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New microorganism isolation techniques with emphasis on laser printing

Figure 2. Gelatin-based micro-three-dimensional printing in the presence of bacteria. (Left) engineering polymicrobial communities (right)
“Reprinted from (Connell J.L., Ritschdorff E.T., Whiteley M., Shear J.B. 3D printing of microscopic bacterial communities // Proc. Natl.
Acad. Sci. - 2013. - Vol. 110 - № 46. - P. 18380–18385).”

plates. Thus, this method is important in the fields of be crosslinked into porous, durable, and biocompatible
chemical biology, chemistry of natural products, as well hydrogels, was used to create microscopic 3D bacterial
as for the discovery and characterization of biologically chambers. Some separate bacteria were enclosed in
active secondary metabolites. the BSA chambers with the volume of 1 pl and were
The approaches mentioned above have been useful then grown in clonal populations. Due to the diffusion
for limiting the size, shape, and physical attributes of biologically significant molecules and antibiotics
(microhabitat); however, none of them have provided the through the walls of BSA, as well as the exchange of
opportunity to determine the 3D geometry of bacterial quorum-sensitive signals, the social behavior of bacterial
aggregates or the orientation of multiple populations communities was studied in relation to the size and
arbitrarily. In addition, the process of cell encapsulation density of the population, the shape of the container, and
in ultra-low volume cavities often limits mass transport, the flow rate of the environment.
leading to conditions that are incompatible with growth Within the human body, bacteria usually exist in
and signaling between physically isolated populations. structured 3D communities consisting of several bacterial
A growing number of proofs highlight the importance of species. To get detailed information about the effect of
microcolonies in bacterial reproduction ; however, the geometry on pathogenicity, a 3D printing strategy is
[27]
lack of tools for systematic assessment of cell behavior described in Connell et al. for bacterial communities,
[52]
in such communities is observed. New strategies for the in which physically distinct but chemically interactive
creation of a 3D cultural environment on a microscopic populations of a certain size, shape, and density can
scale can play a crucial role in identifying how bacteria be organized in any form essentially (Figure 2). Using
manage antibiotic resistance and other social behaviors in this approach, it was shown that the stability of a single
small dense aggregates. pathogenic species of bacteria to the antibiotic might
In Connell et al. and Connell et al. [46,47] , the method of enhance the resistance of the second species because of
laser formation of microscopic 3D chambers based on their 3D relations. With the help of laser lithographic
multiphoton lithography (MPL) [48-51] is described. The technique, microscopic containers of up to 1 pl volume
MPL method has high-throughput capacity and the ability with a container wall thickness of up to 2 µm around
to produce arbitrary patterns. It offers opportunities for the selected bacteria suspended in gelatin were formed by
industrial production of 3D microdevices such as micro- cross-linking of the polypeptide molecules in the focal
optical components, scaffolds for tissue engineering, region due to non-linear absorption of laser radiation by
and microfluidic chips. In Connell et al. study , bovine photosensitizer molecules. The result of this multiphoton
[52]
serum albumin (BSA), a highly soluble protein that can absorption is the formation of singlet oxygen, which

4 International Journal of Bioprinting (2019)–Volume 5, Issue 1