Characterization of the Desiccation Tolerance and Potential for Fomite Spread of Streptococcus pneumoniae.
Streptococcus pneumoniae is the causative agent of diseases such as otitis media,
pneumonia, and bacterial meningitis. Historically, published literature asserted that
pneumococcal transmission occurs via direct contact with respiratory secretions from
infected individuals, yet spread of other respiratory tract pathogens has been linked to
fomites. As the only known reservoir of S. ... read morepneumoniae is the human upper respiratory
tract, little work has investigated its capacity to survive in the environment.
Environmental persistence often depends on an ability to withstand desiccation. A
knowledge gap exists regarding the viability of vegetative cells, particularly those of
major human pathogens, post dehydration. In an era where nosocomial infections are
increasingly common, this has become an important area of study. The goal of this work
is to characterize the ability of S. pneumoniae to withstand desiccation. We hypothesize
that fomites serve as an alternate source of pneumococcal transmission. To test our
hypothesis, we ask three central questions. Can S. pneumoniae survive desiccation? Is S.
pneumoniae infectious after desiccation? And which genes or pathways are involved in
desiccation tolerance? Here, we describe the development of a desiccation protocol using
plate-grown S. pneumoniae. We show that pneumococci can survive at least 28 days of
desiccation under ambient conditions, survival is independent of polysaccharide capsule,
and desiccation is a more complicated phenomenon than starvation alone. Multiple
serotypes survive desiccation, suggesting that this is a species-wide ability encoded by
the core pneumococcal genome. Finally, S. pneumoniae retains infectivity in a mouse
model of colonization, lending support to the fomite transmission hypothesis. To explore
the pathways correlated with desiccation tolerance, we constructed a transposon
insertion library, which was desiccated for two and four days, after which we performed
the Tn-seq method, a high throughput sequencing and quantitation of the transposon
junctions. By comparing the frequency of reads for each insertion in the pre-and
post-desiccation samples, we determined the importance of each gene for S. pneumoniae's
desiccation survival. Of 12 genes selected for further study, mutants in genes encoding
the UvrABC nucleotide excision repair complex, SpxB (pyruvate oxidase), SodA (superoxide
dismutase), MscS (mechanosensitive channel of small conductance), and cardiolipin
synthase all display phenotypes in stress assays. We demonstrate that S. pneumoniae's
desiccation tolerance is tied to its ability to withstand stresses, including those
imposed by osmotic changes and reactive oxygen species, leading to a need for DNA damage
Thesis (Ph.D.)--Tufts University, 2015.
Submitted to the Dept. of Molecular Microbiology.
Advisor: Andrew Camilli.
Committee: Ralph Isberg, Carol Kumamoto, and Joan Mecsas.
Keywords: Microbiology, and Molecular biology.read less