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ACUTE FLACCID PARALYSIS SURVEILLANCE

Vol 39 No. 6 November 2008 1033

Correspondence: TS Saraswathy, Virology Unit,Infectious Disease Research Center, Institute forMedical Research, Jalan Pahang, 50588 KualaLumpur, Malaysia.Tel: 603 26162671; Fax: 603 26938094E-mail: [email protected]

ACUTE FLACCID PARALYSIS SURVEILLANCE:LOOKING BEYOND THE GLOBAL POLIOMYELITIS

ERADICATION INITIATIVE

TS Saraswathy1, H Nor Zahrin2, MY Apandi1, D Kurup2, J Rohani2, S Zainah1

and NS Khairullah3

1Virology Unit, Infectious Disease Research Center, Institute for Medical Research, KualaLumpur; 2Surveillance Unit, Disaese Control Division, Ministry of Health, Kuala Lumpur;

39BIO Sdn Bhd, Kuala Lumpur, Malaysia

Abstract. In 1992 surveillance of acute flaccid paralysis (AFP) cases was introduced in Malay-sia along with the establishment of a national referral laboratory at the Institute for MedicalResearch. The objective of this study was to determine the incidence, viral etiology and clinicalpicture of AFP cases below 15 years of age, reported from 2002 to 2007. Six hundred sev-enty-eight of 688 reported cases were confirmed as AFP by expert review. The clinical presen-tation of acute flaccid paralysis in these cases was diverse, the most commonly reportedbeing Guillian-Barre syndrome (32.3%). Sixty-nine viruses were isolated in this study. Theywere Sabin poliovirus (25), Echovirus (22), Cocksackie B (11), EV71 (5), Cocksackie A (1), anduntypable (5). Malaysia has been confirmed as free from wild polio since the surveillance wasestablished.

INTRODUCTION

Acute flaccid paralysis (AFP) is a clinicalsyndrome characterized by rapid onset weak-ness of limbs, often with weakness of respira-tory muscles and difficulty in swallowing. Ifuntreated, AFP may lead to death due to fail-ure of respiratory muscles.

Surveillance for AFP is an important ac-tivity of public health programs in many coun-tries because it is the key strategy used in theGlobal Polio Eradication Initiative by the WorldHealth Organization (WHO, 1988). WHO’scommitment to the Global Polio EradicationInitiative, has achieved impressive progresswith only four countries remaining endemic forpolio. These countries are India, Pakistan, Ni-

geria and Afghanistan (WHO, 2008). WHO’ssuccess has been attributed largely to a sen-sitive surveillance strategy, which requirescountries to conduct surveillance of all AFPcases using a standard case definition. Ac-tive AFP surveillance in children below 15 yearsof age is ongoing until global eradication isachieved, to monitor risk of importation of wildpolioviruses into non-polio endemic countries.With near eradication of polio, AFP surveillancehas directed the focus on other causes of thisalarming clinical syndrome.

AFP is caused by many conditions, in-cluding viral infections, notably, poliomyelitis,Guillain Barre syndrome (GBS), transversemyelitis, metabolic neuropathies and trauma.In poliomyelitis, poliovirus invasion of the an-terior horn cells of the spinal cord can mani-fest with asymmetrical weakness of limbs withno sensory symptoms or signs. Non-polioenteroviruses (NPEV) may also cause paralyticdisease, but it is usually less severe than po-liomyelitis. Most cases are due to enterovirus

SOUTHEAST ASIAN J TROP MED PUBLIC HEALTH

1034 Vol 39 No. 6 November 2008

70, enterovirus 71, coxsackie A7 and echovi-ruses (Solomon and Wilson, 2003). Manystudies have found GBS to be a leading causeof AFP (Marx et al, 2000). Also known as AcuteInflammatory Demyelinating Polyneuropathy,GBS is an immunologically mediated para-in-fectious or post-infectious process causingdamage to the lower motor neurons in theperipheral nerves or nerve roots. The underly-ing pathology, precise cellular basis or patho-physiologic mechanisms for some causes ofAFP are still not clearly understood. Accuratediagnosis of the cause for AFP has importantimplications for therapy and prognosis ofcases.

