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Desiccation-tolerance of cells of four strains of Mycobacterium chimaera and individual strains of Mycobacterium avium, Mycobacterium intracellulare, Mycobacterium abscessus, and Mycobacterium chelonae were measured by two methods. The survival of water-acclimated cells both in filter paper and on the surface of stainless-steel coupons were measured. In filter paper at 40% relative humidity at 25 °C, survival of patient isolates of M. avium and M. chimaera cells was 28% and 34% after 21 days of incubation, whereas it was 100% for the Sorin 3T isolate of M. chimaera. On stainless-steel biofilms after 42 days of incubation at 40% relative humidity at 25 °C, survival of water-acclimated cells of M. intracellulare was above 100%, while M. chelonae cells did not survive beyond 21 days, and survival of water-acclimated cells of M. avium and M. abscessus was 18% and 14%, respectively. On stainless-steel coupons, survival of patient and Sorin 3T isolates of M. chimaera was quite similar, specifically between 14% and 28% survival, after 42 days of incubation at 40% relative humidity at 25 °C. The experiments would support the hypothesis that some nontuberculous mycobacterial species are relatively desiccation-tolerant, whereas others are not. Further, long-term survival of the two M. chimaera strains is consistent with the presence of that species in Sorin 3T heater-coolers shipped throughout the world.

Nontuberculous mycobacteria (NTM) represent an important group of environmentally saprophytic and potentially pathogenic bacteria that can cause serious mycobacterioses in humans and animals. The sources of infections often remain undetected except for soil- or water-borne, water-washed, water-based, or water-related infections caused by groups of the Mycobacterium (M.) avium complex; M. fortuitum; and other NTM species, including M. marinum infection, known as fish tank granuloma, and M. ulcerans infection, which is described as a Buruli ulcer. NTM could be considered as water-borne, air-borne, and soil-borne pathogens (sapronoses). A lot of clinically relevant NTM species could be considered due to the enormity of published data on permanent, periodic, transient, and incidental sapronoses. Interest is currently increasing in mycobacterioses diagnosed in humans and husbandry animals (esp. pigs) caused by NTM species present in peat bogs, potting soil, garden peat, bat and bird guano, and other matrices used as garden fertilizers. NTM are present in dust particles and in water aerosols, which represent certain factors during aerogenous infection in immunosuppressed host organisms during hospitalization, speleotherapy, and leisure activities. For this Special Issue, a collection of articles providing a current view of the research on NTM—including the clinical relevance, therapy, prevention of mycobacterioses, epidemiology, and ecology—are addressed.

Antimicrobial coatings are one method to reduce the spread of microbial diseases. Transparent coatings preserve the visual properties of surfaces and are strictly necessary for applications such as antimicrobial cell phone screens. This work describes transparent coatings that inactivate microbes within minutes. The coatings are based on a polydopamine (PDA) adhesive, which has the useful property that the monomer can be sprayed, and then the monomer polymerizes in a conformal film at room temperature. Two coatings are described (1) a coating where PDA is deposited first and then a thin layer of copper is grown on the PDA by electroless deposition (PDA/Cu) and (2) a coating where a suspension of Cu2O particles in a PDA solution is deposited in a single step (PDA/Cu2O). In the second coating, PDA menisci bind Cu2O particles to the solid surface. Both coatings are transparent and are highly efficient in inactivating microbes. PDA/Cu kills >99.99% of Pseudomonas aeruginosa and 99.18% of methicillin-resistant Staphylococcus aureus (MRSA) in only 10 min and inactivates 99.98% of SARS-CoV-2 virus in 1 h. PDA/Cu2O kills 99.94% of P. aeruginosa and 96.82% of MRSA within 10 min and inactivates 99.88% of SARS-CoV-2 in 1 h.

Spontaneous mutants of Mycobacterium smegmatis strain mc(2)155 resistant to 1-PG (iridium-L-phenylglycine complex), an antimycobacterial antibiotic, were isolated. Based on the discovery that some 1-PG-resistant mutants (1-PG(R)) were also resistant to high concentrations of clarithromycin (>= 250 mu g/ml), but no other anti-mycobacterial antibiotics, the 23S rRNA region spanning the peptidyl transferase domain was sequenced and mutations shown to be localized in the peptidyl transferase domain of the 23S rRNA gene. Measurements showed that 1-PG bound to ribosomes isolated from the 1-PG-sensitive parental strain, but the ribosome binding values for the 1-PG(R) mutant reduced.

Legionella spp. and other opportunistic premise plumbing pathogens (OPPPs), including Pseudomonas aeruginosa, Mycobacterium avium, Stenotrophomonas maltophilia, and Acinetobacter baumannii, are normal inhabitants of natural waters, drinking water distribution systems and premise plumbing. Thus, humans are regularly exposed to these pathogens. Unfortunately, Legionella spp. and the other OPPPs share a number of features that allow them to grow and persist in premise plumbing. They form biofilms and are also relatively disinfectant-resistant, able to grow at low organic matter concentrations, and able to grow under stagnant conditions. Infections have been traced to exposure to premise plumbing or aerosols generated in showers. A number of measures can lead to reduction in OPPP numbers in premise plumbing, including elevation of water heater temperatures.

