Advanced Techniques in Diagnostic Microbiology

9. Pulsed-Field Gel Electrophoresis 151
(VRE). Nasal carriage of S. aureus occurs in 20–60% of the general population,
and methicillin-resistant S. aureus (MRSA) pose a particular risk for nosocomial
transmission (Kluytmans et al., 1997). It is clear that MRSA can be transferred
between patients in the hospital setting and cause nosocomial infections. Rapidly
assessing the clonal relatedness of these isolates is critical in determining the
extent of transmission during an outbreak and in measuring the strategies for
its containment. PFGE is often used to examine the genetic identity of MRSA
isolates. In a recent publication from our medical center, PFGE results showed
that vertical transmission of one clone of MRSA occurred from a mother to
her preterm infants, followed by horizontal spread to other infants in the same
neonatal intensive care unit (Morel et al., 2002). We demonstrated that the most
likely reservoir of the MRSA clone were the colonized infants (Graham et al.,
2002). It has been reported that some of the MRSA clones, endemic in hospitals
in different countries, may have a high degree of genetic variation, leading to
the appearance of multiple subtype variations (Aires de Sousa and de Lencastre,
2004).
In recent years, the increased prevalence of community-associated (CA)-MRSA
has become a major public health concern (Saiman et al., 2003). In contrast to
hospital-associated (HA)-MRSA, CA-MRSA strains are commonly susceptible to
many antibiotics. Also CA-MRSA appears to have a distinct exotoxin Panton–
Valentine Leukocidin (PVL), which has been associated with severe infections
(Centers for Disease Control, 1999; Baba et al., 2002; Kazakova et al., 2005).
Using PFGE, investigators have found that the typing patterns of CA-MRSA were
distinct from those of HA-MRSA, indicating that different clonal populations
have been successfully propagated in the community. Investigators also reported
that MSSA strain 476 shared an identical PFGE pattern with a CA-MRSA strain,
termed MW2. The only significant difference between the chromosomes of these
two strains was the presence of type IV SCCmec in MW2, suggesting that the
progenitor was the MSSA strain from which MW2 was generated by acquiring
type IV SCCmec (Okuma et al., 2002).
Application to Gram-Negative Bacteria
PFGE has been successfully used as a tool to investigate the epidemiological
relatedness of strains of Gram-negative bacteria including Acinetobacter, Enterobacter,
E. coli, Klebsiella, Pseudomonas, Salmonella, and Serratia within a hospital,
a community, or school or daycare center (Durmaz et al., 2003; Pavlopoulou
et al., 2004). We have reported an outbreak of extended-spectrum beta-lactamase
(ESBL)-producing Klebsiella pneumoniae infections in our neonatal intensive care
unit (NICU) (Gupta et al., 2004). A total of 19 infants were either infected or colonized
with K. pneumoniae, in which 9 of 19 infants developed invasive disease.
Surveillance cultures revealed that two health care workers carried K. pneumoniae
on their hands. The PFGE patterns of isolates from health care workers were compared
with 19 isolates recovered from the infants. Results indicated that the one
clone was shared by both the infants and the hands of health care workers. One