In Malaysia, the Virology Unit at the Insti-tute for Medical Research in Kuala Lumpurwas designated as the National ReferenceLaboratory for Pol iomyel it is Eradication(NRLPE) by the WHO in 1992. Under the AFPsurveillance program, clinicians are requiredto notify every case of AFP, in children underthe age of 15 years to the State Health Officeusing a standard AFP surveil lance form,wherein the relevant clinical and epidemiologi-cal data of the patient is recorded. Two freshstool specimens collected 24 hours apart andwithin 14 days of onset of paralysis are re-quired to be sent in appropriate transport con-ditions to the NRLPE. Each year a significantnumber of samples are received by our labo-ratory from hospitals throughout Malaysia forlaboratory investigation of AFP cases. In thisstudy, we analyzed AFP surveillance data fromJanuary 2002 to December 2007, to deter-mine the incidence, viral etiology and clinicalpicture of AFP, in children below 15 years.

MATERIALS AND METHODS

Samples

Between January 2002 and December2007 the NRLPE received 1,437 stool speci-mens from 688 reported AFP cases sent fromhospitals throughout Malaysia. Another 138

specimens, comprised of throat swabs andcerebrospinal fluid, were also received fromthese cases. The specimens were accompa-nied by an AFP notification form with detailsof patient personal and clinical history.

Cell cultures

All stool specimens were processed withchloroform before inoculation into RD andL20B cell lines from our laboratory stock heldin liquid nitrogen at low passage. The cultureswere grown in monolayers in 75 cm2 tissueculture flasks in Earles basal medium supple-mented with L-glutamine, Hepes buffer, pH7.2, sodium bicarbonate, 10% fetal calf se-rum, 100 IU /ml penicillin G and 100 µg/mlstreptomycin sulfate. Cells from confluent cul-tures were resuspended in maintenance me-dium and the concentration was adjusted to6 X 105 cells/ml for use in the assay. Inocu-lated cell cultures were examined daily forcytopathological effects (CPE). The WHO stan-dard protocols for pre-treatment of stoolspecimens and virus isolation by cell culturewere followed (WHO, 2004).

Microneutralization test procedure

Positive cell cultures were confirmed bymicroneutralization assay using standard WHOantisera. WHO standard enterovirus antiserumpools (0.05 ml) were distributed into wells ofa microtiter plate followed by 0.05 ml of virusdilutions from 10-1 to 10-7. The plates werecovered and incubated at 36ºC in a carbondioxide incubator for 1 hour. After incubation0.01 ml of cell culture suspension, containingapproximately 1.5 x 105 were added to allwells. The antiserum pools that prevent thedevelopment of CPE indicated the identity ofthe virus isolated. Poliovirus isolates were sentto the Victorian Infectious Disease ReferenceLaboratory (VIDRL) in Melbourne, Australia forfurther identification and intratypic differentia-tion.

Controls

The controls incorporated in the assay



included cell control to check for normal cellmorphology and virus control to detect virusinfectivity with complete CPE. Back titrationwas included to confirm that the amount ofvirus used in the assay was within an accept-able range.

AFP clinical picture

All AFP reported cases were followed upfor 60 days to ascertain residual paralysis. Thisactivity was monitored by the SurveillanceUnit, Disease Control Division of the Ministryof Health by communication with cliniciansattending to the AFP cases and from monthlynotification data obtained from State HealthDepartments. Clinical assessment of all re-ported AFP cases were also reviewed at theExpert Polio Review Meetings.

RESULTS

During the period of this study, 688 AFPcases below age 15 years were reported toour laboratory. Following review by the PolioExpert Group, 10 cases were classified asnon-AFP by reassessment of clinical findings

and only 678 were confirmed as AFP. The an-nual incidence of reported AFP cases andspecimens received are shown in Table 1.This may not indicate the actual incidence ofAFP, as many cases may not have been re-ported to our laboratory for viral investigation.Since AFP notification is part of the poliomy-elitis eradication exercise some cases mayhave been omitted when a definite diagnosisby history, for example trauma, did not indi-cate the need for laboratory investigation atthe time of clinical assessment. Enteroviruseswere found in 69 cases (4.9%) out of 1,416stool specimens received from these AFPcases. Polioviruses was found in 25 cases(36.2%). All these polioviruses were confirmedas vaccine related Sabin-like strains by theVIDRL, the WHO Reference Laboratory inMelbourne, Australia. The remaining 44 virusisolates were non-polio enteroviruses. Theidentification and distribution of these virusesare shown in Figs 1 and 2. Echoviruses ac-counted for the majority of non-enterovirusisolated from the AFP cases in this study(50%), followed by coxsackie B viruses (25%)

Year Reported Confirmed Incidence rate Stool EnterovirusesAFP cases AFP cases (Per 100,000) specimens isolated

2002 107 107 1.3 222 10 PVSL11 NPEV

2003 100 98 1.18 191 6 PVSL10 NPEV

2004 120 114 1.35 243 8 NPEV2005 138 137 1.61 292 5 PVSL

5 NPEV2006 116 116 1.34 239 4 PVSL

4 NPEV2007 107 106 1.21 229 6 NPEVTotal 688 678 Range1.2~1.6 1,416 69 Enteroviruses

Table 1Annual reported AFP cases in children below 15 years of age and specimens received for

laboratory investigation.