Members of the Mycobacterium avium complex (MAC) are waterborne, opportunistic pathogens whose characteristics make urban water distribution systems and household plumbing ideal habitats for their survival, persistence and growth. Rather than contaminants, MAC are colonists of drinking water systems. MAC are normal inhabitants of natural soils and water, and enter drinking water treatment systems through surface sources. A proportion of MAC survive transmission through the treatment plant, and regrow in the distribution system and household plumbing. Once within household plumbing, MAC adhere to surfaces and form biofilms, thus preventing their washout. The thermal tolerance of MAC leads to growth in water heating systems. Stagnation does not reduce MAC numbers, as MAC can grow at low oxygen levels. MAC present challenges to current water monitoring approaches as their numbers do not correlate with E. coli, fecal coliforms or heterotrophic plate count bacteria.

Mycobacteria are unique in many aspects of their biology. The development of genetic tools to identify genes critical for their growth by forward genetic analysis holds great promises to advance our understanding of their cellular, physiological and biochemical processes. Here we report the development of a novel transposon, MycoTetOP(2), to aid the identification of such genes by direct transposon mutagenesis. This mariner-based transposon contains nested anhydrotetracycline (ATc)-inducible promoters to drive transcription outward from both of its ends. In addition, it includes the Escherichia coli R6K gamma origin to facilitate the identification of insertion sites. MycoTetOP(2) was placed in a shuttle plasmid with a temperature-sensitive DNA replication origin in mycobacteria. This allows propagation of mycobacteria harboring the plasmid at a permissive temperature. The resulting population of cells can then be subjected to a temperature shift to select for transposon mutants. This transposon and its delivery system, once constructed, were tested in the fast-growing model Mycobacterium smegmatis and 13 mutants with ATc-dependent growth were isolated. The identification of the insertion sites in these mutants led to nine unique genetic loci with genes critical for essential processes in both M. smegmatis and Mycobacterium tuberculosis. These results demonstrate that MycoTetOP(2) and its delivery vector provide valuable tools for the studies of mycobacteria by forward genetics.

Nontuberculous mycobacteria (NTM) that cause human disease can be isolated from household tap water. Easy-to-use physical methods to reduce NTM from this potential source of exposure are needed. Filters and UV disinfection have been evaluated for their ability to reduce numbers of waterborne non-NTM organisms from drinking water, but their efficacy in reducing NTM counts are not well-established. Thus, five commercially available disinfection methods were evaluated for their potential as practical, efficient, and low-cost methods to reduce NTM from tap water. First, suspensions of tap water-adaptedMycobacterium smegmatiswere passed through either a point-of-use, disposable, 7-day or 14-day Pall-Aquasafe filter. The 7-day filter prevented passage ofM. smegmatisin effluent water for 13 days, and the 14-day filter prevented the passage ofM. smegmatisfor 25 days. Second, a granular activated carbon filter system failed to significantly reduceMycobacterium abscessusandMycobacterium aviumnumbers. Third, suspensions of tap water-adaptedM. abscessus, M. avium, andM. chimaera("MycoCocktail") were passed through the "LifeStraw GO" hollow-fiber, two-stage membrane filtration system. LifeStraw GO prevented passage of the MycoCocktail suspension for the entire 68-day evaluation period. Finally, two different water bottle UV sterilization systems, "Mountop" and "SteriPEN," were evaluated for their capacity to reduce NTM numbers from tap water. Specifically, MycoCocktail suspensions were dispensed into Mountop and SteriPEN water bottles and UV treated as per the manufacturer instructions once daily for 7 days, followed by a once weekly treatment for up to 56 days. After 4 days of daily UV treatment, both systems achieved a >4 log reduction in MycoCocktail CFU. After the 56-day evaluation period, suspension and biofilm-associated CFU were measured, and a >4 log reduction in CFU was maintained in both systems. Taken together, physical disinfection methods significantly reduced NTM n

Legionella spp. and other opportunistic premise plumbing pathogens (OPPPs), including Pseudomonas aeruginosa, Mycobacterium avium, Stenotrophomonas maltophilia, and Acinetobacter baumannii, are normal inhabitants of natural waters, drinking water distribution systems and premise plumbing. Thus, humans are regularly exposed to these pathogens. Unfortunately, Legionella spp. and the other OPPPs share a number of features that allow them to grow and persist in premise plumbing. They form biofilms and are also relatively disinfectant-resistant, able to grow at low organic matter concentrations, and able to grow under stagnant conditions. Infections have been traced to exposure to premise plumbing or aerosols generated in showers. A number of measures can lead to reduction in OPPP numbers in premise plumbing, including elevation of water heater temperatures.

Members of the Mycobacterium avium complex (MAC) are waterborne, opportunistic pathogens whose characteristics make urban water distribution systems and household plumbing ideal habitats for their survival, persistence and growth. Rather than contaminants, MAC are colonists of drinking water systems. MAC are normal inhabitants of natural soils and water, and enter drinking water treatment systems through surface sources. A proportion of MAC survive transmission through the treatment plant, and regrow in the distribution system and household plumbing. Once within household plumbing, MAC adhere to surfaces and form biofilms, thus preventing their washout. The thermal tolerance of MAC leads to growth in water heating systems. Stagnation does not reduce MAC numbers, as MAC can grow at low oxygen levels. MAC present challenges to current water monitoring approaches as their numbers do not correlate with E. coli, fecal coliforms or heterotrophic plate count bacteria.

Mycobacterium avium is capable of an adaptive, reversible response to high-temperature survival depending on its growth temperature. Trehalose concentrations of M. avium cells grown at 42 °C were significantly higher compared to those of cells grown at 25 °C. Further, the survival of cells of M. avium grown at 42 °C and exposed to 65 °C were significantly higher than the survival of cells grown at 25 °C. This adaptive response to growth temperature may play a role in the persistence of M. avium in premise plumbing.