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The mechanics of adhesion polymers and their role in bacterial attachment
Umeå University, Faculty of Science and Technology, Department of Physics. (Biophysics and biophotonics group)
2015 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Bacterial resistance to antibiotics is increasing at a high rate in both developing and developed countries. To circumvent the problem of drug-resistant bacterial pathogens, we need to develop new effective methods, substances, and materials that can disarm and prevent them from causing infections. However, to do this we first need to find new possible targets in bacteria to approach and novel strategies to apply.Escherichia coli (E. coli) bacteria is a normal member of the intestinal microflora of humans and mammals, but frequently cause diverse intestinal and external diseases by means of virulence factors, which leads to hundreds of million sick people each year with a high mortality rate. An E. coli bacterial infection starts with adhesion to a host cell using cell surface expressed adhesion polymers, called adhesion pili. Depending on the local environment different types of pili are expressed by the bacteria. For example, bacteria found in the gastrointestinal tract commonly express different pili in comparison to those found in the urinary tract and respiratory tract. These pili, which are vital for bacterial adhesion, thereby serve as a new possible approach in the fight against bacterial infections by targeting and disabling these structures using novel chemicals. However, in order to develop such chemicals, better understanding of these pili is needed.Optical tweezers (OT) can measure and apply forces up to a few hundred pN with sub-pN force resolution and have shown to be an excellent tool for investigating mechanical properties of adhesion pili. It has been found that pili expressed by E. coli have a unique and complex force-extension response that is assumed to be important for the ability of bacteria to initiate and maintain attachment to the host cells. However, their mechanical functions and the advantage of specific mechanical functions, especially in the initial attachment process, have not yet been fully understood.In this work, a detailed description of the pili mechanics and their role during cell adhesion is presented. By using results from optical tweezers force spectroscopy experiments in combination with physical modeling and numerical simulations, we investigated how pili can act as “shock absorbers” through uncoiling and thereby lower the fluid force acting on a bacterium. Our result demonstrate that the dynamic uncoiling capability of the helical part of the adhesion pili modulate the force to fit the optimal lifetime of its adhesin (the protein that binds to the receptor on the host cell), ensuring a high survival probability of the bond.iiiSince the attachment process is in proximity of a surface we also investigated the influence of tether properties and the importance of different surface corrections and additional force components to the Stokes drag force during simulations. The investigation showed that the surface corrections to the Stokes drag force and the Basset force cannot be neglected when simulating survival probability of a bond, since that can overestimate the probability by more than an order of magnitude.Finally, a theoretical and experimental framework for two separate methods was developed. The first method can detect the presence of pili on single cells using optical tweezers. We verified the method using silica microspheres coated with a polymer brush and E. coli bacteria expressing; no pili, P pili, and type 1 pili, respectively. The second method was based on digital holography microscopy. Using the diffraction of semi-transparent object such as red blood cells, we showed that this method can reconstruct the axial position and detect morphological changes of cells.

Place, publisher, year, edition, pages
Umeå: Umeå universitet , 2015. , 65 p.
Keyword [en]
Pili, optical tweezers, bacterial adhesion, fimbriae, uncoiling
National Category
Other Physics Topics Biophysics
Identifiers
URN: urn:nbn:se:umu:diva-109524ISBN: 978-91-7601-331-1 (print)OAI: oai:DiVA.org:umu-109524DiVA: diva2:857819
Public defence
2015-10-23, Naturvetarhuset, N420, Umeå universitet, Umeå, 13:00 (English)
Opponent
Supervisors
Available from: 2015-10-02 Created: 2015-09-30 Last updated: 2015-10-02Bibliographically approved
List of papers
1. Biomechanical and Structural features of CS2 fimbriae of Enterotoxigenic Escherichia coli 
Open this publication in new window or tab >>Biomechanical and Structural features of CS2 fimbriae of Enterotoxigenic Escherichia coli 
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2015 (English)In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 109, no 1, 49-56 p.Article in journal (Refereed) Published
Abstract [en]

