Demystifying Multiple Neisseria gonorrhoeae Testing Methods and Results

Transcription

1 INFECTIOUS DISEASES MARCH 2019 Demystifying Multiple Neisseria gnrrheae Testing Methds and Results Fr many years, culture has been the standard test fr the identificatin f Neisseria gnrrheae. The develpment f nnculture methds, such as nucleic acid amplificatin tests (NAATs), allws fr direct mlecular detectin with greater sensitivity, specificity and ease f specimen transprt. The CDC Recmmendatins fr Labratry-Based Detectin f Chlamydia trachmatis and Neisseria gnrrheae 1 utlines the recmmendatin fr the use f NAATs t detect N. gnrrheae based n the imprvements t verall sensitivity, specificity and specimen transprt cnditins cmpared t ther methds. With additinal mnitring fr N. gnrrheae antimicrbial resistance, there is an increased chance that a specimen submitted fr N. gnrrheae screening r diagnsis may be subjected t testing by multiple methds. Fr example, specimens submitted fr N. gnrrheae screening may receive a NAAT t diagnse the infectin and may be sent fr culture and antimicrbial susceptibility testing fr surveillance. If the NAAT and culture have discrepant results, it can be cnfusing t interpret. Hwever, as with all testing, discrepancies between test methds can ccur, and it is imprtant fr labratrians and clinicians t understand test limitatins. Misdiagnsis based n misinterpretatin f labratry results culd lead t untreated disease prgressin, transmissin f disease due t unknwn diagnsis, r unnecessary and inapprpriate treatment. While there are additinal diagnstic methds fr N. gnrrheae, they are nt cvered in this dcument as they are used rarely in the US. The table and scenaris cntained in this dcument have been designed t imprve understanding and t help trublesht discrepant results between culture and NAAT testing fr N. gnrrheae. Table 1: Imprtant Aspects f the Cmmn N. gnrrheae Testing Methdlgies Categry Culture NAAT Reference Test Sensitivity* Specificity* Specimen Cllectin and Transprt Histrically, still required fr antibitic susceptibility testing and sequencing Recmmended by CDC and APHL fr labratry diagnsis f urgenital and extragenital N. gnrrheae infectins 1 Less sensitive than nucleic acid amplificatin testing Mst sensitive diagnstic methd fr N. (NAAT) 1 gnrrheae Other rganisms (such as Neisseria cinerea) may be misidentified as N. gnrrheae unless additinal testing is perfrmed 1 Specimen cllectin technique, strage cnditins, and transprtatin envirnment may impact viability 7 Specimen cllectin swabs, such as thse with wd shafts and cttn tips, may be inhibitry t grwth 1 Cnsult with labratry fr apprpriate specimen cllectin methd and transprt cnditins Sme cmmercial NAATs might detect nngncccal Neisseria species and ther cmmensals 1 Specimen cllectin kit typically cntains nucleic acid stabilizers resulting in lnger stability PCR can be inhibited by certain substances which culd be in the specimen cllectin swab 8 Specimen cllectin (swabs) and transprt cnditins vary by manufacturer-refer t package insert fr recmmended practices APHL N. gnrrheae Testing Methdlgies Fact Sheet 1

2 Categry Culture NAAT Viable Organisms (N. gnrrheae) Viable rganisms are required fr successful N. gnrrheae has demanding nutritinal and envirnmental grwth requirements, e.g., CO 2 - candle jar Fr maximum recvery f N. gnrrheae, bth selective and nnselective media shuld be used Specimen Types Culture can be perfrmed n all specimen types Testing Fr Sexual Assault Antimicrbial Susceptibility Testing Cnsult the labratry that is perfrming testing t cnfirm specimen types that are accepted Preferred methd fr diagnsing bys, detecting infectin in extragenital specimens, as well as frm vaginal swabs in girls and bth urine and urethral swabs in bys. 1,9 Critical fr detectin and mnitring antimicrbial resistance Viable rganisms nt required fr detectin f pathgen. Methd detects nucleic acids which may persist fllwing apprpriate antimicrbial therapy Limited t specimen types that are FDAcleared: urine, endcervical, urethral and vaginal swabs. First catch urine specimen in females may detect up t 10% fewer infectins as cmpared t vaginal and endcervical swab samples Cnsult the labratry that is perfrming testing t cnfirm specimen types that are accepted. Nt all labratries are validated and able t test extra-genital specimens. NAAT can be used t test vaginal and urine specimens frm girls but shuld be dne after cnsultatin with an expert t ensure prper testing and test interpretatin. Data are insufficient t recmmend use f NAAT in bys and extragenital sites frm either gender. 1,9 N FDA-cleared NAAT fr detectin f antibitic resistance Test f Cure- If treated with an alternative regimen, the patient shuld return 14 days after treatment fr a test Pharyngeal f cure using either culture r NAAT. 9 If the NAAT is psitive, effrts shuld be made t perfrm Gnrrhea a cnfirmatry culture befre retreatment. All psitive cultures fr test-f-cure shuld underg antimicrbial susceptibility testing. 9 *Sensitivity and Specificity vary by methd, cnsult manufacturer package insert fr direct cmparisns f each assay with Frequently Asked Questins 1. A physician requested testing fr N. gnrrheae and the culture came back negative but the NAAT was psitive why wuld this be the case? Answer: While NAAT is mre sensitive than culture, there are several factrs that culd accunt fr the discrdant test results in this case. Befre we address the differences in hw the different methds detect N. gnrrheae, it is imprtant t knw that in mst cases in rder t perfrm bth culture and NAAT tw different samples (swabs and/r urine) are cllected. Many f the explanatins f discrdance culd be traced back t this detail. Fr example, even if the exact same anatmic lcatin is swabbed, there culd be inadequate sampling with ne f the swabs that culd impact the testing. Alternatively, it is pssible that the NAAT was perfrmed n a urine specimen and the culture was perfrmed n a swab, leading t discrdant results merely due t the specimen type used. In the bullets belw and in the subsequent examples, we prvide ther ptential explanatins fr discrdance. Please review the list and cnsider using it in future cases f discrdance yu may encunter. The mst prbable explanatin is that culture is less sensitive than NAAT. 1 The patient culd have been infected with N. gnrrheae which was detected by the NAAT but it was nt detected by culture due t the lwer sensitivity. APHL N. gnrrheae Testing Methdlgies Fact Sheet 2

