Epidemic-Associated Neisseria meningitidis Detected by
Multilocus Enzyme Electrophoresis

In Oregon and parts of Washington State, the incidence of
serogroup B meningococcal disease increased substantially
in 1994 (1). Multilocus enzyme electrophoresis (MEE)
subtyping of N. meningitidis serogroup B strains
collected in these areas during 1993 and 1994 suggested
that these increases were due to a group of genetically
related strains of the enzyme type-5 (ET-5) complex. ET-5
N. meningitidis serogroup B were first recognized in
Norway in 1974 as the cause of a meningococcal disease
epidemic that persisted through 1991. Since 1974,
serogroup B meningococci of the ET-5 complex have caused
epidemics in Europe, Cuba, and South America; these
epidemics elevated disease rates for many years in the
affected areas (2,3) and led to sustained efforts for
vaccine development. This report describes the use of MEE
to compare invasive N. meningitidis serogroup B
meningococcal strains from Oregon and Washington with
epidemic serogroup B strains from other countries and
with serogroup B strains that have caused endemic disease
in other parts of the United States.

MEE, first described in 1966 as a molecular approach to
the study of genetic variation in eukaryotic systems, has
only gradually been adopted by microbiologists and
epidemiologists. The fundamental concept underlying MEE
is that differences in the electrophoretic mobility of
constitutive enzymes (resulting from amino acid
substitutions) reflect the chromosomal genotype of
strains and thereby allow the calculation of a
genetic-relatedness index (Figure 1. [cannot viewd in 
ASCII]). As recently as 1984, only one bacterial species, 
Escherichia coli, had been studied by MEE. Since then, 
however, MEE has been used to characterize genetic 
variation among populations of Legionella spp., 
Bordetella spp., Haemophilus influenzae, Streptococcus 
spp., Listeria monocytogenes, Neisseria meningitidis, 
and other bacteria (4).

To carry out MEE, crude aqueous extracts of bacteria are
electrophoresed in a block of 11% to 12% starch in the
presence of a dilute buffer (pH 8.0). The block is then
cut into thin slices, which are stained to detect
specific enzymes. The distance traveled by each enzyme is
used to create a series of numbers representing the set
of enzyme mobilities characteristic of individual
strains. The number of enzymes used is somewhat arbitrary
and varies between organisms; 15 to 24 enzymes have
usually been adequate to characterize genetic diversity
among bacterial populations. For this investigation,
electrophoretic variations in 24 enzymes were used to
describe genetic variability among isolates of N.
meningitidis serogroup B. N. meningitidis strains used
for this analysis were collected from Oregon (1993-1994,
n=64) and part of Washington State (1993-1994, n=17;
1992, n=2; 1990, n=1; unknown, n=2); serogroup B
meningococcal epidemics outside the United States
(1976-1993; Norway n=1; Cuba n=1; Brazil n=1; and Chile
n=2); and active population-based surveillance for
meningococcal disease in selected areas of the United
States (1991-1994, from the San Francisco Bay area,
Georgia, Maryland, Oklahoma, and Tennessee, n=57). The
epidemic strains tested from Norway and Cuba are the type
used for the outer membrane protein vaccines developed
and tested in these countries.

The MEE data analyzed here (Figure 1 cannot be viewed 
in ASCII) suggest that the increased rates of disease
in Oregon and part of Washington are caused by highly
genetically related N. meningitidis serogroup B strains 
of the ET-5 complex. These strains have been relatively 
rare in the United States. Oregon and Washington strains 
match a strain isolated in Santiago, Chile, during 1993. 
The prolonged duration of some ET-5 serogroup B 
meningococcal epidemics in large regions (e.g., Brazil, 
Argentina, and Chile) demands careful monitoring of this 
organism in the United States. Efforts to identify 
potentially modifiable risk factors for the disease and 
develop a vaccine have been intensified. MEE will 
continue to be the primary means for epidemiologic 
tracking and surveillance of ET-5 complex N. 
meningitidis serogroup B in the United States.

Michael W. Reeves,* Bradley A. Perkins*
Marion Diermayer+, and Jay D. Wenger*
*National Center for Infectious Diseases, Centers for
Disease Control and Prevention (CDC),
Atlanta, Georgia, USA
+Emerging Infections Program, Oregon Dept. of Health,
Portland, Oregon, USA, and Epidemiology Program Office,
CDC, Atlanta, Georgia, USA

References

1. Centers for Disease Control and Prevention. Serogroup
B meningococcal disease--Oregon, 1994. MMWR 1995;44:121-4.
2. Caugant DA, Froholm LO, Bovre K, Holten E, Frasch CE,
Mocca LF, Zollinger WD, Selander RK. Intercontinental
spread of a genetically distinctive complex of clones of
Neisseria meningitidis causing epidemic disease. Proc
Natl Acad Sci USA 1986;83:4927-31.
3. Sacchi CT, Pessoa LL, Ramos SR, Milagres LG, Camargo
MCC, Hidalgo NTR, Melles CEA, Caugant DA, Frasch CE.
Ongoing group B Neisseria meningitidis epidemic in São
Paulo, Brazil, due to increased prevalence of a single
clone of the ET-5 complex. J Clin Microbiol
1992;30:1734-8.
4. Selander RK, Caugant DA, Ochman H, Musser JM, Gilmour
MN, Whittam TS. Methods of multilocus enzyme
electrophoresis for bacterial population genetics and
systematics. Appl Environ Microbiol 1986;51:873-84.