Enterotoxigenic Escherichia coli (ETEC) are a major cause of diarrhea worldwide, and infection of children in underdeveloped countries often leads to high mortality rates. Isolated ETEC express a plethora of colonization factors (fimbriae/pili), of which CFA/I and CFA/II that are assembled via the alternate chaperone pathway (ACP), are amongst the most common. Fimbriae are filamentous structures, whose shafts are primarily composed of helically arranged single pilin-protein subunits, with a unique biomechanical capability allowing them to unwind and rewind. A sustained ETEC infection, under adverse conditions of dynamic shear forces, is primarily attributed to this biomechanical feature of ETEC fimbriae. Recent understandings about the role of fimbriae as virulence factors are pointing to an evolutionary adaptation of their structural and biomechanical features. In this work, we investigated the biophysical properties of CS2 fimbriae from the CFA/II group. Homology modelling its major structural subunit CotA reveals structural clues and these are related to the niche in which they are expressed. Using optical tweezers force spectroscopy we found that CS2 fimbriae unwind at a constant force of 10 pN and have a corner velocity of 1300 nm/s, i.e., the velocity at which the force required for unwinding rises exponentially with increased speed. The biophysical properties of CS2 fimbriae assessed in this work classify them into a low-force unwinding group of fimbriae together with the CFA/I and CS20 fimbriae expressed by ETEC strains. The three fimbriae are expressed by ETEC, colonize in similar gut environments, and exhibit similar biophysical features, but differ in their biogenesis. Our observation suggests that the environment has a strong impact on the biophysical characteristics of fimbriae expressed by ETEC.

Place, publisher, year, edition, pages
Elsevier, 2015
Keyword
pili, optical tweezers, bacteria, pathogenesis, virulence factors
National Category
Other Physics Topics
Research subject
Physics
Identifiers
urn:nbn:se:umu:diva-103247 (URN)10.1016/j.bpj.2015.05.022 (DOI)
Funder
Swedish Research Council, 621-2013-5379
Note

This work was supported by NIH (GM05722 and RR025434 to E.B.), the Swedish Research Council (621-2013-5379 to M.A. and the Carl Trygger foundation to M.A.

Available from: 2015-05-19 Created: 2015-05-19 Last updated: 2017-12-04Bibliographically approved
2. Rigid multibody simulation of a helix-like structure: the dynamics of bacterial adhesion pili
Open this publication in new window or tab >>Rigid multibody simulation of a helix-like structure: the dynamics of bacterial adhesion pili
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2015 (English)In: European Biophysics Journal, ISSN 0175-7571, E-ISSN 1432-1017, Vol. 44, no 5, 291-300 p.Article in journal (Refereed) Published
Abstract [en]

We present a coarse-grained rigid multibody model of a subunit assembled helix-like polymer, e.g., adhesion pili expressed by bacteria, that is capable of describing the polymer's force-extension response. With building blocks representing individual subunits, the model appropriately describes the complex behavior of pili expressed by the gram-negative uropathogenic Escherichia coli bacteria under the action of an external force. Numerical simulations show that the dynamics of the model, which include the effects of both unwinding and rewinding, are in good quantitative agreement with the characteristic force-extension response as observed experimentally for type 1 and P pili. By tuning the model, it is also possible to reproduce the force-extension response in the presence of anti-shaft antibodies, which dramatically changes the mechanical properties. Thus, the model and results in this work give enhanced understanding of how a pilus unwinds under the action of external forces and provide a new perspective of the complex bacterial adhesion processes.