PVSL = Vaccine related Sabin-like polioviruses; NPEV = Non-polio enteroviruses



and enterovirus 71 (11.4%). Table 2 shows theclinical diagnosis of the 678 confirmed AFPcases included in the study. For convenienceof presentation, some of less frequent clinicalpresentations have been grouped using aneuro-anatomical approach. Fourteen of the

678 AFP cases did not have a specific clinicaldiagnosis other than a report of AFP. Whenthe available clinical data was inadequate,the AFP cases were followed up with the at-tending pediatrician in charge to get a full casereport. These cases either had adequate

Table 2Clinical diagnosis of confirmed AFP cases from 2002 to 2007.

ConditionGuillain-Barre syndrome (Including acute and chronic inflammatory 219 32.3 demyelinating polyneuropathy, post-infection demyelination)Transverse myelitis 36 5.3Meningitis / meningoencephalitis 85 12.5Viral encephalitis 26 3.8Vaccine associated poliomyelitis 3 0.4Enteroviral parainfectious flaccid paralysis 2 0.3Critical illness (acute excerbation of bronchial asthma due 4 0.6 to pneumonia, upper respiratory tract infection with hyperemesis, bronchopneumonia and diarrhea)

Site / Factor (Contition)Brain (encephalomyelitis / myelitis , cervical cord tumor, cerebral tumor, 114 16.8 cerebral atropy acute lymphoblastic leukemia, rheumatic chorea, recurrent transient ischemic attack, cerebral ataxia, right sided thalamic infarct, extrapyramidal cerebral palsy, partial seizures, Moya-moya disease with intracranial hemorrhage, cerebral edema due to fall, acute demyelinating encephalomyelitis (ADEM, multiple sclerosis, cerebrovascular accident, mitochondrial myopathy)

Musculoskeletal (viral myositis, septic arthritis, transient synovitis, 62 9.1 musculodystrophy, viral myalgia)

Nerve (peripheral neuropathy, traumatic neuritis, post-infective neuritis, 23 3.4 neuropraxia secondary to infection, vasculitis, Miller-Fisher Syndrome)

Neuropathies associated with systemic / metabolic disorders 63 9.3 (hypokalemic periodic paralysis, nephrotic syndrome, porphyric crisis, renal tubular acidosis, acute urinary retention) hypokalemia due to diarrhea and malnutrition, Bartter’s Syndrome, Inborn error of metabolism

Spinal cord (spinal cord tumor, spinal cord injury, extradural compression, 16 2.4 syringomyelia, spinal dural AV fistula

Neuromuscular junction (myasthenia gravis, botulism, jelly fish sting) 6 0.9

Nerve root (brachial plexitis) 5 0.7

Not specified 14 2.1

Total 678 100

Clinical diagnosis N %

ADEM- acute demyelinating encephalomyelitis



stools which were confirmed as non-polio bylaboratory investigations or fully recoveredmotor function. Since this AFP reporting sys-tem was based on the national polio eradica-tion program, the Expert Review Group de-termined these cases as non-polio AFP.

DISCUSSION

The AFP surveillance system provides asensitive tool for investigating AFP cases inchildren, with careful clinical evaluation of thedifferential diagnosis and expert review ofcases. Accurate diagnosis requires a preciseknowledge of the etiology and underlyingpathophysiology. Analysis of the clinical find-ings and diagnosis reported for the 678 casesin this study showed that the underlyingcauses were diverse.

No wild poliovirus was isolated during thisperiod. Malaysia has been free of wild poliovi-rus circulation since the mid 1980s and is oneof the countries listed in the Western PacificRegion as nonendemic for poliovirus. The lastmajor outbreak of poliomyelitis in Malaysiaoccurred in 1977 with 121 cases, including 4deaths (IMR, 1977). The number of poliomy-elitis cases decreased dramatically from 1978following an effective National Oral Polio Vac-cine Immunization Program introduced in1972. However, three cases of poliomyelitiswere reported in 1992 probably due to impor-tation of wild poliovirus (IMR, 1992/1993).Since 1993, no wild poliovirus has been iden-tified; our success being attributed to goodimmunization coverage and an effective AFPsurveillance system. Three of the cases in thestudy were identified as possible vaccine as-sociated paralytic poliomyelitis (VAPP). VAPPis diagnosed when a vaccine strain of poliovi-rus is isolated from the stool of an AFP case,where oral polio vaccine was given in the last30 days preceeding onset of paralysis. Sabin-like poliovirus type 1 and type 3 were isolatedfrom these cases.

