the measure of initial disease transmission [Formula: see text] and the endemic equilibrium are determined. Both [Formula: see text] and the disease prevalence in mosquitoes are more sensitive to the natural mosquito death rate, d(m). The disease prevalence in livestock and humans are more sensitive to livestock and human recruitment rates, [Formula: see text] and [Formula: see text], respectively, suggesting isolation of livestock from humans is a viable preventive strategy during an outbreak. Numerical simulations support the analytical results in further exploring theoretically the long-term dynamics of the disease at the population level.     

Exploiting magnetic resonance angiography imaging improves model estimation of BOLD signal

The change of BOLD signal relies heavily upon the resting blood volume fraction ([Formula: see text]) associated with regional vasculature. However, existing hemodynamic data assimilation studies pretermit such concern. They simply assign the value in a physiologically plausible range to get over ill-conditioning of the assimilation problem and fail to explore actual [Formula: see text]. Such performance might lead to unreliable model estimation. In this work, we present the first exploration of the influence of [Formula: see text] on fMRI data assimilation, where actual [Formula: see text] within a given cortical area was calibrated by an MR angiography experiment and then was augmented into the assimilation scheme. We have investigated the impact of [Formula: see text] on single-region data assimilation and multi-region data assimilation (dynamic cause modeling, DCM) in a classical flashing checkerboard experiment. Results show that the employment of an assumed [Formula: see text] in fMRI data assimilation is only suitable for fMRI signal reconstruction and activation detection grounded on this signal, and not suitable for estimation of unobserved states and effective connectivity study. We thereby argue that introducing physically realistic [Formula: see text] in the assimilation process may provide more reliable estimation of physiological information, which contributes to a better understanding of the underlying hemodynamic processes. Such an effort is valuable and should be well appreciated.     

Architecture of the Outer Membrane of Escherichia coli III. Protein-Lipopolysaccharide Complexes in Intramembraneous Particles

In a previous paper (A. Verkleij, L. van Alphen, J. Bijvelt, and B. Lugtenberg, Biochim. Biophys. Acta 466:269-282, 1977) we have hypothesized that particles on the outer fracture face of the outer membrane ([Formula: see text]), with corresponding pits on the inner fracture face of the outer membrane ([Formula: see text]), consist of lipopolysaccharide (LPS) aggregates stabilized by divalent cations and that they might contain protein and/or phospholipid. In the present paper the roles of LPS, cations, and proteins in these [Formula: see text] particles are described more extensively, using a strain that lacks the major outer membrane proteins, b, c, and d (b(-) c(-) d(-)), and has a reduction in the number of [Formula: see text] particles of 75%. To study the role of divalent cations in the formation of [Formula: see text] particles, these b(-) c(-) d(-) cells were grown or incubated with Ca(2+), Mg(2+), or putrescine. The presence of Ca(2+) resulted in the appearance of many [Formula: see text] particles and [Formula: see text] pits. Mg(2+) and putrescine were less effective than Ca(2+). Introduction of these particles was not accompanied by alterations in the relative amounts of LPS and cell envelope proteins. Ca(2+) treatment of a heptoseless derivative of a b(-) c(-) d(-) strain did not result in morphological changes. Incubation of Ca(2+)-treated cells with ethylenediaminetetraacetate caused the disappearance of the introduced particles as well as the release of more than 60% of the cellular LPS. These results strongly support the hypothesis that LPS is involved in the formation of [Formula: see text] particles and [Formula: see text] pits. The roles of various outer membrane proteins in the formation of [Formula: see text] particles were studied by comparing the freeze-fracture morphology of b(-) c(-) d(-) cells with that of cells which contain one of the outer membrane proteins b, c, d, and e or the receptor protein for bacteriophage lambda. The results showed that the presence of any of these five proteins in a b(-) c(-) d(-) background resulted in a large increase in the number of [Formula: see text] particles and [Formula: see text] pits, indicating that these proteins are, independent of each other, involved in the formation of [Formula: see text] particles and [Formula: see text] pits. The simplest explanation for the results is that in wild-type cells each particle consists of LPS complexed with some molecules of a single protein species, stabilized by either divalent cations or polyamines. It is hypothesized that the outer membrane of the wild-type cell contains a heterogeneous population of particles, of which 75% consists of protein b-LPS, protein c-LPS, and protein d-LPS particles. A function of these particles as aqueous pores is proposed.     