3 The site f cllectin may impact the results. Fr example, pharyngeal samples may nt grw as well in culture but there was enugh nucleic acid present fr the NAAT methd t detect the N. gnrrheae. The specimen cllectin swab itself might nt have been apprpriate fr the methd (either it was inhibitry t the grwth 1 and/r there was nt enugh f the rganism n the cllectin swab). The rganism may n lnger be viable but the nucleic acid frm the rganisms was still detected by the NAAT. Sme reasns the rganism may n lnger be viable include: ineffective transprt media 7 transprt delays inapprpriate specimen transprt cnditins the patient having received antibitic therapy prir t sample cllectin If the NAAT is perfrmed within a certain time frame (refer t manufacturer s package insert fr exact time frame) f receiving antibitic therapy it can still detect residual nucleic acid despite successful treatment. This means that the patient is n lnger infected and n viable rganism can be cultured but the NAAT is psitive because nucleic acid is still present. T prevent this utcme refer t the manufacture s package insert fr the intended use and limitatins f the assay. The persn culd have been truly negative fr N. gnrrheae but the NAAT detected a nngncccal Neisseria species. This is cnsidered a false-psitive result because the persn was nt infected with N. gnrrheae, but the NAAT result indicates that the patient is psitive fr N. gnrrheae. This is a limitatin f the assay design. See the manufacturer s package insert fr assay-specific limitatins. Being aware f the ptential reasns fr the discrdant results and explring whether any f these scenaris culd have cntributed t the result is an imprtant next step in determining whether the patient may have gnrrhea. The clinician s decisin t initiate treatment is dependent n many factrs including labratry findings, any ptential fllw-up testing, clinical examinatin and risk factrs, public health department case definitins and the CDC STD Treatment Guidelines We just received test results fr a patient suspected f having gnrrhea the NAAT was negative but the culture was psitive, shuld we be cncerned? Answer: Discrepant results are nt uncmmn. While we nrmally think f the NAAT assay as being mre sensitive, there are ptential explanatins fr these results. As in the example abve, let us examine sme f the reasns that culd cntribute t the discrdant test results frm this patient. Neisseria meningitidis There has been an increase in cases f N. meningitidis, a nrmal clnizer f the naspharynx identified in persns presenting with signs and symptms indicative f gnrrhea. 2 5 Due t the ptential bisafety cncerns, 6 labratries wrking up genital and extragenital cultures, including presumptive islates f N. gnrrheae, shuld be aware f the ptential presence f N. meningitidis in these cultures. Labratries shuld cnsult with a bisafety expert and/ r perfrm a risk assessment f their testing prcedures t ensure safe testing practices. The site f cllectin may impact the results. The ptimal specimen type fr detectin f N. gnrrheae infectin by NAAT are vaginal swabs frm wmen and first catch urine frm men. The use f ther specimen types can result in decreased sensitivity in the NAAT resulting in a negative NAAT result frm a patient that has a psitive Als, there are sme substances present in patient specimens that may be inhibitry t NAAT 8,10 but nt t grwth in The specimen cllectin swab itself might nt have been apprpriate fr the methd (either it was inhibitry t the PCR 8 and/r there was nt enugh f the rganism n the cllectin swab) resulting in a negative NAAT despite grwth in NAAT assays are designed t be very specific t the rganism being tested, therefre any mutatins in the pathgen that ccur within the target area f the assay culd result in the NAAT being negative despite the pathgen being present and being able t be cultured. While this is likely t be rare, it is pssible. Technical errr, specimen mix-up, instrumentatin errr, r target levels belw the assay s limit f detectin may result in a negative NAAT despite grwth in APHL N. gnrrheae Testing Methdlgies Fact Sheet 3

