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KNOW YOUR ENEMY: Characterizing Pathogenic Biomaterials Using Laser Tweezers
Umeå University, Faculty of Science and Technology, Department of Physics.
2022 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Diseases caused by pathogenic agents such as bacteria and viruses result in devastating costs on personal and societal levels. However, it is not just the emergence of new diseases that is problematic. Antibiotic resistance among bacteria makes uncomplicated infections difficult and lethal. Resilient disease-causing spores spread in hospitals, the food industry, and water supplies requiring effective detection and disinfection methods. Further, we face complex neurological diseases where no effective treatment or diagnostic methods exist. Thus, we must increase our fundamental understanding of these diseases to develop effective diagnostic, detection, disinfection, and treatment methods.

Classically, the methods used for detecting and studying the underlying mechanics of pathogenic agents work on a large scale, measuring the average macroscopic behavior and properties of these pathogens. However, just as with humans, the average behavior is not always representative of individual behavior. Therefore, it is also essential to investigate the characteristics of these pathogens on a single cell or particle level. 

This thesis develops and applies optical techniques to characterize pathogenic biomaterial on a single cell or particle level. At the heart of all these studies is our Optical Tweezers (OT) instrument. OT are a tool that allows us to reach into the microscopic world and interact with it. Finally, by combining OT with other experimental techniques, we can chemically characterize biomaterials and develop assays that mimic different biological settings. Using these tools, we investigate bacterial adhesion, disinfection, and detection of pathogenic spores and proteins.

Hopefully, the insights of these studies can lessen the burden on society caused by diseases by helping others develop effective treatment, diagnostic, detection, and disinfection methods in the future. 

Place, publisher, year, edition, pages
Umeå: Umeå universitet , 2022. , p. 73
Keywords [en]
Optical Tweezers, Laser Tweezers, Raman Spectroscopy, Bacterial Adhesion, Biophysics, Pili, Bacterial Spores, Endospores, Oocysts, Cryptosporidium, Optics
National Category
Biophysics Atom and Molecular Physics and Optics
Research subject
biology; Physics
Identifiers
URN: urn:nbn:se:umu:diva-192471ISBN: 978-91-7855-726-4 (print)ISBN: 978-91-7855-727-1 (electronic)OAI: oai:DiVA.org:umu-192471DiVA, id: diva2:1637609
Public defence
2022-03-11, NAT.D.410, Naturvetarhuset, Umeå, 09:00 (English)
Opponent
Supervisors
Available from: 2022-02-18 Created: 2022-02-14 Last updated: 2025-02-20Bibliographically approved
List of papers
1. Cooke-Triplet-Tweezers: More compact, robust and efficient optical tweezers
Open this publication in new window or tab >>Cooke-Triplet-Tweezers: More compact, robust and efficient optical tweezers
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2018 (English)In: Optics Letters, ISSN 0146-9592, E-ISSN 1539-4794, Vol. 43, no 9, p. 1990-1993Article in journal (Refereed) Published
Abstract [en]

We present a versatile three-lens optical design to improve the overall compactness, efficiency, and robustness for optical tweezers based applications. The design, inspired by the Cooke–Triplet configuration, allows for continuous beam magnifications of 2–10× , and axial as well as lateral focal shifts can be realized without switching lenses or introducing optical aberrations. We quantify the beam quality and trapping stiffness and compare the Cooke–Triplet design with the commonly used double Kepler design through simulations and direct experiments. Optical trapping of 1 and 2 μm beads shows that the Cooke–Triplet possesses an equally strong optical trap stiffness compared to the double Kepler lens design but reduces its lens system length by a factor of 2.6. Finally, we demonstrate how a Twyman–Green interferometer integrated in the Cooke–Triplet optical tweezers setup provides a fast and simple method to characterize the wavefront aberrations in the lens system and how it can help in aligning the optical components perfectly.

