Retrospective survey for sialidase activity in Mycoplasma pneumoniae isolates from cases of community-acquired pneumonia
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Title: Retrospective survey for sialidase activity in Mycoplasma pneumoniae isolates from cases of community-acquired pneumonia
Series Title: BMC Research Notes 2011, 4:195
Physical Description: Journal Article
Creator: Brown, Daniel R.
May, Meghan
Publisher: BioMed Central
Place of Publication: United Kingdom
Publication Date: 15 June 2011
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Abstract: Background: Sialidase is a well-known virulence factor of other respiratory pathogens, but was only recently documented to occur in some species of Mycoplasma. The sialidase activity expressed can vary quantitatively among strains within a species of mycoplasma, from undetectable to amounts that correlate positively with strain virulence. Very few isolates of Mycoplasma pneumoniae had ever been examined for sialidase activity, so it was unknown whether sialidase may contribute to diseases involving this species. Findings: No sialidase activity was detected by spectrofluorometric assay of 15 laboratory strains and 91 clinical isolates of M. pneumoniae banked over many years from patients having radiologically-confirmed, uncomplicated community-acquired pneumonia. Conclusions: The annotated genome of strain M129 (GenBank NC_000912, ATCC 29342), also isolated from a patient with pneumonia, accurately represents the absence of sialidase genes from strains of M. pneumoniae typically associated with uncomplicated community-acquired pneumonia. A possible involvement of sialidase in neurologic or other extra-respiratory manifestations of M. pneumoniae mycoplasmosis remains to be investigated.
Acquisition: Collected for University of Florida's Institutional Repository by the UFIR Self-Submittal tool. Submitted by Dan Brown.
Publication Status: Published
Funding: Publication of this article was funded in part by the University of Florida Open-Access Publishing Fund.
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Source Institution: University of Florida Institutional Repository
Holding Location: University of Florida
Rights Management: All rights reserved by the submitter.
System ID: IR00000563:00001

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SHORTREPORTOpenAccess Retrospectivesurveyforsialidaseactivityin Mycoplasmapneumoniae isolatesfromcases ofcommunity-acquiredpneumonia MeghanMay 1 † andDanielRBrown 2* † Abstract Background: Sialidaseisawell-knownvirulencefactorofotherrespiratorypathogens,butwasonlyrecently documentedtooccurinsomespeciesof Mycoplasma .Thesialidaseactivityexpressedcanvaryquantitatively amongstrainswithinaspeciesofmycoplasma,fromundetectabletoamountsthatcorrelatepositivelywithstrain virulence.Veryfewisolatesof Mycoplasmapneumoniae hadeverbeenexaminedforsialidaseactivity,soitwas unknownwhethersialidasemaycontributetodiseasesinvolvingthisspecies. Findings: Nosialidaseactivitywasdetectedbyspectrofluorometricassayof15laboratorystrainsand91clinical isolatesof M.pneumoniae bankedovermanyyearsfrompatientshavingradiologically-confirmed,uncomplicated