Acute
bacterial meningitis continues to be a life-threatening neurological
emergency that warrants rapid diagnosis and management to prevent
mortality and serious neurological disability. The August 1966 issue of
Indian Pediatrics had a Research article on ‘Bacteriologic Methods in
the diagnosis of Acute Bacterial Meningitis.’ Through this
communication, we present the changes in epidemiology and advances in
the diagnosis of acute bacterial meningitis in last 50 years.
The Past
The reported article by Hughes, et al. [1]
describes the CSF findings of three consecutive cases of acute bacterial
meningitis, out of total 653 admissions to the Institute of Child Health
during 13-month period from January 1965 through 1966. In this paper,
the authors demonstrated the efficiency of improved bacteriologic
cultures over routine cultures in isolating the causative agents of
acute bacterial meningitis. The special bacteriological procedures were
carried out in assistance with Baltimore Biological Laboratory Division
(Baltimore, Maryland) and Difco laboratories (Detroit, Michigan). The
three children described herein had CSF cell count >1000 cell/mm3
with predominance of polymorphonuclear cells.
Apart from CSF cell count, biochemistry, gram stain and routine cultures
(use of sheep blood agar incubated in atmospheric air at 37
0 C for 24h), specialized
bacteriological cultures using sheep blood agar in trypticase soy agar
base (BA) and chocolate agar (CA) plates were employed for isolating the
etiological agents. ‘Candle Jar’ cultures were performed by placing
culture plates or slants in biscuit tin having tight fitting lid which
was sealed while the candle was left inside burning. All the cultures
were maintained for at least 48 hours prior to being discarded as
negative. Selected strains were further typed by agglutination with
commercial antiserum.
The CSF from first two patients ( age 6 mo and 11 mo)
were non-contributory on Gram stain and failed to demonstrate growth on
routine blood agar, except for positive satellitism. However, abundant
growth was noted on CA after incubation in both atmospheric air and
candle jar, and the colonies were characterized as Haemophilus
influenzae. In Case 2, serological identification was established by
agglutination with H. influenzae type b. The third case was a
20-day-old boy, and his CSF revealed Gram positive cocci on gram stain
but failed to grow on ordinary BA incubated in air at 370C.
The use of CA supported growth more than BA under similar incubation
conditions but growth augmentation and appearance of
b hemolysis was
facilitated by incubation in candle jar. The colonies were moderately
sensitive to bacitracin and designated as group A Streptococcus after
precipitation in capillary tubes containing group A antiserum.
The reported article discussed the etiologic agents
of acute bacterial meningitis from Children’s Hospital, Boston (447
cases over a period 1956-60) which revealed H. influenzae as the
most important bacteriological agent implicated. Indian studies in that
era also reported it as the leading cause of non-tuberculous bacterial
meningitis across all age groups [2]. It was realized that H.
influenzae is fastidious organism and its isolation can be
facilitated by the use of CA under candle jar conditions. This medium
also supported the growth and hemolysis of Group A
b- hemolytic
Streptococci, Neisseria meningitides and some strains of
Diplococci pneumoniae (Streptococcus pneumoniae). CA cultures
incubated in candle jar were recommended as the most economical,
convenient and efficient media over routine culture methods used for CSF
analysis in cases of acute bacterial meningitis.
Historical Background and past knowledge:
The earliest treatise on the association of meningitis with bacterial
(meningococcal) infection was by an Austrian bacteriologist Anton
Weichselbaumin 1887, though the first reported outbreak of meningitis
occurred much earlier in Geneva in 1805.The use of lumbar puncture for
early CSF analysis was introduced by Heinrich Quincke in 1891 [3]. The
etiological agents responsible for acute bacterial meningitis received
recognition in the late 19th century [3]. In the pre-antibiotic era,
bacterial meningitis had a uni-formly fatal outcome, until the use of
penicillin in 1944 [4].
The Present
Worldwide, the various epidemiological studies
conducted after publication of the reported article have indicated H.
influenzae type b (Hib), Streptococcus pneumoniae and
Neisseria meningitidis to be the commonest etiological agents of
acute bacterial meningitis in children. Till the end of the 20th
century, the global burden of meningitis due to H. influenzae was
huge, and it contributed a large proportion of under-five mortality and
morbidity [5]. A multi-centre sentinel surveillance study from India
found H. influenzae to be the predominant cause (70%) of bacterio-logically
confirmed meningitis in children under two years of age, while S.
pneumoniae and group B Streptococcus were identified in 13% and 8%
cases, respectively [6]. Introduction of Hib conjugate vaccine has led
to virtual elimination of invasive disease due to H. influenzae
from developed countries and a substantial decline in the incidence of
meningitis in developing countries that have adopted it as part of
routine immunization program. The widespread use of pneumococcal and
meningococcal conjugate vaccines have further influenced the
epidemiology of meningitis in developed countries.