National Category
Atom and Molecular Physics and Optics
Identifiers
urn:nbn:se:umu:diva-145899 (URN)10.1364/OL.43.001990 (DOI)000431179400013 ()29714728 (PubMedID)2-s2.0-85046663259 (Scopus ID)
Available from: 2018-03-21 Created: 2018-03-21 Last updated: 2023-09-07Bibliographically approved
2. Step-by-Step Guide to 3D Print Motorized Rotation Mounts for Optical Applications
Open this publication in new window or tab >>Step-by-Step Guide to 3D Print Motorized Rotation Mounts for Optical Applications
2021 (English)In: Applied Optics, ISSN 0003-6935, E-ISSN 1539-4522, Vol. 60, no 13, p. 3764-3771Article in journal (Refereed) Published
Abstract [en]

Motorized rotation mounts and stages are versatile instruments that introduce computer control to optical systems, enabling automation and scanning actions. They can be used for intensity control and position adjustments, etc. However, these rotation mounts come with a hefty price tag, and this limits their use. This work shows how to build two different types of motorized rotation mounts for 1" optics, using a 3D printer and off-the-shelf components. The first is intended for reflective elements, like mirrors and gratings, and the second for transmissive elements, like polarizers and retarders. We evaluate and compare their performance to commercial systems based on velocity, resolution, precision, backlash, and axis wobble. Also, we investigate the angular stability using Allan variance analysis. The results show that our mounts perform similar to systems costing more than 2000 Euro, while also being quick to build and costing less than 200 Euro. As a proof of concept, we show how to control lasers used in an optical tweezers and Raman spectroscopy setup. When used for this, the 3D printed motorized rotational mounts provide intensity control with a resolution of 0.03 percentage points or better.

Place, publisher, year, edition, pages
Optical Society of America, 2021
National Category
Other Physics Topics Atom and Molecular Physics and Optics
Research subject
Physics
Identifiers
urn:nbn:se:umu:diva-181868 (URN)10.1364/AO.422695 (DOI)000645865100027 ()2-s2.0-85105102116 (Scopus ID)
Available from: 2021-03-30 Created: 2021-03-30 Last updated: 2023-09-05Bibliographically approved
3. Optical design for laser tweezers Raman spectroscopy setups for increased sensitivity and flexible spatial detection
Open this publication in new window or tab >>Optical design for laser tweezers Raman spectroscopy setups for increased sensitivity and flexible spatial detection
2021 (English)In: Applied Optics, ISSN 1559-128X, E-ISSN 2155-3165, Vol. 60, no 16, p. 4519-4523Article in journal (Refereed) Published
Abstract [en]

We demonstrate a method to double the collection efficiency in Laser Tweezers Raman Spectroscopy (LTRS) by collecting both the forward and back-scattered light in a single-shot multitrack measurement. Our method can collect signals at different sample volumes, granting both the pinpoint spatial selectivity of confocal Raman and the bulk sensitivity of non-confocal Raman simultaneously. Further, we display that our approach allows for reduced detector integration time and laser power. To show this, we measure the Raman spectra of both polystyrene beads and bacterial spores. For spores, we can trap them at 2.5 mW laser power and acquire a high signal-to-noise ratio Power spectrum of the CaDPA peaks using an integration time of 2 x 30 seconds. Thus, our method will enable the monitoring of biological samples sensitive to high intensities for longer times. Additionally, we demonstrate that by a simple modification, we can add polarization sensitivity and retrieve extra biochemical information. 