Trans-Theta Logistics: A New Family of Population Growth Sigmoid Functions

Sigmoid functions have been applied in many areas to model self limited population growth. The most popular functions; General Logistic (GL), General von Bertalanffy (GV), and Gompertz (G), comprise a family of functions called Theta Logistic ([Formula: see text] L). Previously, we introduced a simple model of tumor cell population dynamics which provided a unifying foundation for these functions. In the model the total population (N) is divided into reproducing (P) and non-reproducing/quiescent (Q) sub-populations. The modes of the rate of change of ratio P/N was shown to produce GL, GV or G growth. We now generalize the population dynamics model and extend the possible modes of the P/N rate of change. We produce a new family of sigmoid growth functions, Trans-General Logistic (TGL), Trans-General von Bertalanffy (TGV) and Trans-Gompertz (TG)), which as a group we have named Trans-Theta Logistic (T [Formula: see text] L) since they exist when the [Formula: see text] L are translated from a two parameter into a three parameter phase space. Additionally, the model produces a new trigonometric based sigmoid (TS). The [Formula: see text] L sigmoids have an inflection point size fixed by a single parameter and an inflection age fixed by both of the defining parameters. T [Formula: see text] L and TS sigmoids have an inflection point size defined by two parameters in bounding relationships and inflection point age defined by three parameters (two bounded). While the Theta Logistic sigmoids provided flexibility in defining the inflection point size, the Trans-Theta Logistic sigmoids provide flexibility in defining the inflection point size and age. By matching the slopes at the inflection points we compare the range of values of inflection point age for T [Formula: see text] L versus [Formula: see text] L for model growth curves.     

Location of the Fracture Faces Within the Cell Envelope of Acinetobacter Species Strain MJT/F5/5

The cell wall of the gram-negative bacterium Acinetobacter species strain MJT/F5/5 shows in thin section an external “additional” layer, an outer membrane, an intermediate layer, and a dense layer. Negatively stained preparations showed that the additional layer is composed of hexagonally arranged subunits. In glycerol-treated preparations, freeze-etching revealed that the cell walls consist of four layers, with the main plane of fracture between layers cw 2 and cw 3. The surface of [Formula: see text] 2 consisted of densely packed particles, whereas [Formula: see text] 3 appeared to be fibrillar. In cell envelopes treated with lysozyme by various methods, the removal of the dense layer has detached the outer membrane and additional layer from the underlying layers, as shown in thin sections. When freeze-etched in the absence of glycerol, these detached outer membranes with additional layers fractured to reveal both the faces [Formula: see text] 2 and [Formula: see text] 3 with their characteristic surface structures, and, in addition, both the external and internal etched surfaces were revealed. This experiment provided conclusive evidence that the main fracture plane in the cell wall lies within the interior of the outer membrane. This and other evidence showed that the corresponding layers in thin sections and freeze-etched preparations are: the additional layer, cw 1; the outer membrane, cw (2 + 3); and the intermediate and dense layers together from cw 4. Because of similarities in structure between this Acinetobacter and other gram-negative bacteria, it seemed probable that the interior of the outer membrane is the plane most liable to fracture in the cell walls of most gram-negative bacteria.     

The dynamics, causes and possible prevention of hepatitis e outbreaks

Rapidly spreading infectious diseases are a serious risk to public health. The dynamics and the factors causing outbreaks of these diseases can be better understood using mathematical models, which are fit to data. Here we investigate the dynamics of a Hepatitis E outbreak in the Kitgum region of northern Uganda during 2007 to 2009. First, we use the data to determine that [Formula: see text] is approximately 2.25 for the outbreak. Secondly, we use a model to estimate that the critical level of latrine and bore hole coverages needed to eradicate the epidemic is at least [Formula: see text] and [Formula: see text] respectively. Lastly, we further investigate the relationship between the co-infection factor for malaria and Hepatitis E on the value of [Formula: see text] for Hepatitis E. Taken together, these results provide us with a better understanding of the dynamics and possible causes of Hepatitis E outbreaks.     

Copper(I)-alkyl sulfide and -cysteine tri-nuclear clusters as models for metallo proteins: a structural density functional analysis

