Synchrotron X-ray diffraction and fluorescence reveals more about the nature of biofilms

 

Image by Kaspar Kallip - Own work, CC BY-SA 4.0, https://commons.wikimedia.org/w/index.php?curid=45334128

Biofilms are multicellular microbial communities made of bacterial cells surrounded by a network of secreted biopolymers. Biofilms enable bacteria to attach on surfaces and the extracellular matrix barrier structurally isolates bacterial cells from external environmental fluctuations. For instance, biofilms can confer bacterial cell protection against mechanical stress and toxins like antimicrobials and disinfectants.

 

While the bacterial cells are genetically identical, their different phenotypes within biofilms allow the communities to respond differently to nutrients and oxygen levels, as well as threats.

 

Spatiotemporal gene expression analyses are the most common methods used to understand biofilm-associated phenotypic heterogeneity at the molecular level. Spatiotemporal gene expression is the activation of genes within specific tissues of an organism at specific times during development. Gene activation patterns vary widely in complexity.

 

Despite the method, it remains unknown how the elemental and structural properties of biofilms differ across space and time because most studies digest biofilms before analyzing the isolated molecules for their quantities.

 

New methods can reveal the importance of the physicochemical properties of the molecules secreted by the cells in a biofilm. This leads to the differential distribution of nutrients that propagate through macroscopic length scales and hence different biofilm properties.

 

An alternative technique is X-ray diffraction. This method can help to determine the structures of molecules present in a sample using electron densities of diffracted X-rays. A second method is X-ray fluorescence, which can be used to detect emissions of characteristic fluorescent X-rays from a material that is bombarded with high energy X-rays for elemental analysis.

 

Such technologies have shown how TasA fibers can greatly influence how stable the biofilms are and explain for differences in structural stability between old and young biofilms. TasA proteins can form fibers into three different morphologies: aggregate in acidic conditions, form thin and long fibrils at high salt concentration, and establish fiber bundles at high protein and salt concentrations. Thin and long fibrils were most common in young biofilms (1-2-day-old), showing that as biofilm ages, the proportion of TasA fibers could change a biofilm’s mechanical integrity.

 

Researchers have made use of both methods to scan intact Bacillus subtilis biofilm samples of different ages. This revealed spatiotemporal division of four main components including water, spores, extracellular matrix, and metal ion. The data shows how these components interact to affect processes like sporulation and polymer fibril formation.

 

One observation made is where water is bound in biofilm wrinkles and the role of wrinkles acting as water channels for nutrient transport and waste removal. Biofilm wrinkles could serve as channels filled with water to transfer nutrients across biofilms, and the water in wrinkles is not freely floating.

 

Importantly, bacterial spores can survive long periods of dehydration without detectable damage, and the presence of water can trigger germination. In terms of signalling when germination may be possible, spore coat protein swelling in biofilms may serve as humidity sensors for spore germination.

 

Posted by Dr. Tim Sandle, Pharmaceutical Microbiology Resources (http://www.pharmamicroresources.com/)

Understanding biofilms: The life and death of an 'altruistic' bacterium

Unknown author. Source: http://ober-proteomics.pnnl.gov/bioapps/organisms.stm

Public Domain: https://commons.wikimedia.org/w/index.php?curid=39355477   

 

A new study from the University of Montreal shows how some bacteria living in a biofilm sacrifice themselves to ensure the survival of the community.

   

Living in a biofilm provides numerous advantages to bacteria: things like resource sharing, shelter from predators, and increased resistance to toxic compounds such as antibiotics. Yet the option to leave the biofilm when environmental conditions deteriorate can also be of benefit to bacteria, allowing them to relocate to a more hospitable environment.

 

 

For the bacterium Caulobacter crescentus, the biofilm becomes a kind of prison in perpetuity: once cells are attached to a surface through a strong adhesive at one end of the cell, they cannot leave the biofilm. However, when these attached cells divide, their unattached 'daughter' cells have a choice of joining the biofilm or moving away.

 

Caulobacter crescentus is a Gram-negative, oligotrophic bacterium widely distributed in fresh water lakes and streams. The bacterium has a characteristic crescent shape. Caulobacter was the first asymmetric bacterium shown to age. Reproductive senescence was measured as the decline in the number of progenies produced over time.

 

What is of interest to microbiologists is with how cells decide to stay or leave the biofilm. When Caulobacter cells die in the biofilm, they release their DNA, which inhibits daughter cells from joining the biofilm, hence promoting relocation from environments where death rate increases.

 

The researchers have set out to determine if cell death occurred randomly as the environmental quality declined or if it was a regulated process responding to a specific signal. This reveals that Caulobacter uses a programmed cell death mechanism that causes some cells to sacrifice themselves when the conditions inside the biofilm deteriorate. This is known as a toxin-antitoxin system.

 

The mechanism uses a toxin that targets a vital function and its associated antidote, the antitoxin. The toxin is more stable than the antitoxin and when programmed cell death is initiated, the amount of antitoxin is reduced, resulting in cell death. The toxin-antitoxin system is activated when oxygen becomes sparse as the biofilm becomes larger and cells compete for the available oxygen.

 

The resulting death of a subset of cells releases DNA, which promotes the dispersal of new cells to potentially more hospitable environments, thereby preventing overcrowding that would further reduce environmental quality in the biofilm.

 

The reference is:

 

Cecile Berne, Sebastien Zappa, Yves V Brun. eDNA-stimulated cell dispersion from Caulobacter crescentus biofilms upon oxygen limitation is dependent on a toxin-antitoxin system. eLife, 2022; 11 DOI: 10.7554/eLife.80808

Posted by Dr. Tim Sandle, Pharmaceutical Microbiology Resources (http://www.pharmamicroresources.com/)