Place, publisher, year, edition, pages
Optical Society of America, 2021
Keywords
Raman, optical tweezers
National Category
Atom and Molecular Physics and Optics Analytical Chemistry
Research subject
Physics
Identifiers
urn:nbn:se:umu:diva-182971 (URN)10.1364/AO.424595 (DOI)000658194300003 ()2-s2.0-85106550983 (Scopus ID)
Funder
Swedish Research Council, 2019-04016Swedish Foundation for Strategic Research
Available from: 2021-05-11 Created: 2021-05-11 Last updated: 2023-09-05Bibliographically approved
4. 3D printed water-soluble scaffolds for rapid production of PDMS micro-fluidic flow chambers
Open this publication in new window or tab >>3D printed water-soluble scaffolds for rapid production of PDMS micro-fluidic flow chambers
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2018 (English)In: Scientific Reports, E-ISSN 2045-2322, Vol. 8, no 1, article id 3372Article in journal (Refereed) Published
Abstract [en]

We report a novel method for fabrication of three-dimensional (3D) biocompatible micro-fluidic flow chambers in polydimethylsiloxane (PDMS) by 3D-printing water-soluble polyvinyl alcohol (PVA) filaments as master scaffolds. The scaffolds are first embedded in the PDMS and later residue-free dissolved in water leaving an inscription of the scaffolds in the hardened PDMS. We demonstrate the strength of our method using a regular, cheap 3D printer, and evaluate the inscription process and the channels micro-fluidic properties using image analysis and digital holographic microscopy. Furthermore, we provide a protocol that allows for direct printing on coverslips and we show that flow chambers with a channel cross section down to 40 x 300 μm can be realized within 60 min. These flow channels are perfectly transparent, biocompatible and can be used for microscopic applications without further treatment. Our proposed protocols facilitate an easy, fast and adaptable production of micro-fluidic channel designs that are cost-effective, do not require specialized training and can be used for a variety of cell and bacterial assays. To help readers reproduce our micro-fluidic devices, we provide: full preparation protocols, 3D-printing CAD files for channel scaffolds and our custom-made molding device, 3D printer build-plate leveling instructions, and G-code.

Place, publisher, year, edition, pages
Nature Publishing Group, 2018
National Category
Other Materials Engineering Other Engineering and Technologies Other Physics Topics
Identifiers
urn:nbn:se:umu:diva-144631 (URN)10.1038/s41598-018-21638-w (DOI)000425500300044 ()2-s2.0-85042263864 (Scopus ID)
Funder
Swedish Research Council, 2013-5379The Kempe Foundations, JCK-1622
Available from: 2018-02-08 Created: 2018-02-08 Last updated: 2025-02-10Bibliographically approved
5. Step-by-step guide to reduce spatial coherence of laser light using a rotating ground glass diffuser
Open this publication in new window or tab >>Step-by-step guide to reduce spatial coherence of laser light using a rotating ground glass diffuser
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2017 (English)In: Applied Optics, ISSN 1559-128X, E-ISSN 2155-3165, Vol. 56, no 19, p. 5427-5435Article in journal (Refereed) Published
Abstract [en]

Wide field-of-view imaging of fast processes in a microscope requires high light intensities motivating the use of lasers as light sources. However, due to their long spatial coherence length, lasers are inappropriate for such applications, as they produce coherent noise and parasitic reflections, such as speckle, degrading image quality. Therefore, we provide a step-by-step guide for constructing a speckle-free and high-contrast laser illumination setup using a rotating ground glass diffuser driven by a stepper motor. The setup is easy to build, cheap, and allows a significant light throughput of 48%, which is 40% higher in comparison to a single lens collector commonly used in reported setups. This is achieved by using only one objective to collect the scattered light from the ground glass diffuser. We validate our setup in terms of image quality, speckle contrast, motor-induced vibrations, and light throughput. To highlight the latter, we record Brownian motion of micro-particles using a 100x oil immersion objective and a high-speed camera operating at 2000 Hz with a laser output power of only 22 mW. Moreover, by reducing the objective magnification to 50x, sampling rates up to 10,000 Hz are realized. To help readers with basic or advanced optics knowledge realize this setup, we provide a full component list, 3D-printing CAD files, setup protocol, and the code for running the stepper motor.