community-acquiredpneumonia. Conclusions: TheannotatedgenomeofstrainM129(GenBankNC_000912,ATCC29342),alsoisolatedfroma patientwithpneumonia,accuratelyrepresentstheabsenceofsialidasegenesfromstrainsof M.pneumoniae typicallyassociatedwithuncomplicatedcommunity-acquiredpneumonia.Apossibleinvolvementofsialidasein neurologicorotherextra-respiratorymanifestationsof M.pneumoniae mycoplasmosisremainstobeinvestigated. Findings Researchobjective Mycoplasmapneumoniae isprimarilyassociatedwith interstitialpneumonitis,tracheobronchitis,desquamative bronchitisandpharyngitis,collectivelyreferredtoasprimaryatypicalpneumonia(PAP)[1,2].Mycoplasmosis accountsfor20-30%ofcommunity-acquiredpneumonia(CAP)cases,constitutingsignificantdiseaseandeconomicburdensinNorthAmericaandWesternEurope. FollowingitsinitialassociationwithPAP,otherdiseases involving M.pneumoniae invasionofnon-respiratory tissueswerereported[Tabl e1].Neurologic,dermal, hemotropic,cardiac,arthritic,hepatic,pancreatic, nephritic,andmusculoskeletalpathologieshavebeen described[2-12];manyofthosediseasesoccurredsecondarilytoPAP,eitherthroughdisseminationof M.pneumoniae fromtherespiratorytract,orfollowing associatedautoimmunedisorders[3].Primaryextrarespiratory M.pneumoniae infectionsintheabsenceof PAP,includingmeningoencephalitis,hepatitis,and pancreatitis,havealsobeendescribed[13,14],and M.pneumoniae hasbeenisolatedfromtheurogenital tractintheabsenceofclinicalsigns. Themajorityofstudiesonclinicalaspectsof M.pneumoniae mycoplasmosisaddressthediagnosis,treatment andpreventionofCAP,while factorspredisposingto extra-respiratorydiseasesr emainvirtuallyunexplored. Extracellular “ spreadingfactors ” likesialidasearewellknownvirulencedeterminantsofotherpathogenic microorganisms,andaretargetsforchemotherapyin diseasessuchasinfluenza.Sialidaseisassociatedwith systemicdisseminationduringinfectionwithmanybacterialspecies,mostnotably Streptococcuspneumoniae and Clostridiumperfringens ,butsuchglycosidaseswere onlyrecentlydocumentedtooccurincertainspeciesof Mycoplasma .Thesialidaseactivityexpressedbymycoplasmascanvarysignificantlyamongstrainswithina species,fromundetectabletoamountsthatcorrelate positivelywithstrainviru lence.Veryfewisolatesof *Correspondence:drbrown@ufl.edu † Contributedequally 2 DepartmentofInfectiousDiseasesandPathology,CollegeofVeterinary Medicine,UniversityofFlorida,GainesvilleFL,USA Fulllistofauthorinformationisavailableattheendofthearticle MayandBrown BMCResearchNotes 2011, 4 :195 http://www.biomedcentral.com/1756-0500/4/195 2011Brownetal;licenseeBioMedCentralLtd.ThisisanopenaccessarticledistributedunderthetermsoftheCreativeCommons AttributionLicense(http://creativecommons.org/licenses/by/2.0),whichpermitsunrestricteduse,distribution,andreproductionin anymedium,providedtheoriginalworkisproperlycited.