After having set up the computational methodology for Cu(I)-sulfur systems as models for copper proteins, namely using the simple ligands H(2)S, HS(-), CH(3)SH, and CH(3)S(-), the Cu(I)-Cysteine systems have been investigated: [Cu(I)( S -H(2)Cys) (n) ](+) (H(2)Cys, cysteine, NH(2),SH,COOH) [Cu(I)( S -HCys) (n) ](1-) (n) (NH(2),S(-),COOH). Finally, the structures for bi-nuclear [Formula: see text] (Et, CH(2)CH(3)), [Formula: see text] and tri-nuclear [Cu(I)( S -SH)](3), [Formula: see text], [Formula: see text], [Formula: see text], [Formula: see text] (NH(2),SH,COOH), [Formula: see text] (NH(2),S(-),COOH, and NH(2),SH,COO(-)), as well as [Formula: see text] (NH(2),S(-),COO(-)), were also optimized to mimic the active center for a metallo-chaperone copper transport protein (CopZ). The X-ray structures for the biomolecules were matched fairly well as regards the Cu-S bond distances and Cu…Cu contact distances in the case the model cysteine S atom is deprotonated. Upon protonation of ligand S atoms, the conformation of clusters is altered and might bring about the di- and tri-nuclear core breakage. These findings suggest that subtle protonation/deprotonation steps, i.e. small and/or local pH changes play a significant role for copper transport processes.     

The biosynthesis of 4-hydroxycoumarin and dicoumarol by Aspergillus fumigatus Fresenius

A strain of Aspergillus fumigatus Fresenius, isolated from spoiled hay, converts melilotic acid (o-hydroxyphenylpropionic acid) and o-coumaric acid into 4-hydroxycoumarin and dicoumarol. The sequence is shown to be melilotic acid (I) [Formula: see text] coumaric acid (IV) [Formula: see text] beta-hydroxymelilotic acid (II) [Formula: see text] beta-oxomelilotic acid (III) [Formula: see text] 4-hydroxycoumarin (VI), on the basis of (1) studies on the formation of postulated intermediates, (2) experiments with isotopically labelled materials and (3) sequential enzyme induction. In the presence of semicarbazide, o-coumaraldehyde is formed from o-coumaric acid: there is no evidence, however, that this lies on the normal metabolic pathway.     

The effects of dissolved oxygen levels on the metabolic interaction between digestion and locomotion in Cyprinid fishes with different locomotive and digestive performances

To test whether the effects of water oxygen concentration ([O(2)]) on the metabolic interaction between locomotion and digestion differ between fish species with different locomotive and digestive behaviours in normoxia, we investigated the swimming performance of fasted and fed fish at water [O(2)] of 1, 2 and 8 (normoxia) mg L(-1) (2.5, 5 and 20 kPa) at 25°C in three juvenile Cyprinidae fish species: goldfish (Carassius auratus), common carp (Cyprinus carpio) and qingbo (Spinibarbus sinensis). Digestion, taxon and water [O(2)] all had significant effects on the pre-exercise oxygen consumption rate [Formula: see text] and the swimming performance (P < 0.05). Among the three fishes, qingbo showed the highest swimming performance and the lowest feeding [Formula: see text] at the saturated water [O(2)], and its active oxygen consumption rate [Formula: see text] and critical swimming speed (U (crit)) decreased the most with decreases in water [O(2)]. Qingbo exhibited a locomotion-priority metabolic mode at all three water [O(2)]. Digestion was sacrificed to locomotion in a postprandial swimming situation, but fed qingbo could not maintain their U (crit) at water [O(2)] of 2 and 1 mg L(-1). Goldfish showed the lowest swimming performance and the highest feeding [Formula: see text] at the saturated water [O(2)]. They exhibited a digestion-priority metabolic mode at high water [O(2)]. However, with a decrease in water [O(2)], the feeding [Formula: see text] decreased more acutely than the respiratory capacity; thus, digestion and locomotion performed independently in a postprandial swimming situation (i.e., an additive metabolic mode) at a water [O(2)] of 1 mg L(-1). The common carp showed moderate and balanced swimming performance and feeding [Formula: see text] at the saturated water [O(2)], and exhibited an additive metabolic mode at all 3 water [O(2)], because digestion, swimming and respiratory capacities decreased in parallel with the decrease in water [O(2)].     

Fracture Faces in the Cell Envelope of Escherichia coli

Freeze-fracturing of Escherichia coli cells in the presence of 30% (v/v) glycerol resulted in a double cleavage of the cell envelope exposing two convex and two concave fracture faces ([Formula: see text], [Formula: see text] and [Formula: see text], [Formula: see text]) with characteristic patterns. Complementary replicas revealed the relationship of the fracture faces to their corresponding fracture planes. The inner fracture plane splits the plasma membrane at one particular level. Apparently the outer fracture plane was located in the outer part of the wall, as it was separated by a layer ([Formula: see text]) from the fractured profile (CW1) presumably corresponding to the murein layer. The outer fracture plane did alternate toward the cell periphery, exposing complementary smooth areas ([Formula: see text] and [Formula: see text]). When cells were freeze-fractured in the absence of glycerol, the outer cell surface appeared as an etching face rather than a fracture face. A schematic representation of the relative location of the different fracture faces in the E. coli cell envelope is given.