Place, publisher, year, edition, pages
Optical Society of America, 2017
National Category
Atom and Molecular Physics and Optics
Identifiers
urn:nbn:se:umu:diva-135625 (URN)10.1364/AO.56.005427 (DOI)000404745800041 ()2-s2.0-85021648389 (Scopus ID)
Available from: 2017-06-01 Created: 2017-06-01 Last updated: 2024-07-02Bibliographically approved
6. Tunable two-dimensional patterning of a semiconducting Nanometer-Thin C60 fullerene film using a spatial light modulator
Open this publication in new window or tab >>Tunable two-dimensional patterning of a semiconducting Nanometer-Thin C60 fullerene film using a spatial light modulator
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2020 (English)In: ACS Applied Nano Materials, ISSN 2574-0970, Vol. 3, no 6, p. 2574-0970Article in journal (Other academic) Published
Abstract [en]

The photochemical coupling of fullerene molecules into covalently connected oligomeric or polymeric structures can result in drastically lowered solubility in common solvents with retained semiconductor properties. Here, we exploit this combination of properties for the utilization of fullerenes as a negative photoresist material with electronic functionality. Specifically, we develop an easily tunable exposure system, essentially comprising a laser and a computer-controlled spatial light modulator (SLM) featuring >8 million independently controlled pixels, for the spatially selective photochemical transformation of nanometer-thin C60 fullerene films. With a carefully designed laser-SLM-exposure/solvent-development cycle, we are able to realize well-resolved two-dimensional hexagonal or square patterns of circular C60 microdots with a center-to-center distance of 1–5 μm and a maximum thickness of 20–35 nm over several square-millimeter-sized areas on a substrate. The functionality of such a hexagonal C60 pattern was demonstrated by its inclusion in between the transparent electrode and the active material in a light-emitting electrochemical cell, which featured an enhanced light output by >50% in comparison to a reference device void of the patterned C60 layer.

Place, publisher, year, edition, pages
Acoustical Society of America (ASA), 2020
Keywords
C60, fullerenes, tunable and high-resolution 2D patterning, spatial light modulator, negative photoresist, light outcoupling, light-emitting electrochemical cell
National Category
Other Physics Topics
Identifiers
urn:nbn:se:umu:diva-166406 (URN)10.1021/acsanm.0c00793 (DOI)000545689000055 ()2-s2.0-85087440615 (Scopus ID)
Funder
Swedish Foundation for Strategic Research Swedish Research CouncilSwedish Energy AgencyBertil & Britt Svenssons Stiftelse för Belysningsteknik
Note

Previously included in thesis in manuscript form.

Available from: 2019-12-16 Created: 2019-12-16 Last updated: 2023-03-23Bibliographically approved
7. Unveiling molecular interactions that stabilize the chaperone-usher pili rod and their role for mechanical and kinetic properties
Open this publication in new window or tab >>Unveiling molecular interactions that stabilize the chaperone-usher pili rod and their role for mechanical and kinetic properties
(English)Manuscript (preprint) (Other academic)
National Category
Biophysics Other Physics Topics
Identifiers
urn:nbn:se:umu:diva-192463 (URN)
Available from: 2022-02-14 Created: 2022-02-14 Last updated: 2025-02-20
8. Impact of an alpha helix and a cysteine-cysteine disulfide bond on the resistance of bacterial adhesion pili to stress
Open this publication in new window or tab >>Impact of an alpha helix and a cysteine-cysteine disulfide bond on the resistance of bacterial adhesion pili to stress
2021 (English)In: Proceedings of the National Academy of Sciences of the United States of America, ISSN 0027-8424, E-ISSN 1091-6490, Vol. 118, no 21, article id e2023595118Article in journal (Refereed) Published
Abstract [en]