With near eradication of poliomyelitis, vi-ruses other than polioviruses have been re-ported to cause AFP (Solomon and Wilson,2003). Among these viruses are echoviruses,coxsackie viruses and enterovirus 71, whichare also members of the enterovirus genus.Japanese encephalitis virus and West Nile vi-rus have also been reported to cause this syn-drome (Solomon et al, 1998; Glass et al,2002). In our study, 44 non-polio enteroviruses(NPEV) were isolated; the most predominant,as seen from Fig 1, were the echoviruses.Further investigation by immunofluorescentmethod using specific antisera pools showedthat these were echovirus types 3, 6 and 11.The clinical presentations in these cases in-cluded aseptic meningitis, meningoencepha-litis, viral encephalitis and GBS. Five out of44 NPEVs were enterovirus 71 and the clini-

1

5

8

11

22

25

0 5 10 15 20 25 30

CoxsackieA

EV71

Untypable

Coxsackie B

Echovirus

Sabin poliovirus

Sabin type 1Sabin type 2Sabin type 3

Fig 2–Distribution of Sabin-like poliovirus isolatedfrom AFP cases: Sabin type 3 (43.8%) > Sabintype 1 (34.3% ) > Sabin type 2 (21.9%).

Fig 1–Distribution of Enteroviruses isolated fromAFP cases.



cal presentations in these cases includedacute encephalopathy and enterovira lmonoplegia. EV 71 has been reported in coun-tries of the Asia Pacific region to cause largeoutbreaks of hand foot and mouth disease(HFMD) (Cardosa et al, 2003). In MalaysiaHFMD outbreaks occurred in 1997 and in1999, where EV71 was reported to cause theneurological complications in some childrenwhich included acute flaccid paralysis andbrain stem encephalitis (Chan et al, 2000).

Gullain Barre Syndrome (GBS) accountedfor 32.3% of AFP cases in this study with anoverall annual incidence rate of 0.42. In theabsence of wild poliovirus induced poliomy-elitis, GBS is the most common cause of AFPreported in many parts of the world, account-ing for over 50% in many industrialized anddeveloping countries (Marx et al, 2000). It isinteresting to note that in a previous study ofAFP surveillance in Malaysia, data from 1997to 2001 showed that GBS was found in 30.2%of AFP cases, with an annual incidence of0.36% (Hussain et al, 2004). The majority ofGBS cases in the AFP surveillance were diag-nosed based on typical clinical features, suchas progressive ascending and symmetricalparalysis of the limbs with or without cere-brospinal fluid abnormalities. Not all caseswere confirmed by nerve conduction studies,which can only be carried out at regional cen-ters where these facilities and pediatric neu-rologists are available. Hussain et al (2004)found that central nervous system infectionswere more common in Malaysia than othercountries.

Among other significant causes of AFPreported during this period were transversemyelitis (5.3%) and neuropathies occurringfrom metabolic disorders (9.3%). Metabolicdisorders included acute hypokalemic periodicparalysis, hypokalemia from malnutrition andacute gastroenteritis as well as porphyria andrenal tubular acidosis. As seen in Table 2, theother conditions and causes attributed to AFP

during this surveillance period were broad.These conditions included central nervoussystem disorders, infantile botulism, jelly fishstings, and weakness associated with criticalillnesses, such as acute excerbation of bron-chial asthma secondary to pneumonia.

In conclusion, despite the great reduc-tion in the number of poliomyelitis cases fol-lowing the Global WHO Poliomyelitis Cam-paign, AFP continues to be an important neu-rological presentation in children. The list ofunderlying causes of AFP is broad and com-plex. Clinicians need to have a detailed know-ledge of the differential diagnosis to ensureeffective and timely management. The currentWHO AFP surveillance provides a sensitivetool for investigating AFP cases with carefulclinical evaluation of the differential diagnosisand expert review of cases. It is therefore cru-cial that AFP surveillance be conducted evenin the absence of wild poliovirus transmission.

ACKNOWLEDGEMENTS

The authors would like to thank the Di-rector General of the Ministry of Health, Ma-laysia and the Director of the Institute for Medi-cal Research for permission to publish thispaper. Special thanks are extended to thehealth staff of the Ministry of Health, Malaysiawho contribute to the success of the acuteflaccid paralysis surveillance program.

REFERENCES

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