Identification of airborne bacteria from an indoor environment and initial characterisation of a multi-drug resistant Kytococcus sedentarius isolate

Abstract

This study was conducted to identify the diversity of airborne bacteria in the Sheffield university lecture theatre (MBB-F2) through passive monitoring (PM) and active monitoring (AM). A total of 6 and 7 bacteria were isolated from settle plates as a PM and dehumidifier as AM respectively. Samples were taken over an hour a week on each Friday after 17.00 for eight weeks, and each sampler isolated different bacteria. Bacteria isolated from dehumidifier were screened against a selection of antibiotics according to British Society for Antimicrobial Chemotherapy (BCAS). Disc diffusion and MIC methods indicated that Kytococcus sedentarius (designed MBB13) is resistant to gentamicin, ciprofloxacin and erythromycin; and the latter is normally a successful treatment for its infections. Kytococcus sedentarius is found on the skin, mucosae and oropharynx and also an opportunistic pathogen causing pitted keratolysis (PK), valve endocarditis and hemorrhagic pneumonia. Thus, features of this organism, together with the complete genome sequence, annotation and in vitro studies were used to describe this organism. Data confirmed that K. sedentarius MBB13 is a strict aerobe; unable to cause PK due to its inability to degrade keratin; and under salt stress, it is able to produce glycine betaine as a main compatible solute and other solutes such as glutamate, proline, acetate, lactate and ectoine. Kytococcus sedentarius MBB13 is an actinomyces and its genome revealed four WhiB proteins (1, 2, 3 and 7) that are restricted to actinomyces and four sigma factors (A, B, H and J). In Mycobacterium tuberculosis, WhiB proteins act as nitric oxide-responsive transcription regulators at least in part through interaction with the major sigma factor A ‘ σA’. The fundamental role of WhiB proteins in actinomyces’ developmental processes proposed that they could play a role in entry into and emergence from the non-replicative persistent state. The M. tuberculosis WhiB1 and WhiB2 are essential and encode DNA-binding proteins with a nitric oxide (NO) sensitive [4Fe-4S] cluster. NO is an important component of the host response to M. tuberculosis and a high dose of NO generated by macrophages can kill the bacilli but a low dose can promote transition to the dormant non-replicating state. Here, K. 3 sedentarius MBB13 WhiB1 and WhiB2 are NO-sensitive; WhiB1 is O2-insensitive but not WhiB2; and the cluster of WhiB1 is essential for the protein folding. A quantitative real time PCR analysis following exposure to a variety of growth and antibiotics indicates differential in upregulation of whiB genes. In vivo protein-protein interaction indicates that all four WhiB proteins of K. sedentarius MBB13 interact with the C-terminal domain of σA but not with the other sigma factors. The sensitivity of WhiB1 and WhiB2 to NO, differentiation of whiB genes expression due to variety of growth and antibiotics, ability of WhiB proteins to interact with SigA and the structural characterization of folding WhiB1 suggest that those proteins may have a role in regulating the pathogenicity of K. sedentarius MBB13.