Nanomechanical Characterization of Bacillus anthracis Spores by Atomic Force Microscopy

ABSTRACTThe study of structures and properties of bacterial spores is important to understanding spore formation and biological responses to environmental stresses. While significant progress has been made over the years in elucidating the multilayer architecture of spores, the mechanical properties of the spore interior are not known. Here, we present a thermal atomic force microscopy (AFM) study of the nanomechanical properties of internal structures ofBacillus anthracisspores. We developed a nanosurgical sectioning method in which a stiff diamond AFM tip was used to cut an individual spore, exposing its internal structure, and a soft AFM tip was used to image and characterize the spore interior on the nanometer scale. We observed that the elastic modulus and adhesion force, including their thermal responses at elevated temperatures, varied significantly in different regions of the spore section. Our AFM images indicated that the peptidoglycan (PG) cortex ofBacillus anthracisspores consisted of rod-like nanometer-sized structures that are oriented in the direction perpendicular to the spore surface. Our findings may shed light on the spore architecture and properties.IMPORTANCEA nanosurgical AFM method was developed that can be used to probe the structure and properties of the spore interior. The previously unknown ultrastructure of the PG cortex ofBacillus anthracisspores was observed to consist of nanometer-sized rod-like structures that are oriented in the direction perpendicular to the spore surface. The variations in the nanomechanical properties of the spore section were largely correlated with its chemical composition. Different components of the spore materials showed different thermal responses at elevated temperatures.

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The analysis of nanomechanical properties of Candida spp. by atomic force microscopy (AFM) method

Postępy Higieny i Medycyny Doświadczalnej ◽

10.5604/01.3001.0013.3449 ◽

2019 ◽

Vol 73 ◽

pp. 353-358

Author(s):

Małgorzata Tokarska-Rodak ◽

Sławomir Czernik ◽

Marta Chwedczuk ◽

Dorota Plewik ◽

Tomasz Grudniewski ◽

...

Keyword(s):

Atomic Force Microscopy ◽

Candida Albicans ◽

Adhesion Force ◽

Clinical Strain ◽

Candida Spp ◽

Clinical Strains ◽

Force Microscopy ◽

Nanomechanical Properties ◽

Atomic Force ◽

Morphological Differences

The aim of the study was to analyze the selected nanomechanical properties of Candida spp: Candida albicans (standard strain ATCC 10231), Candida albicans (clinical strain, cultured from an oral swab), Candida lipolytica (clinical strain, cultured from a nosal swab) in atomic force microscopy (AFM). The culture Candida spp. was performed of Tryptone Soya Broth (BioMaxima). The topography and sample properties were analysed in AFM (Ntegra Spectra C from NT) and the results were carried out using NOVA 1.1.0.1824 software. C. albicans ATCC 10231 cells were significantly higher 1.81 μm (p = 0.001) from clinical strains: C. albicans (1.30 μm) and C. lipolytica (1.23 μm). C. albicans ATCC 10231 cells, and C. albicans cells of the clinical strain were softer, especially in the top parts of cells, than C. lipolytica cells. Adhesion force measured for C. albicans ATCC 10231 was 62.83 nN, and was significantly higher compared to the values obtained for C. albicans (41.93 nN, p = 0.0002 ) and C. lipolytica (41.78 nN, p = 0.0002 ). The stiffness of the Candida spp. cell surface was comparable and was in the range of 5–6 nA. The differences in height may result from different conditions in which clinical strains grow. Adhesion force can be helpful in the analysis of the degree of destruction of the cell wall by various substances. The conducted analyses showed morphological differences and the differences in mechanical properties of the researched Candida spp. This data may be important in assessing their susceptibility to the effects of various substances of a lytic nature.

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Quantifying the Nanomechanical Properties of Bacillus Anthracis and Implications for Spore Killing

ASME 2010 First Global Congress on NanoEngineering for Medicine and Biology ◽

10.1115/nemb2010-13032 ◽

2010 ◽

Author(s):

Terri A. Camesano ◽

Paola A. Pinzo´n-Arango ◽

Ramanathan Nagarajan

Keyword(s):

Elastic Modulus ◽

Bacillus Anthracis ◽

Hertz Model ◽

Force Microscopy ◽

Nanomechanical Properties ◽

Atomic Force ◽

Spore Killing ◽

Before And After ◽

Bacillus Anthracis Spores ◽

Tensile Elastic Modulus

Atomic force microscopy (AFM) was used to study the changes in the nanomechanical properties of the surface of Bacillus anthracis spores during germination. The Hertz model of continuum mechanics of contact was used to evaluate the Young’s or tensile elastic modulus of the spores before and after germination by applying the model to load-indentation curves obtained during force cycles. The highest elastic modulus was obtained with dormant spores, with average elasticity values of 197 ± 81 MPa. Fully vegetative spores had the lowest elasticity values. The elasticity decreased when spores were incubated with either L-alanine or inosine, and the decrease was greatest when both of these germinants were used in combination. We also found that as the spore elasticity values increasd, the indentation depth of the AFM probe into the surface increased, with vegetative B. anthracis cells having elasticity depths of up to 246.2 nm. We have shown that spore nanomechanical properties change during germination, and depend on the type of germinant that is used. Weakening of the spore cell wall may help explain why germinating cells are much more susceptible to anti-sporal agents. The study of elasticity of spores may be a valuable tool in the development of antibiotics or anti-sporal treatments.

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In Situ Imaging of Candida albicans Hyphal Growth via Atomic Force Microscopy

mSphere ◽

10.1128/msphere.00946-20 ◽

2020 ◽

Vol 5 (6) ◽

Author(s):

Arzu Çolak ◽

Mélanie A. C. Ikeh ◽

Clarissa J. Nobile ◽

Mehmet Z. Baykara

Keyword(s):

Atomic Force Microscopy ◽

Candida Albicans ◽

Adhesion Force ◽

Hyphal Growth ◽

Antifungal Drugs ◽

Content Type ◽

Force Microscopy ◽

Atomic Force ◽

Host Tissues

ABSTRACT Candida albicans is an opportunistic fungal pathogen of humans known for its ability to cause a wide range of infections. One major virulence factor of C. albicans is its ability to form hyphae that can invade host tissues and cause disseminated infections. Here, we introduce a method based on atomic force microscopy to investigate C. albicans hyphae in situ on silicone elastomer substrates, focusing on the effects of temperature and antifungal drugs. Hyphal growth rates differ significantly for measurements performed at different physiologically relevant temperatures. Furthermore, it is found that fluconazole is more effective than caspofungin in suppressing hyphal growth. We also investigate the effects of antifungal drugs on the mechanical properties of hyphal cells. An increase in Young’s modulus and a decrease in adhesion force are observed in hyphal cells subjected to caspofungin treatment. Young’s moduli are not significantly affected following treatment with fluconazole; the adhesion force, however, increases. Overall, our results provide a direct means of observing the effects of environmental factors and antifungal drugs on C. albicans hyphal growth and mechanics with high spatial resolution. IMPORTANCE Candida albicans is one of the most common pathogens of humans. One important virulence factor of C. albicans is its ability to form elongated hyphae that can invade host tissues and cause disseminated infections. Here, we show the effect of different physiologically relevant temperatures and common antifungal drugs on the growth and mechanical properties of C. albicans hyphae using atomic force microscopy. We demonstrate that minor temperature fluctuations within the normal range can have profound effects on hyphal cell growth and that different antifungal drugs impact hyphal cell stiffness and adhesion in different ways.

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