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M.pneumoniae havebeenexaminedforsialidaseactivity,soitwasunknownwhethersialidasemaycontribute eithertoPAPortoextra-respiratorydiseasesinvolving thisspecies.Toestablisht hebaselinefrequencyofits occurrancein M.pneumoniae ,weconductedaretrospectivesurveyforsialidase activityinclinicalisolates associatedwithrespiratorymycoplasmosis.MethodsFifteenlaboratorystrainsof M.pneumoniae ,including thewell-knownvirulentstrainPI1428(AmericanType CultureCollectionacce ssionnumber29085,froma patientwithPAP),SAD03,SAD05,TW11-4,1RCH, 9RCH,11-61,104.2,142-48,256-8,541-6,541-29,1161, 1311and15531,plus91clinicalisolatesfromcasesof radiologically-confirmedin terstitialpneumoniawere tested.Thede-identifiedclinicalisolateshadbeen bankedoveraperiodofmanyyearsfrompatientsdiagnosedwithCAPatvariouslocationsintheUS.They wereisolatedfromfourdistinctsitesintherespiratory tract:61werefromthroatswabs,4fromnasalswabs,17 fromsputum,and9frombronchoalveolarlavagefluid. Thelatterwereconsideredb yclinicianstobeinvasive becausetheywerefrompatientshospitalizedwithsevere pneumonia.Itwasunknownwhetheranyofthepatients hadbeendiagnosedashavingpneumoniaattributable solelyto M.pneumoniae ,orhadbeentreatedempiricallywithviralneuraminidaseinhibitors. Frozenstocksofmycoplasmawerepassagedonetime inSP-4mediumsupplementedwith10%w/vglucose and15%v/vfetalbovineserumat37Cinambientair. Sialidaseactivityof M.pneumoniae cellssuspendedin conditionedmediumwasassessedusingthefluorogenic substrate2 ’ -(4-methylumbelliferyl)a -DN -acetylneuraminicacid(MUAN)aspreviouslydescribed[15].Briefly, 50 lofstationaryphasecultureswereaddedto50 l of0.35%w/vMUANin30mMsodiumacetatebuffer, pH4.5.Following60minincubationatroomtemperature,fluorescenceat450nmwasquantitatedusinga spectrofluorometer(excitationat350nm;cutofffilterat 420nm).Thepositivecontrolswere Mycoplasmaalligatoris strainA21JP2T,whichexpressesacell-associated sialidase[16],andTypeVIsialidasepurifiedfrom Clostridiumperfringens .Thenegativecontrolswere Mycoplasmacrocodyli strainMP145T,whichdoesnotexpress anysialidase,andfreshculturemedium.ResultsNosialidaseactivitywasdetectedinanyspecimen,thus therewasnoevidencethattheenzymecontributesto PAPinvolving M.pneumoniae .However,theundocumentedclinicalsignificanceof M.pneumoniae isolation fromthesepatientsisarecognizedweaknessofthis study.Itseemsunlikelythatpotentialempirictreatment withviralneuraminidaseinhibitorsstrictlyprecluded isolationofsialidase-positivestrainsof M.pneumoniae Table1Diverseoutcomesassociatedwith Mycoplasmapneumoniae infectiona Aural Dermal Musculoskeletal  otitisexterna  bullousexanthema  reactivearthritis  otitismedia  cutaneousvasculitis  rhabdomyolysis  myringitis  erythemanodosum  septicarthritis Cardiac  Stevens-Johnsonsyndrome Ocular  arrhythmia  ulcerativestomatitis  conjunctivitis  myocarditis  urticaria  iritis  pericardialeffusion  vesicularrash  retinitis  pericarditis Gastrointestinal  uveitis Centralnervoussystem  hepatitis Renal  ataxia  pancreatitis  glomerulonephritis  Bell ’ spalsy Hematologic  IgAnephropathy  choreoathetosis  aplasticanemia  interstitialnephritis  encephalitis  hemolyticanemia Respiratory  encephalomyelitis  hemophagocytosis  primaryatypicalpneumonia  Guillain-Barre-syndrome  intravascularcoagulation  meningoencephalitis  lymphadenopathy  meningitis  neutropenia  paralysis  neutrophilia  polyradiculitis  septicemia  psychosis  thrombocytopeniaaTheprimarydiseasestategeneratedby M.pneumoniae infectionisPAP,althoughmanyadditionalclinicalmanifestationshavebeenreported[2]. Mycoplasmosisdueto M.pneumoniae affectsnumerousorgansystemsbygeneratingprimarylesionsinnon-respiratorytissuesorsecondarylesionsfollowing presumedhematogenousspreadfromtherespiratorytract,orbycontributingtoautoimmune-relateddisorders.Differencesinbacterialorhostphe notypesthat explainthediverseclinicaloutcomesremainessentiallyunknown.MayandBrown BMCResearchNotes 2011, 4 :195 http://www.biomedcentral.com/1756-0500/4/195 Page2of3