Escherichia coli express adhesion pili that mediate attachment to host cell surfaces and are exposed to body fluids in the urinary and gastrointestinal tracts. Pilin subunits are organized into helical polymers, with a tip adhesin for specific host binding. Pili can elastically unwind when exposed to fluid flow forces, reducing the adhesin load, thereby facilitating sustained attachment. Here we investigate biophysical and structural differences of pili commonly expressed on bacteria that inhabit the urinary and intestinal tracts. Optical tweezers measurements reveal that Class 1a pili of uropathogenic E. coli (UPEC), as well as Class 1b of enterotoxigenic E. coli (ETEC), undergo an additional conformational change beyond pilus unwinding, providing significantly more elasticity to their structure than ETEC Class 5 pili. Examining structural and steered molecular dynamics simulation data, we find this difference in Class 1 pili subunit behavior originates from an alpha-helical motif that can unfold when exposed to force. A disulfide bond cross-linking beta-strands in Class 1 pili stabilizes subunits, allowing them to tolerate higher forces than Class 5 pili that lack this covalent bond. We suggest that these extra contributions to pilus resiliency are relevant for the UPEC niche since resident bacteria are exposed to stronger, more transient drag forces compared to those experienced by ETEC bacteria in the mucosa of the intestinal tract. Interestingly, Class 1b ETEC pili include the same structural features seen in UPEC pili, while requiring lower unwinding forces that are more similar to those of Class 5 ETEC pili.

National Category
Other Physics Topics Biophysics
Identifiers
urn:nbn:se:umu:diva-181870 (URN)10.1073/pnas.2023595118 (DOI)000659436300016 ()2-s2.0-85106364234 (Scopus ID)
Funder
Swedish Research CouncilThe Kempe Foundations
Available from: 2021-03-30 Created: 2021-03-30 Last updated: 2025-02-20Bibliographically approved
9. Archaic chaperone-usher pilus self-secretes into a superelastic zigzag spring architecture
Open this publication in new window or tab >>Archaic chaperone-usher pilus self-secretes into a superelastic zigzag spring architecture
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(English)Manuscript (preprint) (Other academic)
National Category
Biophysics Biochemistry Molecular Biology Other Physics Topics
Identifiers
urn:nbn:se:umu:diva-192464 (URN)10.21203/rs.3.rs-936177/v1 (DOI)
Available from: 2022-02-14 Created: 2022-02-14 Last updated: 2025-02-20
10. pH induced changes in Raman, UV-Vis absorbance, and fluorescence spectra of dipicolinic acid (DPA)
Open this publication in new window or tab >>pH induced changes in Raman, UV-Vis absorbance, and fluorescence spectra of dipicolinic acid (DPA)
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2022 (English)In: Spectrochimica Acta Part A - Molecular and Biomolecular Spectroscopy, ISSN 1386-1425, E-ISSN 1873-3557, Vol. 271, article id 120869Article in journal (Refereed) Published
Abstract [en]

Dipicolinic acid (DPA) is an essential component for the protection of DNA in bacterial endospores and is often used as a biomarker for spore detection. Depending upon the pH of the solution, DPA exists in different ionic forms. Therefore, it is important to understand how these ionic forms influence spectroscopic response. In this work, we characterize Raman and absorption spectra of DPA in a pH range of 2.0–10.5. We show that the ring breathing mode Raman peak of DPA shifts from 1003 cm−1 to 1017 cm−1 and then to 1000 cm−1 as pH increases from 2 to 5. The relative peak intensities related to the different ionic forms of DPA are used to experimentally derive the pKa values (2.3 and 4.8). We observe using UV–vis spectroscopy that the changes in the absorption spectrum of DPA as a function of pH correlate with the changes observed in Raman spectroscopy, and the same pKa values are verified. Lastly, using fluorescence spectroscopy and exciting a DPA solution at between 210–330 nm, we observe a shift in fluorescence emission from 375 nm to 425 nm between pH 2 and pH 6 when exciting at 320 nm. Our work shows that the different spectral responses from the three ionic forms of DPA may have to be taken into account in, e.g., spectral analysis and for detection applications.

