A sequential utilization of the UV-A (365 nm) fluence rate for disinfection of water, contaminated with Legionella pneumophila and Legionelladumoffii.

Legionella species are the etiological agent of Legionnaires' disease, a pathology easily contracted from water circuits and by the inhalation of aerosol droplets. This bacterium mainly proliferates in water: Legionella pneumophila is the most commonly isolated specie in water environments and consequently in water system, although further Legionella species have frequently been isolated, including Legionella dumoffii. The simultaneous presence of the two species in the water system can therefore lead to the simultaneous infection of several people, giving rise to harmful outbreaks. Ultraviolet inactivation of waterborne microorganisms offers a rapid and effective treatment technique and recently is getting more attention mostly to eliminate unsafe level of contamination. To tackle the issue, the inactivation of the two species of Legionella spp., namely L. pneumophila and L. dumoffii, by means of UV-A light emitting diodes (UV-A LED) system is explored. We used a commercially available UV-A LED at 365 nm wavelength, and the UV-A dose is given incrementally to the Legionellae with a concentration of 106 CFU/mL in 0.9% NaCl (aq) solution. In this study, with a UV-A-dose of 1700 mJ/cm2, the log-reduction of 3-log (99.9% inactivation) for L. pneumophila and 2.1-log (99.1% inactivation) for L. dumoffii of the contaminated water are achieved. The Electrical Energy per Order (EEO) is evaluated and showed this system is more economic and efficient in comparison with UV-C and UV-B LEDs. Following the support of this preliminary study with additional tests, aiming to validate the technology, we expect this device may be installed in water plants such as cooling systems or any water purification station in either industrial or home scales to reduce the risk of this infectious disease, preventing consumers' health.

Genetic Manipulation of Non-pneumophila Legionella: Protocols Developed for Legionella longbeachae.

Current biomedical research into Legionnaires' disease is dominated by studies of Legionella pneumophila, largely because this pathogen is responsible for approximately 90% of clinical disease worldwide. However, in certain geographical regions, infections with non-pneumophila species are responsible for a significant proportion of diagnosed Legionnaires' disease. Understanding the pathogenesis of these non-pneumophila species of Legionella is an important step toward clinical intervention. The capacity to genetically manipulate these pathogens is essential in order to understand the genetic factors that contribute to infection and the environmental life cycle of these bacteria. The capacity to delete, mutate, and relocate genetic regions of interest allows molecular research into gene function and importance. In this chapter, methods are outlined to introduce plasmids into Legionella by electroporation. This technique is particularly useful as it is often the essential preliminary step to experiments that observe the behavior of the bacterium under altered conditions, for example, the transformation of bacteria with reporter plasmids to monitor Dot/Icm effector translocation. Electroporation is a well-established method for transformation of competent bacteria, and here specific protocols are provided, suiting a range of materials and conditions that have been successfully applied to L. longbeachae and L. dumoffii. Additionally, a homologous recombination approach to delete genetic regions of interest in L. longbeachae is outlined. The application of these techniques allows for identification of the genetic determinants of non-pneumophila Legionella virulence and for important comparative studies with other Legionella species.