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fromanyofthesepatients,butthisremainstobedefinitivelyestablished. Theabilityofcertain M.pneumoniae strainstocause extra-respiratorymycoplasmo sismayrequireadditional cellularfunctions,whichcanbegeneticallyacquired in vivo fromthepolymicrobialcommunitywithinthe humanhost.Thepresenceofbacterialspreadingfactors suchassialidasemightexplainthedifferencesinclinical manifestationsofinfectionwithcertainstrainsof M. pneumoniae .Sialidaseisaprominentcandidateforthis becauseofitswell-documentedtendencytobehorizontallytransferredamongbacteria[17].Inaddition,quantitativevariationofsialida seactivityamongstrainsof otherpathogenicmycoplasmasrangesfromundetectable toextremelyhighinsignificantcorrelationwithstrain invasiveness[18].Apossiblei nvolvementofsialidasein neurologicorothermanifestationsofinvasivedisease remainstobeinvestigated,butabankofthoseless commonspecimenswillberequiredtodetermine whetherPAP-associatedstrainsarelesslikelytoexpress sialidaseactivitythanstrainsassociatedwithotheroutcomesof M.pneumoniae mycoplasmosis.Weconclude thattheannotatedgenomeofthewell-knownvirulent strainM129(GenBankacce ssionnumberNC_000912), alsoisolatedfromapatientwithpneumonia(American TypeCultureCollectionaccessionnumber29342),accuratelyrepresentstheabsenceofsialidasegenesfrom strainsof M.pneumoniae typicallyassociatedwith uncomplicatedPAP. Acknowledgements Theclinicalisolatesof M.pneumoniae wereobtainedfromDr.KenB.Waites oftheUniversityofAlabamaatBirmingham,BirminghamAL,USA,and handledinaccordancewithaprotocolapprovedbytheUniversityof Florida ’ sInstitutionalReviewBoard(UFIRB#89-2009).WethankDina Demcovitz,AmyRatliffandLynnDuffyfortechnicalassistance.Thiswork wassupportedbyPublicHealthServicegrant1R01GM076584fromtheUS NationalInstituteofGeneralMedicalSciences(DRB).Publicationofthis articlewasfundedinpartbytheUniversityofFloridaOpen-Access PublishingFund. Authordetails 1 DepartmentofBiologicalSciences,TowsonUniversity,TowsonMD,USA. 2 DepartmentofInfectiousDiseasesandPathology,CollegeofVeterinary Medicine,UniversityofFlorida,GainesvilleFL,USA. Authors ’ contributions MMjointlyconceivedthestudyandparticipatedinstudydesign;collected, analyzedandinterpretedthedata;anddraftedthemanuscript.DRBjointly conceivedthestudyandparticipatedinstudydesign;reviewedthedata analysisandinterpretation;andrevisedthemanuscriptforimportant intellectualcontent.Bothauthorshavereadandapprovedthefinalversion ofthemanuscript. Competinginterests Theauthorsdeclarethattheyhavenocompetinginterests. Received:5January2011Accepted:15June2011 Published:15June2011 References 1.KrauseDC,Taylor-RobinsonD: Mycoplasmaswhichinfecthumans. In Mycoplasmas:MolecularBiologyandPathogenesis. Editedby:ManiloffJ. Washington,DC,AmericanSocietyforMicrobiologyPress;1992:417-444. 