Keyword
Fimbriae, Escherichia coli, Optical tweezers, Simulations, Force spectroscopy
National Category
Biophysics
Identifiers
urn:nbn:se:umu:diva-106000 (URN)10.1007/s00249-015-1021-1 (DOI)000356143100002 ()25851543 (PubMedID)
Available from: 2015-07-07 Created: 2015-07-03 Last updated: 2017-12-04Bibliographically approved
3. Helix-like bio-polymers can act as effective dampers for bacteria in flows
Open this publication in new window or tab >>Helix-like bio-polymers can act as effective dampers for bacteria in flows
2012 (English)In: European Biophysics Journal, ISSN 0175-7571, E-ISSN 1432-1017, Vol. 41, no 6, 551-560 p.Article in journal (Refereed) Published
Abstract [en]

Biopolymers are vital structures for many liv- ing organisms; for a variety of bacteria, adhesion polymers play a crucial role for the initiation of colonization. Some bacteria express, on their surface, attachment organelles (pili) that comprise subunits formed into stiff helix-like structures that possess unique biomechanical properties. These helix-like structures possess a high degree of flexi- bility that gives the biopolymers a unique extendibility. This has been considered beneficial for piliated bacteria adhering to host surfaces in the presence of a fluid flow. We show in this work that helix-like pili have the ability to act as efficient dampers of force that can, for a limited time, lower the load on the force-mediating adhesin-receptor bond on the tip of an individual pilus. The model presented is applied to bacteria adhering with a single pilus of either of the two most common types expressed by uropathogenic Escherichia coli, P or type 1 pili, subjected to realistic flows. The results indicate that for moderate flows (~25 mm/s) the force experienced by the adhesin-receptor interaction at the tip of the pilus can be reduced by a factor of ~6 and ~4, respectively. The uncoiling ability pro- vides a bacterium with a ‘‘go with the flow’’ possibility that acts as a damping. It is surmised that this can be an important factor for the initial part of the adhesion process, in particular in turbulent flows, and thereby be of use for bacteria in their striving to survive a natural defense such as fluid rinsing actions.

Place, publisher, year, edition, pages
Springer: , 2012
Keyword
fimbriae, pili, uncoiling, damping, bacterial adhesion
National Category
Other Physics Topics Biophysics
Research subject
Physics; biology
Identifiers
urn:nbn:se:umu:diva-54018 (URN)10.1007/s00249-012-0814-8 (DOI)
Available from: 2012-04-11 Created: 2012-04-11 Last updated: 2017-12-07Bibliographically approved
4. The shaft of the type 1 fimbriae regulates an externalforce to match the FimH catch bond
Open this publication in new window or tab >>The shaft of the type 1 fimbriae regulates an externalforce to match the FimH catch bond
2013 (English)In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 104, no 10, 2137-2148 p.Article in journal (Refereed) Published
Abstract [en]

Type 1 fimbriae mediate adhesion of uropathogenic Escherichia coli to host cells. It has been hypothesized that due to their ability to uncoil under exposure to force, fimbriae can reduce fluid shear stress on the adhesin-receptor interaction by which the bacterium adheres to the surface. In this work, we develop a model that describes how the force on the adhesin-receptor interaction of a type 1 fimbria varies as a bacterium is affected by a time-dependent fluid flow mimicking in vivo conditions. The model combines in vivo hydrodynamic conditions with previously assessed biomechanical properties of the fimbriae. Numerical methods are used to solve for the motion and adhesion force under the presence of time-dependent fluid profiles. It is found that a bacterium tethered with a type 1 pilus will experience significantly reduced shear stress for moderate to high flow velocities and that the maximum stress the adhesin will experience is limited to ∼120 pN, which is sufficient to activate the conformational change of the FimH adhesin into its stronger state but also lower than the force required for breaking it under rapid loading. Our model thus supports the assumption that the type 1 fimbria shaft and the FimH adhesin-receptor interaction are optimized to each other, and that they give piliated bacteria significant advantages in rapidly changing fluidic environments.