Place, publisher, year, edition, pages
Elsevier, 2022
Keywords
Bacterial spores, DPA, Biomarker, Raman spectra, UV–vis absorption spectra, Fluorescence spectra
National Category
Biophysics Other Physics Topics Inorganic Chemistry
Identifiers
urn:nbn:se:umu:diva-191504 (URN)10.1016/j.saa.2022.120869 (DOI)000751812400019 ()35065519 (PubMedID)2-s2.0-85122995846 (Scopus ID)
Funder
Swedish Research Council, 2019–04016The Kempe Foundations, JCK-1916.2
Available from: 2022-01-19 Created: 2022-01-19 Last updated: 2025-09-30Bibliographically approved
11. Biophysical Fingerprinting of Single Bacterial Spores using Laser Raman Optical Tweezers
Open this publication in new window or tab >>Biophysical Fingerprinting of Single Bacterial Spores using Laser Raman Optical Tweezers
2020 (English)In: Proceedings Volume 11416, Chemical, Biological, Radiological, Nuclear, and Explosives (CBRNE) Sensing XXI, 2020, SPIE - International Society for Optical Engineering, 2020, article id 1141601Conference paper, Published paper (Refereed)
Abstract [en]

Spore-forming bacteria that cause diseases pose a danger in our society. When in spore form, bacteria can survive high temperatures and resist a plethora of disinfection chemicals. Effective disinfection approaches are thus critical. Since a population of bacterial spores is heterogeneous in many aspects, single spore analyzing methods are suitable when heterogeneous information cannot be neglected. We present in this work a highresolution Laser Raman optical tweezers that can trap single spores and characterize their Raman spectra. We first evaluate our system by measuring Raman spectra of spores, and purified DNA and DPA. Thereafter, we expose Bacillus thuringiensis spores to peracetic acid, chlorine dioxide, and sodium hypochlorite, which are common disinfection chemicals. The data reveals how these agents change the constitutes of a spore over time, thus improving on the mode of action of these disinfection chemicals.

Place, publisher, year, edition, pages
SPIE - International Society for Optical Engineering, 2020
National Category
Atom and Molecular Physics and Optics Biophysics
Identifiers
urn:nbn:se:umu:diva-170121 (URN)10.1117/12.2558102 (DOI)000590002900014 ()2-s2.0-85089074190 (Scopus ID)
Conference
SPIE Defense + Commercial Sensing, 2020, Online Only, California, United States, April 27, 2020.
Funder
Swedish Research Council, 2019-04016The Kempe Foundations, JCK-1916.2
Available from: 2020-04-27 Created: 2020-04-27 Last updated: 2025-02-20Bibliographically approved
12. Mode of action of Disinfection chemicals  on the bacterial spore structure and their Raman spectra
Open this publication in new window or tab >>Mode of action of Disinfection chemicals  on the bacterial spore structure and their Raman spectra
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2021 (English)In: Analytical Chemistry, ISSN 0003-2700, E-ISSN 1520-6882, Vol. 93, no 6, p. 3146-3153Article in journal (Refereed) Published
Abstract [en]

Contamination of toxic spore-forming bacteria is problematic since spores can survive a plethora of disinfection chemicals and it is hard to rapidly detect if the disinfection chemical has inactivated the spores. Thus, robust decontamination strategies and reliable detection methods to identify dead from viable spores are critical. In this work, we investigate the chemical changes of Bacillus thuringiensis spores treated with sporicidal agents such as chlorine dioxide, peracetic acid, and sodium hypochlorite using laser tweezers Raman spectroscopy. We also image treated spores using SEM and TEM to verify if we can correlate structural changes in the spores with changes to their Raman spectra. We found that over 30 min, chlorine dioxide did not change the Raman spectrum or the spore structure, peracetic acid showed a time-dependent decrease in the characteristic DNA/DPA peaks and ∼20% of the spores were degraded and collapsed, and spores treated with sodium hypochlorite showed an abrupt drop in DNA and DPA peaks within 20 min and some structural damage to the exosporium. Structural changes appeared in spores after 10 min, compared to the inactivation time of the spores, which is less than a minute. We conclude that vibrational spectroscopy provides powerful means to detect changes in spores but it might be problematic to identify if spores are live or dead after a decontamination procedure.