2.WaitesK,TalkingtonD: Newdevelopmentsinhumandiseasesdueto mycoplasmas. In Mycoplasmas:MolecularBiology,Pathogenicity,and StrategiesforControl. Editedby:BlanchardA,BrowningGF.Norfolk,Horizon Bioscience;2005:289-354. 3.AtkinsonTP,BalishMF,WaitesKB: Epidemiology,clinicalmanifestations, pathogenesisandlaboratorydetectionof Mycoplasmapneumoniae infections. FEMSMicrobiolRev 2008, 32 :956-973. 4.FreemanR,McMahonM: Acutepancreatitisandserologicalevidenceof infectionwith Mycoplasmapneumoniae Gut 1978, 19 :367-370. 5.Arav-BogerR,AssiaA,SpirerZ,BujanoverY,ReifS: Cholestatichepatitisas amainmanifestationof Mycoplasmapneumoniae infection. JPediatr GastroenterolNutr 1995, 21 :459-460. 6.DaxboeckF,BlackyA,SeidlR,KrauseR,AssadianO: Diagnosis,treatment, andprognosisof Mycoplasmapneumoniae childhoodencephalitis: systematicreviewof58cases. JChildNeurol 2004, 19 :865-871. 7.al-AbassiA: Acutepancreatitisassociatedwith Mycoplasmapneumoniae : acasereportofmisseddiagnosis. MedPrincPract 2002, 11 :112-115. 8.VicP,BlondinG,BlayoM,FinelE,DaaboulM,QueinnecC,BroussineL: Pancratiteaiguetinfection Mycoplasmapneumoniae [Acute pancreatitisand Mycoplasmapneumoniae infection]. ArchPediatr 2004, 11 :154. 9.DelCastilloM,D ’ GianoC,GoicoecheaM,MorelloF,SalsamendiP,MoraA: Mycoplasmapneumoniae meningoencephalitisinayoungadult. Medicina(BAires) 2005, 65 :338-340. 10.StammB,MoschopulosM,HungerbuehlerH,GuarnerJ,GenrichG,ZakiS: Neuroinvasionby Mycoplasmapneumoniae inacutedisseminated encephalomyelitis. EmergInfectDis 2008, 14 :641-643. 11.WalterND,GrantGB,BandyU,AlexanderNE,WinchellJM,JordanHT, SejvarJJ,HicksLA,GiffordDR,AlexanderNT,ThurmanKA,SchwartzSB, DennehyPH,KhetsurianiN,FieldsBS,DillonMT,ErdmanDD,WhitneyCG, MooreMR: Communityoutbreakof Mycoplasmapneumoniae infection: school-basedclusterofneurologicdiseaseassociatedwithhousehold transmissionofrespiratoryillness. JInfectDis 2008, 198 :1365-1374. 12.LeeS,YangS,ChangC,YehH,ChowW: Mycoplasmapneumoniae associatedacutehepatitisinanadultpatientwithoutlunginfection. J ChinMedAssoc 2009, 72 :204-206. 13.NaritaM,YamadaS,NakayamaT,SawadaH,NakajimaM,SageshimaS: Two casesoflymphadenopathywithliverdysfunctiondueto Mycoplasma pneumoniae infectionwithmycoplasmalbacteraemiawithout pneumonia. JInfect 2001, 42 :154-156. 14.Romero-GmezM,OteroMA,Snchez-MuozD,Ramrez-ArcosM, LarraonaJL,SurezGarcaE,Vargas-RomeroJ: Acutehepatitisdueto Mycoplasmapneumoniae infectionwithoutlunginvolvementinadult patients. JHepatol 2006, 44 :827-828. 15.MayM,BrownDR: Secretedsialidaseactivityofcaninemycoplasmas. Vet Microbiol 2009, 137 :380-383. 16.BrownDR,ZacherLA,FarmerieWG: Spreadingfactorsof Mycoplasma alligatoris ,aflesh-eatingmycoplasma. JBacteriol 2004, 186 :3922-3927. 17.RoggentinP,SchauerR,HoyerL,VimrE: Thesialidasesuperfamilyandits spreadbyhorizontalgenetransfer. MolMicrobiol 1993, 9 :915-921. 18.MayM,KlevenSH,BrownDR: Sialidaseactivityin Mycoplasmasynoviae AvianDis 2007, 51 :829-833. doi:10.1186/1756-0500-4-195 Citethisarticleas: MayandBrown: Retrospectivesurveyforsialidase activityin Mycoplasmapneumoniae isolatesfromcasesofcommunityacquiredpneumonia. BMCResearchNotes 2011 4 :195. MayandBrown BMCResearchNotes 2011, 4 :195 http://www.biomedcentral.com/1756-0500/4/195 Page3of3


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