Keyword
type 1, E coli, pili, optical tweezers, FimH
National Category
Biophysics Physical Sciences
Research subject
Physics
Identifiers
urn:nbn:se:umu:diva-64463 (URN)10.1016/j.bpj.2013.03.059 (DOI)000319318400008 ()23708354 (PubMedID)
Available from: 2013-01-29 Created: 2013-01-29 Last updated: 2017-12-06Bibliographically approved
5. Tethered cells in fluid flows: beyond the Stokes’ drag force approach
Open this publication in new window or tab >>Tethered cells in fluid flows: beyond the Stokes’ drag force approach
2015 (English)In: Physical Biology, ISSN 1478-3967, E-ISSN 1478-3975, Vol. 12, 056006Article in journal (Refereed) Published
Abstract [en]

Simulations of tethered cells in viscous sub-layers are frequently performed using the Stokes' drag force, but without taking into account contributions from surface corrections, lift forces, buoyancy, the Basset force, the cells' finite inertia, or added mass. In this work, we investigate to what extent such contributions, under a variety of hydrodynamic conditions, influence the force at the anchor point of a tethered cell and the survival probability of a bacterium that is attached to a host by either a slip or a catch bond via a tether with a few different biomechanical properties. We show that a consequence of not including some of these contributions is that the force to which a bond is exposed can be significantly underestimated; in general by similar to 32-46%, where the influence of the surface corrections dominate ( the parallel and normal correction coefficients contribute similar to 5-8 or similar to 23-26%, respectively). The Basset force is a major contributor, up to 20%, for larger cells and shear rates. The lift force and inertia contribute when cells with radii >3 mu m have shear rates>2000 s(-1). Buoyancy contributes significantly for cells with radii > 3 mu m for shear rates<10 s(-1). Since the lifetime of a bond depends strongly on the force, both the level of approximation and the biomechanical model of the tether significantly affect the survival probability of tethered bacteria. For a cell attached by a FimH-mannose bond and an extendable tether with a shear rate of 3000 s(-1), neglecting the surface correction coefficients or the Basset force can imply that the survival probability is overestimated by more than an order of magnitude. This work thus shows that in order to quantitatively assess bacterial attachment forces and survival probabilities, both the fluid forces and the tether properties need to be modeled accurately.

National Category
Other Physics Topics
Identifiers
urn:nbn:se:umu:diva-106875 (URN)10.1088/1478-3975/12/5/056006 (DOI)000362005900010 ()26331992 (PubMedID)
Funder
Swedish Research Council, 2013-5379Swedish Research Council, 621-2008-3280
Available from: 2015-08-11 Created: 2015-08-11 Last updated: 2017-12-04Bibliographically approved
6. Detecting the presence of surface organelles at the single cell level, a novel cell sorting approach
Open this publication in new window or tab >>Detecting the presence of surface organelles at the single cell level, a novel cell sorting approach
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(English)Manuscript (preprint) (Other academic)
Keyword
Optical tweezers, drag force, E. coli, pili, fimbriae
National Category
Other Physics Topics
Identifiers
urn:nbn:se:umu:diva-109521 (URN)
Available from: 2015-09-30 Created: 2015-09-30 Last updated: 2015-09-30
7. Cell shape identification using digital holographic microscopy
Open this publication in new window or tab >>Cell shape identification using digital holographic microscopy
2015 (English)In: Applied Optics, ISSN 1559-128X, E-ISSN 2155-3165, Vol. 54, no 24, 7442-7448 p.Article in journal (Refereed) Published
Abstract [en]

We present a cost-effective, simple and fast digital holographic microscopy method based upon Rayleigh-Sommerfeld back propagation for identification of the geometrical shape of a cell. The method was tested using synthetic hologram images generated by ray-tracing software and from experimental images of semi-transparent spherical beads and living red blood cells. Our results show that by only using the real part of the back-reconstructed amplitude the proposed method can provide information of the geometrical shape of the object and at the same time accurately determine the axial position of the object under study. The proposed method can be used in flow chamber assays for pathophysiological studies where fast morphological changes of cells are studied in high numbers and at different heights.

Place, publisher, year, edition, pages
Optical Society of America, 2015
Keyword
Microscopy, Digital holography, Optical tweezers or optical manipulation
National Category
Atom and Molecular Physics and Optics
Identifiers
urn:nbn:se:umu:diva-106588 (URN)10.1364/AO.54.007442 (DOI)000360190200039 ()26368783 (PubMedID)
Available from: 2015-07-22 Created: 2015-07-22 Last updated: 2017-12-04Bibliographically approved

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