3D printing by two-photon polymerization enables the fabrication of microstructures with complex shapes and critical dimensions of a few hundreds of nanometers. On state-of-the art commercial two-photon polymerization systems, an immense 3D design freedom can be put into practice by direct laser writing using a precise fabrication technology, which makes this approach highly attractive for different applications on the microscale, such as microrobotics, micro-optics, or biosensing. However, navigating the different possible configurations and selecting the optimal parameters for the fabrication process often requires intensive testing and optimization. In addition to the more established acrylate-based resins, there is a growing interest in the use of soft materials. In this paper, we demonstrate the fabrication of various microscale structures by two-photon polymerization using a Nanoscribe Photonic Professional GT+ commercial system. Furthermore, we describe the different configurations of the system and parameter selection, as well as commercial resins and their chemical and mechanical properties. Finally, we provide a short guide aiming to serve as starting point for the two-photon polymerization-based fabrication of various microscale architectures with distinct characteristics.

The disinfection efficiency of synthetic and real wastewater by means of UV-A and UV-C irradiation in the presence or absence of TiO₂ was investigated. A reference strain of Escherichia coli suspended in sterile 0.8% (w/v) NaCl aqueous solution was used as a synthetic wastewater, while real wastewater samples were collected from the outlet of the secondary treatment of a municipal wastewater treatment plant. E. coli inactivation was monitored both by the conventional culture technique and by the real-time PCR method. Culture method showed that UV-C irradiation (11 W lamp) achieved total E. coli inactivation of 100% within 3 min of photolytic treatment. On the other hand, UV-A (9 W lamp)/TiO₂; [TiO₂] = 200 mg L⁻¹ (i.e. best operating conditions) required 60 min to achieve total disinfection of the synthetic wastewater. Real time PCR revealed compatible results, regarding the better efficiency of UV-C. However, it showed different times of bacterial inactivation, probably due to the phenomenon of “viable but not culturable bacteria”. Disinfection durability tests in the dark and under natural sunlight irradiation showed that there is cell repair when UV-C irradiation is used for synthetic wastewater disinfection. Regarding real wastewater it was observed that only UV-C irradiation was capable of totally inactivating E. coli population in short time. Comparing results obtained from both methods, real time PCR proved to be more reliable and accurate, concerning the bacterial detection and enumeration in aquatic samples after the application of UV irradiation.

Mobilizable plasmids lack necessary genes for complete conjugation and are therefore non-self-transmissible. Instead, they rely on the conjugation system of conjugal plasmids to be horizontally transferred to new recipients. While community permissiveness, the fraction of a mixed microbial community that can receive self-transmissible conjugal plasmids, has been studied, the intrinsic ability of a community to mobilize plasmids that lack conjugation systems is unexplored. Here, we present a novel framework and experimental method to estimate the mobilization potential of mixed communities. We compare the transfer frequency of a mobilizable plasmid to that of a mobilizing and conjugal plasmid measured for a model strain and for the assayed community. With Pseudomonas putida carrying the gfp-tagged mobilizable RSF1010 plasmid as donor strain, we conducted solid surface mating experiments with either a P. putida strain carrying the mobilizing plasmid RP4 or a model bacterial community that was extracted from the inner walls of a domestic shower conduit. Additionally, we estimated the permissiveness of the same community for RP4 using P. putida as donor strain. The permissiveness of the model community for RP4 (at 1.16x10-4 transconjugants per recipient (T/R)) was similar to that previously measured for soil microbial communities. RSF1010 was mobilized by the model community at a frequency of 1.16x10-5 T/R, only one order of magnitude lower than its permissiveness to RP4. This mobilization frequency is unexpectedly high considering that (i) mobilization requires the presence of mobilizing conjugal plasmids within the permissive fraction of the recipients; (ii) in pure culture experiments with P. putida retromobilization of RSF1010 through RP4 only took place in approximately half of the donors receiving the conjugal plasmid in the first step. Further work is needed to establish how plasmid mobilization potential varies within and across microbial communities.

Several marine bacteria of the Roseobacter group can inhibit other microorganisms and are especially antagonistic when growing in biofilms. This aptitude to naturally compete with other bacteria can reduce the need for antibiotics in large-scale aquaculture units, provided that their culture can be promoted and controlled. Micropatterned surfaces may facilitate and promote the biofilm formation of species from the Roseobacter group, due to the increased contact between the cells and the surface material. Our research goal is to fabricate biofilm-optimal micropatterned surfaces and investigate the relevant length scales for surface topographies that can promote the growth and biofilm formation of the Roseobacter group of bacteria. In a preliminary study, silicon surfaces comprising arrays of pillars and pits with different periodicities, diameters, and depths were produced by UV lithography and deep reactive ion etching (DRIE) on polished silicon wafers. The resulting surface microscale topologies were characterized via optical profilometry and scanning electron microscopy (SEM). Screening of the bacterial biofilm on the patterned surfaces was performed using green fluorescent staining (SYBR green I) and confocal laser scanning microscopy (CLSM). Our results indicate that there is a correlation between the surface morphology and the spatial organization of the bacterial biofilm.