Methicillin-resistant Staphylococcus aureus (MRSA) is a global public health
problem and is a major cause of morbidity and mortality worldwide, imposing serious
economic costs on patients and hospitals. Prior to the mid-1990s, MRSA was largely
a healthcare-associated pathogen, causing infection predominantly in people with
frequent or recent contact with healthcare facilities (HA-MRSA). Since then,
community-associated MRSA (CA-MRSA), which often causes infection among
healthy children and young adults with no exposure to the healthcare setting, has
become increasingly prevalent. Worryingly, there is evidence that CA-MRSA is
penetrating the healthcare MRSA reservoir, and even replacing traditional HA-MRSA
strains. This highlights the need to keep abreast of the changing epidemiology of
MRSA in order to implement effective infection control strategies. To investigate the
composition of the healthcare MRSA reservoir and ascertain the extent to which CAMRSA
has penetrated this reservoir, a countywide, population-based cohort study of
MRSA in hospital inpatients and nursing home residents was conducted in Orange
County (OC), California, covering a total of 46 facilities. CA-MRSA was found to be
fully mixed with HA-MRSA in the hospital setting. The predominant CA-MRSA
clone in the US, USA300, was the most commonly isolated MRSA clone in OC
hospitals. In OC nursing homes, HA-MRSA (specifically a variant of USA100 that is
also very common in OC hospitals but has not been reported elsewhere)
predominates, but USA300 made up just over a quarter of the isolates and was the
second most frequently isolated clone. Both OC hospitals and nursing homes were
dominated by the same three strains: USA300, USA100 and a variant of USA100.
Not only are community-based infection control strategies needed to stem the influx
of community associated strains, in particular USA300, into the hospital setting, but
also strategies tailored to the complex problem of MRSA transmission and infection in nursing homes, to minimise the impact of the unique nursing home MRSA
reservoir on overall regional MRSA burden. A key component of effective infection
control strategies is prompt isolation of MRSA carriers, facilitated by rapid
diagnostics. PCR-based methods of MRSA detection offer a much faster alternative to
traditional culture techniques, but are expensive and often complex to operate. A
novel nucleic acid amplification technique developed by my industrial sponsor,
TwistDx Ltd, called recombinase polymerase amplification (RPA), has been
incorporated into a probe based detection system called TwistAmp MRSA, and offers
a simple and cheap alternative to current commercial PCR-based assays, amplifying
MRSA to detectable levels within 20 minutes. I tested the assay with diverse
collections of MRSA and discovered that 4% of isolates from a UK MRSA collection
could not be detected by the assay. I subsequently developed RPA primers for their
detection. Nonetheless, TwistAmp MRSA was able to detect most MRSA strains, and
was comparable to current commercial assays in this respect. Despite a very high
analytical sensitivity of approximately 20 CFU/swab, the clinical sensitivity of
TwistAmp MRSA was lower than expected with respect to the current market leader,
Xpert MRSA. I investigated lysis and filtration methods to improve the assay's
clinical sensitivity, but found that such methods did not currently warrant inclusion in
the TwistAmp MRSA protocol. While TwistAmp MRSA performance is in line with
current assays, and is a faster, cheaper and simpler assay, a problem faced by all
molecular methods of MRSA detection is the constant emergence of undetectable
MRSA strains, necessitating continual assay evaluation and improvement where
possible.