This document discusses primary amoebic meningoencephalitis (PAM) caused by the brain-eating amoeba Naegleria fowleri. It describes two cases of PAM identified in Pakistan in 2008. Cerebrospinal fluid analysis found elevated protein and neutrophils in both cases. From April to June 2009, 11 additional PAM cases were identified and treated, but none survived. Molecular diagnosis using PCR and fluorescent probes confirmed N. fowleri infection in three patients. The increasing PAM cases in Karachi were associated with swimming in poorly chlorinated fresh water.

1. PRIMARY AMOEBIC
MENINGOENCEPHALITIS CAUSED
BY NAEGLERIA FOWLERI,
KARACHI, PAKISTAN
PRESENTED BY :DARAKHSHAN SALEEM
MS MICROBIOLOGY
FUUASTE

2. Brain-Eating Amoeba
Brain-Eating Amoeba
 So-called Brain-eating amoeba is a species
discovered in 1965.
 There are several species of Naegleria but only
the fowleri species causes human disease.
 Naegleria loves very warm water. It can
survive in water as hot as 113 degrees
Fahrenheit.

4.  N. fowleri amoebas are relatively common, they only
rarely cause brain disease.
 N. fowleri disease is known as Primary Amoebic
Meningoencephalitis (PAM).
 It occurs from zero to eight times a year, almost
always from July to September

5. SYMPTOMS :
Headache
fever
stiff neck
loss of appetite
vomiting
altered mental state
seizures
coma
There may also be hallucinations, drooping eyelid,
blurred vision, and loss of the sense of taste.

6. Two cases were studied at AKUH
In June 2008
•2 DAYS HISTORY
•HGF
•SEIZURES
•COMATOSE WITH FIXED AND DIALATED
LEFT EYE PUPIL

7. IN SEPTEMBER 2008
PATEINT WAS 25 YEARS OLD
• 24 HRS HISTORY
• FEVER
• VOMITING
• NECK RIGIDITY

8. CSF ANALYSIS IN BOTH CASES
• ELEVATED PROTEIN LEVEL
• ELEVATED NEUTROPHILIC PLEOCYTOSIS
• HYPOGLYCORRHACHIA
• GS AND LA WERE NEGATIVE

9. TILL APRIL2009 NO PAM DETECTED
IN APRIL 2009 THERE WERE 11
PATIENTS WITH PAM REFERRED TO
AKU

10. THOSE PATEINTS WERE TREATED WITH
AMPHOTERECIN B
RIFAMPIN,FLUCONAZOLNZOOLRIFA
MOITHEY WERE INCUBATED AND
VANTILATED AFTRE HOSPITAL
ADMISSION
BUT NO ONE SURVIVED

11. BECAUSE…..GS AND LA WERE
NEGATIVE
CSF CULTURED ON NON NUTRIENT AGAR ON A
LAWN OF E.COLI ,OBSERVED CULTURED
AMOEBA..WHICH LATER ON PRODUCED
FLAGELA

12. ON THE BASIS OF THESE DIAGNOSIS
WET MOUNT DONE BY LAB

13. MOLECULAR DIAGNOSIS:
IN JUNE 2009 CSF COLLECTED FROM 3
PATIENTS AND CONFIRMED THE INFECTION
BY REAL TIME PCR

14. o Separation of DNA double-stranded template
o Primer formation
o Extension of new DNA strands by a DNA
polymerase and deoxyribonucleoside
triphosphates (dNTPs)
o Other proteins involved
PRINCIPLE

15. CONT….
PRIMER NaegF192 (3’-GTG CTG AAA CCT
AGC TAT TGT AAC TCA GT-5’) AND
NaegIR344 (5’-CAC TAG AAA AAG CAA
ACC TGA AAG G-3’) WERE USED TO
AMPLIFY A 153-bp FRAGMENT

16. • THIS WAS DETECTED BY
HEXACHLOROFLUORESCEIN(HEX)
LABELED PROBE Nfowl(5’-HEX-AT AGC
AAT ATA TTC AGG GGA GCT GGG C-
BHQ1-3’)

17. PCR PERFORMANCE
2 INITIAL INCUBATIONS
AT 50 C WITH URACIL-DNA-
GLYCOSYLASE ACTIVITY FOR 2
MINS

18. 2ND
AT 95 C WITH ACTIVATION
OF PLATINIUM Taq DNA
POLYMERASE)
FLOURESECNCE WAS MEASURED

19. CONCLUSIONS
• PAM CASES INCREASE IN KARACHI AND
MOST PATIENTS WERE YOUNG HEALTHY
ADULTS .
• PAM DIAGNOSIS SHOULD BE IMPROVE BUT
THIS IS NOT THE SOLE REASON
• PAM IS ASSOCIATED WITH FRESH WATER
SWIMMING POOL HAVING POORLY
CHLORINATED WATER