4 The specimen culd be negative fr N. gnrrheae, as the NAAT result indicated but a nngncccal Neisseria species was cultured that based n initial identificatin methds appeared t be N. gnrrheae. As discussed in the first example, in rder t perfrm bth culture and NAAT, tw different swabs are cllected. Discrepant results culd be the result f inadequate sampling with ne f the swabs. Being aware f the ptential reasns fr the discrdant results and explring whether any f these scenaris culd have cntributed t the result is an imprtant next step in determining whether the patient may have gnrrhea. The determinatin f the diagnsis and whether t initiate treatment will be dependent n many factrs including labratry findings, any ptential fllw-up testing, the clinical examinatin and risk factrs, public health department case definitins and the CDC STD Treatment Guidelines I received a preliminary reprt frm the labratry that indicated that the patient sample was presumptive negative fr gnrrhea but n the final reprt the NAAT was psitive fr N. gnrrheae and the culture was cnfirmed N. gnrrheae psitive. Hw culd this happen? Answer: A presumptive result based n clny mrphlgy grwth n selective media, Gram stain and xidase alne may be evaluated at 24 hurs and cnsidered presumptive negative if the clnies examined d nt fit the bichemical prfile. Hwever, grwth f Neisseria species may be delayed. In the labratry, the plates are incubated fr 48 hurs t check fr final grwth. It is pssible and nt uncmmn fr a culture t be negative at 24 hurs but psitive at 48 hurs. In additin, N. meningitidis resembles N. gnrrheae using initial screening tests and can nly be differentiated using further cnfirmatry testing. Sme rganisms may take lnger t grw and require additinal measures, such as enhanced culturing techniques, t prmte grwth. Testing methds require pure grwth f an rganism and this can be time-cnsuming. REFERENCES 1. Centers fr Disease Cntrl and Preventin. Recmmendatins fr the labratry-based detectin f Chlamydia trachmatis and Neisseria gnrrheae MMWR Recmm Rep Mrb Mrtal Wkly Rep Recmm Rep Mar 14;63(RR-02): Retchless AC, Kretz CB, Chang H-Y, Bazan JA, Abrams AJ, Nrris Turner A, et al. Expansin f a urethritis-assciated Neisseria meningitidis clade in the United States with cncurrent acquisitin f N. gnrrheae alleles. BMC Genmics ;19(1): Ma KC, Unem M, Jeverica S, Kirkcaldy RD, Takahashi H, Ohnishi M, et al. Genmic Characterizatin f Urethritis-Assciated Neisseria meningitidis Shws that a Wide Range f N. meningitidis Strains Can Cause Urethritis. J Clin Micrbil. 2017;55(12): Bazan JA, Petersn AS, Kirkcaldy RD, Briere EC, Maierhfer C, Turner AN, et al. Ntes frm the Field: Increase in Neisseria meningitidis-assciated Urethritis Amng Men at Tw Sentinel Clinics - Clumbus, Ohi, and Oakland Cunty, Michigan, MMWR Mrb Mrtal Wkly Rep Jun 3;65(21): Bazan JA, Turner AN, Kirkcaldy RD, Retchless AC, Kretz CB, Briere E, et al. Large Cluster f Neisseria meningitidis Urethritis in Clumbus, Ohi, Clin Infect Dis Off Publ Infect Dis Sc Am ;65(1): Centers fr Disease Cntrl and Preventin (CDC). Labratry-acquired meningcccal disease--united States, MMWR Mrb Mrtal Wkly Rep Feb 22;51(7): Papp JR, Henning T, Khubbar M, Kalve V, Bhattacharyya S, Travanty E, et al. Recvery f Neisseria gnrrheae frm 4 cmmercially available transprt systems. Diagn Micrbil Infect Dis Oct;86(2): Schrader C, Schielke A, Ellerbrek L, Jhne R. PCR inhibitrs - ccurrence, prperties and remval. J Appl Micrbil Nv;113(5): Centers fr Disease Cntrl and Preventin. Sexually Transmitted Diseases Treatment Guidelines, MMWR Recmm Rep Mrb Mrtal Wkly Rep Recmm Rep Jun 5;64(RR-3): APHL N. gnrrheae Testing Methdlgies Fact Sheet 4

5 10. Ng L-K, Martin IE. The labratry diagnsis f Neisseria gnrrheae. Can J Infect Dis Med Micrbil. 2005;16(1): Acknwledgments This dcument was develped by APHL s Sexually Transmitted Diseases Subcmmittee. Assciatin f Public Health Labratries The Assciatin f Public Health Labratries (APHL) wrks t strengthen labratry systems serving the public s health in the US and glbally. APHL s member labratries prtect the public s health by mnitring and detecting infectius and fdbrne diseases, envirnmental cntaminants, terrrist agents, genetic disrders in newbrns and ther diverse health threats. This prject was 100% funded with federal funds frm a federal prgram f $485,382. This publicatin was supprted by Cperative Agreement #5NU60OE frm the US Centers fr Disease Cntrl and Preventin (CDC). Its cntents are slely the respnsibility f the authrs and d nt necessarily represent the fficial views f CDC r the Department f Health and Human Services Gergia Avenue, Suite 700 Silver Spring, MD Phne: Fax: , Assciatin f Public Health Labratries. All Rights Reserved.