Place, publisher, year, edition, pages
American Chemical Society (ACS), 2021
National Category
Other Physics Topics Biophysics
Identifiers
urn:nbn:se:umu:diva-179119 (URN)10.1021/acs.analchem.0c04519 (DOI)000620922300011 ()2-s2.0-85100614040 (Scopus ID)
Funder
Swedish Research Council, 2019-04016The Kempe Foundations, (JCK-1916.2
Available from: 2021-01-26 Created: 2021-01-26 Last updated: 2025-02-20Bibliographically approved
13. Laser induced degradation of bacterial spores during micro-Raman spectroscopy
Open this publication in new window or tab >>Laser induced degradation of bacterial spores during micro-Raman spectroscopy
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2022 (English)In: Spectrochimica Acta Part A - Molecular and Biomolecular Spectroscopy, ISSN 1386-1425, E-ISSN 1873-3557, Vol. 265, article id 120381Article in journal (Refereed) Published
Abstract [en]

Micro-Raman spectroscopy combined with optical tweezers is a powerful method to analyze how the biochemical composition and molecular structures of individual biological objects change with time. In this work we investigate laser induced effects in the trapped object. Bacillus thuringiensis spores, which are robust organisms known for their resilience to light, heat, and chemicals are used for this study. We trap spores and monitor the Raman peak from CaDPA (calcium dipicolinic acid), which is a chemical protecting the spore core. We see a correlation between the amount of laser power used in the trap and the release of CaDPA from the spore. At a laser power of 5 mW, the CaDPA from spores in water suspension remain intact over the 90 min experiment, however, at higher laser powers an induced effect could be observed. SEM images of laser exposed spores (after loss of CaDPA Raman peak was confirmed) show a notable alteration of the spores' structure. Our Raman data indicates that the median dose exposure to lose the CaDPA peak was ∼60 J at 808 nm. For decontaminated/deactivated spores, i.e., treated in sodium hypochlorite or peracetic acid solutions, the sensitivity on laser power is even more pronounced and different behavior could be observed on spores treated by the two chemicals. Importantly, the observed effect is most likely photochemical since the increase of the spore temperature is in the order of 0.1 K as suggested by our numerical multiphysics model. Our results show that care must be taken when using micro-Raman spectroscopy on biological objects since photoinduced effects may substantially affect the results.

Place, publisher, year, edition, pages
Elsevier, 2022
Keywords
Bacteria, Decontamination, Multiphysics modelling, Optical tweezers, Raman spectroscopy, Spores
National Category
Biophysics Atom and Molecular Physics and Optics Cell Biology
Identifiers
urn:nbn:se:umu:diva-187284 (URN)10.1016/j.saa.2021.120381 (DOI)000709268600014 ()34562861 (PubMedID)2-s2.0-85115389622 (Scopus ID)
Funder
Swedish Research Council, 2019-04016The Kempe Foundations, JCK-1916.2Swedish Armed Forces, 470-A400821
Available from: 2021-09-07 Created: 2021-09-07 Last updated: 2025-09-30Bibliographically approved
14. Reference Raman Spectrum and Mapping of Cryptosporidium parvum Oocysts
Open this publication in new window or tab >>Reference Raman Spectrum and Mapping of Cryptosporidium parvum Oocysts
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2022 (English)In: Journal of Raman Spectroscopy, ISSN 0377-0486, E-ISSN 1097-4555, Vol. 53, no 7, p. 1293-1301Article in journal (Refereed) Published
Abstract [en]

