Antimicrobial wings
Cicada wings are covered with bactericidal microsculpture
It turned out that this effect is determined not by the biological or chemical properties of the wing surface, but by its specific relief: a carpet of microscopic spikes. In February of this year, the same group of scientists put forward a hypothesis explaining how the deadly microsculpture works. Modern nanotechnology can easily reproduce it, so in the near future this principle may lead to the creation of a new class of artificial bactericidal materials.
Fig. 1. A. Song cicada Psaltoda claripennis (photo from the website www.pbase.com ). B. Cicada wing surface in a scanning electron microscope; the length of the scale ruler is 2 microns (photo from the work discussed by Ivanov et al., 2012). C. Pseudomonas aeruginosa bacteria in a scanning electron microscope (photo from the website commons.wikimedia.org ). D. Bacteria of this species that died on the surface of the cicada wing, photographed in a scanning electron microscope (photo from the work discussed by Ivanova et al., 2012)The wings of the singing cicada Psaltoda claripennis look transparent, like glass (Fig. 1A), but at high magnification it can be seen that their surface is densely studded with spikes about 200 nm high and a base diameter of 100 nm (Fig. 1B).
A group of scientists from several institutes in Australia and Spain became interested in whether such a nanorelief could prevent bacteria from sticking in the aquatic environment (protecting surfaces from microbial fouling is an urgent task in engineering and medicine). The wing was immersed in a solution containing Pseudomonas aeruginosa, a ubiquitous rod–shaped bacterium capable of causing some human diseases (Fig. 1C). Despite the uneven surface, bacteria stuck to the wings in large quantities, however, as a rule, they died within 5 minutes after contact. This process could be observed using a laser confocal microscope in the presence of fluorescent dyes, which bind living, dying and dead cells differently, causing them to glow in different colors.
When the wings were dried and examined in a scanning electron microscope, it turned out that they were plastered with empty bacterial shells pressed into a carpet of spikes (Fig. 1D). Since the examination in a scanning microscope is carried out in a vacuum and requires preliminary drying of the sample, it could be suspected that the bacteria acquire this appearance in preparation for microscopy. However, the researchers were able to register the destruction of the bacterium upon contact with the wing and its indentation into the relief directly in an aqueous medium using an atomic force microscope (see Fig. 2 for details). It is noteworthy that this also happened when the thinnest (10 nm) film of gold was sprayed onto the wings with a magnetron before the experiment. Since the sculpture of gilded wings practically did not differ from the natural one, and their chemical properties radically changed, this experiment proved that it is the sculpture of the surface that destroys bacteria.
Fig. 2. Destruction of a bacterial cell on the spines of a cicada wing, traced directly in an aqueous medium (not shown) using an atomic force microscope (AFM). The position of the probe in the form of a thin elastic rod with a nanoscale tip at the end in contact with the bacterium was recorded. The graph shows that after the initial period of smooth lowering, when the bacterium was gradually pressed into the carpet of spikes, the probe abruptly failed at 200 nm, which corresponds to the height of the spikes, i.e. the bacterium burst. Figures from the article under discussion by Ivanova et al., 2012Since the XIX century, bacteria have been divided into two groups – gram-positive and gram-negative; the first group is stained by the Gram method, and the second, respectively, is not stained, which reflects the differences in the structure and composition of the bacterial cell wall of these groups.
In their next work, the authors compared the effect of Psaltoda claripennis wings on 4 types of gram-negative and 3 types of gram-positive bacteria. It turned out that the bactericidal effect is observed only in relation to gram-negative and does not depend on the shape of bacterial cells (rods or cocci). Since the shells of most gram-positive bacteria are more durable, the authors concluded that the phenomenon is based on the interaction of the wing surface with the bacterial shell.
In their last article, the authors proposed a hypothetical explanation of the effect they discovered and supported it with calculations. Since the spikes are much smaller than the bacterium, the proposed model (Fig. 3) ignores the shape of the latter and describes the interaction of a carpet of spikes with a flat surface of the bacterium. On the other hand, since the thickness of the bacterial shell – about 10 nm – is small compared to the height of the spikes, this shell can be conditionally considered an elastic membrane. It is assumed that after the initial contact of the bacteria with the tops of the spikes, the adhesion forces (gluing) tend to increase the contact area. The spines are gradually drawn into the bacterium, its shell is deformed, and the areas of the shell in the spaces between the spines are stretched until there is a rupture. Calculations have shown that in order for the spikes in this model to simply pierce the shell (like a hedgehog balloon), their vertices should be much sharper, with a radius of rounding about 1 nm.
Fig. 3. A hypothesis explaining how the bacterial membrane is deformed and destroyed on a cicada wing covered with micro-spikes with rounded tops (the aqueous medium is not shown). The green color shows the areas of the bacterial shell stuck to the spikes, and the orange color shows the areas in the spaces between the spikes. The growth of the contact areas leads to a rupture of the shell between the spikes (lower figure). Figure from the article under discussion Pogodin et al., 2013The behavior of this system is determined by the strength of the adhesive interaction, the geometry of the spikes, as well as the strength and flexibility of the bacterial wall.
(Note that the model ignores the interaction of both surfaces with the surrounding fluid, which can be a serious disadvantage.) The flexibility of the wall depends on intracellular pressure, that is, turgor: the higher it is, the stronger the shell resists deformation. To test this idea, the authors irradiated three types of gram–positive bacteria with microwaves - normally resistant to deadly sculpture. As a result of this treatment, the bacterial shells become permeable for a while, which leads to a partial loss of turgor. Indeed, irradiated bacteria lost their resistance and died on the carpet of spikes in the same way as gram-negative forms.
If the new effect is confirmed by independent groups of researchers, then artificial surfaces with bactericidal microsculpture will undoubtedly find a variety of applications, despite the fact that they do not solve the original task of researchers – to protect the surface from fouling with microbial biofilm. On the contrary, the authors observed that the surface of the cicada's wing was covered with empty bacterial shells layer by layer.
In conclusion, it is worth remembering the singing cicadas themselves. Most likely, the described bactericidal properties of the surface of the wings of Psaltoda claripennis are not a protective biological adaptation of these insects. Firstly, the penetration of pathogenic microbes into the insect through the wing membrane (actually a dead organic film that does not give access to the body cavity and living tissues) is almost impossible. Secondly, adult singing cicadas are terrestrial insects that keep their wings dry. Recent studies (see: Sun et al., 2012. Influence of Cuticle Nanostructuring on the Wetting Behavior/States on Cicada Wings) have shown that it is precisely the sculpture of microscopic spikes that makes the wings of singing cicadas exceptionally water-repellent ("lotus effect"). It should be assumed that the authors were lucky enough to find a useful application for this structure outside the biological context in which it originated and exists in nature.
Sources:
1) Ivanova et al., Natural bactericidal surfaces: mechanical rupture of Pseudomonas aeruginosa cells by cicada wings // Small. 2012.
2) Hasan et al., Selective bactericidal activity of nanopatterned superhydrophobic cicada Psaltoda claripennis wing surfaces // Applied microbiology and biotechnology. 2012.
3) Pogodin et al., Biophysical model of bacterial cell interactions with nanopatterned cicada wing surfaces // Biophysical journal. 2013.
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