Diagnosis of acute bacterial meningitis relies
heavily on Gram stain, culture and antigen detection by latex
agglutination testing. Culture of CSF continues to remain gold standard
for diagnosis of acute bacterial meningitis. The age old candle jar
continues to be in use, but has given way to special CO2
incubators in modern settings. In India, the isolation rates of these
pathogens is low as compared to West. Tropical environmental conditions
in India probably do not allow the survival of relatively more fragile
bacteria like H. influenzae, N. meningitidis, S.
pneumoniae, S. agalactiae and Listeria monocytogenes
over hardy pathogens. The yield of CSF culture is further reduced in
patients who have received antibiotic before lumbar puncture, delay in
processing of samples or due to storage of these samples in refrigerator
[7]. Recently MALDI-TOF MS (Matrix assisted laser desorption ionization-
time of flight mass spectrometry) has emerged as a rapid automated, low
cost and reliable tool based on mass spectroscopy for microbial
identification and detection of antibiotic resistance [8].
In recent times, molecular diagnostics have
drastically reduced the turnaround time for diagnosis. Nucleic acid
detection techniques such as PCR can detect bacteria in culture negative
patients. Broad range PCR and multiplex PCR have shown high sensitivity
and specificity for detection of H. influenzae, N. meningitides,
Group B Streptococci and S. pneumoniae [9]. Recently FDA has
approved a fully automated DNA extraction and amplification system which
has drastically reduced turnaround time for detection of DNA to just one
hour. This system requires 200 µL of CSF to test for a multiplex panel
of six bacteria (H. influenzae, N. meningitides, Group B
Streptococci, S. pneumoniae, Listeria monocytogenes and E coli
K1), eight viruses and two fungi [10]. However, their utilization in
routine diagnostic practice is hindered by the high running cost.
Overall, etiological diagnosis of acute bacterial meningitis still
remains a challenge, especially in low- and middle-income countries.
References
1. Hughes R, Brahmachari S. Bacteriologic Methods in
the diagnosis of acute bacterial meningitis. Indian
Pediatr.1966;3:281-5.
2. Grant KB, Ichaporia RN, Joshi SK, Wadia RS.
Meningitis due to Haemophilus Influenza. J Assoc Physicians India.
1965;13:619-26.
3. Tyler KL. Chapter 28: a history of bacterial
meningitis. Handb Clin Neurol. 2010;95:417-33.
4. Rosenberg DH, Arling PA. Penicillin in the
treatment of meningitis. JAMA.1944;125: 1011-7.
5. Watt JP, Wolfson LJ, O’Brien KL, Henkle E, Deloria-Knoll
M, McCall N, et al. Burden of diseases caused by Haemophilus
influenzae type b in children younger than 5 years: global
estimates. Lancet. 2009;374:903-11.
6. Ramachandran P, Fitzwater SP, Aneja S, Verghese
VP, Kumar V, Nedunchelian K, et al. Prospective multi-centre
sentinel surveillance for Haemophilus influenzae type b & other
bacterial meningitis in Indian children. Indian J Med Res.
2013;137:712-20.
7. Nigrovic LE, Malley R, Macias CG, Kanegaye JT,
Moro-Sutherland DM, Schremmer RD, et al. Effect of antibiotic
pre-treatment on cerebrospinal fluid profiles of children with bacterial
meningitis. Pediatrics. 2008;122:726-30.
8. Singhal N, Kumar M, Kanaujia PK, Virdi JS.
MALDI-TOF mass spectrometry: an emerging technology for microbial
identification and diagnosis. Front Microbiol. 2015; 6:791. doi:
10.3389/fmicb.2015.00791. eCollection 2015
9. Brouwer MC, Tunkel AR, van de Beek D.
Epidemiology, diagnosis, and antimicrobial treatment of acute bacterial
meningitis. Clin Microbiol Rev. 2010;23:467-92.