Cryptosporidium parvum is a protozoan parasite and among the most infectious diarrhea-causing pathogens, leading to severe health problems for malnourished children and immunocompromised individuals. Outbreaks are common even in developed countries, originating from water or food contamination and resulting in suffering and large costs for society. Therefore, robust, fast and highly specific detection strategies of Cryptosporidium are needed. Label-free detection techniques such as Raman spectroscopy have been suggested, however high-resolution reported spectra in the literature are limited. In this work, we report reference Raman spectra at 3 cm-1 resolution for viable and inactivated Cryptosporidium oocysts of the species C. parvum, gathered at a single oocyst level using a laser tweezers Raman spectroscopy system. We furthermore provide tentative Raman peak assignments for the Cryptosporidium oocysts, along with Raman mapping of the oocysts’ heterogeneous internal structure. Finally, we compare the C. parvum Raman spectrum with other common enterotoxigenic pathogens: Escherichia coli, Vibrio cholerae, Bacillus cereus and Clostridium difficile. Our results show a significant difference between C. parvum Raman spectra and the other pathogens.

Place, publisher, year, edition, pages
John Wiley & Sons, 2022
Keywords
Cryptosporidium, laser tweezers Raman spectroscopy, Raman mapping
National Category
Biophysics Other Physics Topics Analytical Chemistry Infectious Medicine
Identifiers
urn:nbn:se:umu:diva-192462 (URN)10.1002/jrs.6361 (DOI)000787096700001 ()2-s2.0-85128826326 (Scopus ID)
Funder
Swedish Research Council, 2019-04016The Kempe Foundations, JCK-1916.2Swedish Armed Forces, A74028
Note

Originally included in thesis in manuscript form. 

Available from: 2022-02-14 Created: 2022-02-14 Last updated: 2025-09-30Bibliographically approved
15. Synthetic NAC 71-82 Peptides Designed to Produce Fibrils with Different Protofilament Interface Contacts
Open this publication in new window or tab >>Synthetic NAC 71-82 Peptides Designed to Produce Fibrils with Different Protofilament Interface Contacts
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2021 (English)In: International Journal of Molecular Sciences, ISSN 1661-6596, E-ISSN 1422-0067, Vol. 22, no 17, article id 9334Article in journal (Refereed) Published
Abstract [en]

Alpha-synucleinopathies are featured by fibrillar inclusions in brain cells. Although α-synuclein fibrils display structural diversity, the origin of this diversity is not fully understood. We used molecular dynamics simulations to design synthetic peptides, based on the NAC 71-82 amino acid fragment of α-synuclein, that govern protofilament contacts and generation of twisted fibrillar polymorphs. Four peptides with structures based on either single or double fragments and capped or non-capped ends were selected for further analysis. We determined the fibrillar yield and the structures from these peptides found in the solution after fibrillisation using protein concentration determination assay and circular dichroism spectroscopy. In addition, we characterised secondary structures formed by individual fibrillar complexes using laser-tweezers Raman spectroscopy. Results suggest less mature fibrils, based on the lower relative β-sheet content for double- than single-fragment peptide fibrils. We confirmed this structural difference by TEM analysis which revealed, in addition to short protofibrils, more elongated, twisted and rod-like fibril structures in non-capped and capped double-fragment peptide systems, respectively. Finally, time-correlated single-photon counting demonstrated a difference in the Thioflavin T fluorescence lifetime profiles upon fibril binding. It could be proposed that this difference originated from morphological differences in the fibril samples. Altogether, these results highlight the potential of using peptide models for the generation of fibrils that share morphological features relevant for disease, e.g., twisted and rod-like polymorphs.

Place, publisher, year, edition, pages
MDPI, 2021
Keywords
α-synuclein, NAC 71-82 peptides, fibril polymorphs
National Category
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urn:nbn:se:umu:diva-186977 (URN)10.3390/ijms22179334 (DOI)000694357900001 ()2-s2.0-85113788315 (Scopus ID)
Available from: 2021-08-29 Created: 2021-08-29 Last updated: 2025-02-20Bibliographically approved

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Dahlberg, Tobias

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