Cracking is an inevitable feature of concrete, typically leading to corrosion of the embedded steel reinforcement and massive deterioration because of the freezing–thawing cycles. Different means have been proposed to increase the serviceability performance of cracked concrete structures. This case study deals with bacteria encapsulated in cementitious materials to “heal” cracks. Such a biological self-healing system requires preserving the bacteria’s viability in the cement matrix. Many embedded bacterial spores are damaged during concrete curing, drastically reducing efficiency. This study investigates the viability of commonly used non-ureolytic bacterial spores when immobilized in calcium alginate microcapsules within self-healing cementitious composites. Three Bacillus species were used in this study, i.e., B. pseudofirmus, B. cohnii, and B. halodurans. B. pseudofirmus demonstrated the best mineralization activity; a sufficient number of bacterial spores remained viable after the encapsulation. B. pseudofirmus and B. halodurans spores retained the highest viability after incorporating the microcapsules into the cement paste, while B. halodurans spores retained the highest viability in the mortar. Cracks with a width of about 0.13 mm were filled with bacterial calcium carbonate within 14 to 28 days, depending on the type of bacteria. Larger cracks were not healed entirely. B. pseudofirmus had the highest efficiency, with a healing coefficient of 0.497 after 56 days. This study also revealed the essential role of the cement hydration temperature on bacterial viability. Thus, further studies should optimize the content of bacteria and nutrients in the microcapsule structure.

One of the biggest challenges in the development of a biological self-healing concrete is to ensure the long-term viability of bacteria that are embedded in the concrete. In the present study, a coated expanded clay (EC) is investigated for its potential use as a bacterial carrier in biological concrete. Eight different materials for coatings were selected considering cost, workability and accessibility in the construction industry. Long-term (56 days) viability analysis was conducted with a final evaluation of each coating performance. Our results indicate that healing efficiency in biological concrete specimens is strongly related to viable bacteria present in the healing agent. More viable bacteria-containing specimens exhibited a higher crack closure ratio. Our data suggest that the additional coating of EC particles improves long-term bacterial viability and, consequently, provides efficient crack healing in biological concrete.

The capacity of biological self-healing concrete (BSHC) to repair cracks relies on the sustained viability and metabolic function of bacteria embedded within the concrete. BSHC structures face significant risk in cold climates due to low temperatures and freeze–thaw (FT) cycles, during which freezing water can generate internal pressure that damages bacterial cells and diminishes their activity. A special feature of this study is the incorporation of bacterial spores within expanded clay aggregates, tested under varying environmental conditions. The viability of bacterial spores was measured under cold and freeze–thaw cycles by counting colony-forming units, and a specific methodology was developed to assess the efficiency of self-healing under rain-simulated conditions. It was demonstrated that bacteria embedded in concrete could endure fluctuations in low temperatures and freeze–thaw cycles, compromising approximately 50% of viable spores. Also, it was found that water immersion during concrete curing can trigger early germination, decreasing viable spore counts by nearly tenfold. Ultimately, it was demonstrated that the healing of cracks in BSHC components is influenced by the conditions under which the specimens are incubated. The results suggest that BSHC can be employed in cold climate areas, given that suitable curing conditions and adequate bacterial protection within the concrete are ensured.

N⁶-methyladenine and N⁶,N⁶-dimethyladenine are the heterocyclic bases present in the RNA of eukaryotic and bacterial cells and play important regulatory roles. How the degradation of such modified nucleic acids, and the subsequent demethylation of modified heterocyclic bases, occurs in the bacterium Escherichia coli is not established. Here, we investigated the growth of adenine auxotroph strains in a minimal M9 medium supplemented with either N⁶-methyladenine or N⁶,N⁶-dimethyladenine. We found that N⁶-methyladenine supported the growth of ∆purH::Km but not that of the ∆purA::Km strain, whereas N⁶,N⁶-dimethyladenine did not support the growth of either adenine auxotroph. Similar experiments performed using structurally related 2-amino-N⁶-methylpurine and 2-amino-N⁶,N⁶-dimethylpurine bases—using ∆guaA::Km, ∆guaB::Km, and ∆purH::Km guanine auxotrophs—demonstrated that growth of only the ∆guaB::Km mutant was supported by 2-amino-N⁶-methylpurine but not by its dimethylated counterpart. We expressed and purified C-teminus 6xHis tagged E. coli adenine/adenosine deaminases AdeC and Add and tested their substrate specificity. We demonstrated that AdeC protein does not catalyse deamination of either N⁶-methyl- or N⁶,N⁶-dimethyladenine, whereas Add catalyses deamination of N⁶-methyl- but not that of N⁶,N⁶-dimethyladenosine. Based on our findings, biochemical pathways leading to the demodification and return into metabolism of N⁶-methyladenine and 2-amino-N⁶-methylpurine in E. coli are proposed.