Comparative diagnostic accuracy study of three multiplex molecular assays for bacterial gastroenteritis Simon Goldenberg Consultant Microbiologist and Infection Control Doctor Guy’s & St Thomas’ NHS Foundation Trust Honorary Senior Lecturer King’s College London 1000 Top 10 most frequent reasons for emergency 900 admission, Guy’s & St Thomas’ Hospital, 800 2014/15 700 600 500 emergency admissions emergency 400 300 200 Number Number of 100 0 Primary diagnosis code Clinical features are not helpful for predicting the etiology of diarrhoea • Duration of diarrhoea • Abdominal pain • Vomiting • Blood stools • Number of bowel movements • Fever • Requirement for IV hydration This emphasizes the need for more rapid, sensitive, and simple methods to improve the diagnostic yield Chan et al. Hong Kong J Emerg Med. 2003;10:162-168 Svenungusson et al. Clin Infect Dis. 2000 30 (5): 770-778 Munk Petersen et al. Scand J Gastroenterol 1996;31:906-11 Svanteson et al. Scand J Infect Dis 1988;20:303-14 aripiprazole, acarbose, acamprosate, acebutolol, aceclofenac, acemetacin, acetazolamide, acyclovir, acitretin, activated charcoal, agalsidase beta, alendronate, alfuzosin, allopurinol, almotriptan, amphotericin, amiloride, aminophylline, amitriptyline, amlodipine, amoxicillin, anastrozole, anidulafungin, allantoin, aspirin, atenolol, atorvastatin, aztreonam, azathioprine, azithromycin, barium sulphate, brinzolamide, baclofen, bendroflumethiazide, belimumab, betaxolol, bezafibrate, bisoprolol, bleomycin, bisacodyl, bosutinib, bromocriptine, buprenorphine, bumetanide, busulfan, buspirone, calcium carbonate, calcium folinate, calcium glubionate, calcium lactobionate, caffeine, colestilan, caspofungin, capecitabine, captopril, carglumic acid, carbamazepine, carboplatin, cetirizine, carvedilol, cefadroxil, cephalexin, cefotaxime, cefradine, ceftazidime, ceftriaxone, cefuroxime, celecoxib, celiprolol, chlorambucil, chlorphenamine, colesevelam, ciprofloxacin, cimetidine, citalopram, cisplatin, clarithromycin, clindamycin, clofarabine, clopidogrel, clozapine, codeine phosphate, colchicine, colestipol, colistimethate, corifollitropin alfa, crisantaspase, cytarabine, dacarbazine, daptomycin, darifenacin, decitabine, dantrolene, daunorubicin, deferasirox, deferiprone, desferrioxamine, desloratadine, dihydrocodeine, diazepam, diclofenac, digoxin, diltiazem, docusate sodium, dipyridamole, disodium pamidronate, docetaxel, domperidone, donepezil, dorzolamide hydrochloride, doxazosin, doxorubicin, doxycycline, ethosuximide, esomeprazole, emtricitabine, enalapril, etanercept, entacapone, epirubicin, epoprostenol, etoposide, epoetin alfa, erdosteine, erythromycin, estradiol, ethambutol, ethosuximide, etodolac, ethinylestradiol, exemestane, fentanyl, fenofibrate, ferrous sulphate, ferrous fumarate, flecainide Electronic Medicines Compendium www.medicines.org.uk aripiprazole, acarbose, acamprosate, acebutolol, aceclofenac, acemetacin, acetazolamide, acyclovir, acitretin, activated charcoal, agalsidase beta, alendronate, alfuzosin, allopurinol, almotriptan, amphotericin, amiloride, aminophylline, amitriptyline, amlodipine, amoxicillin, anastrozole, anidulafungin, allantoin, aspirin, atenolol, atorvastatin, aztreonam, azathioprine, azithromycin, barium sulphate, brinzolamide, baclofen, bendroflumethiazide, belimumab, betaxolol, bezafibrate, bisoprolol, bleomycin, bisacodyl, bosutinib, bromocriptine, buprenorphine, bumetanide, busulfan, buspirone, calcium carbonate, calcium folinate, calcium glubionate, calcium lactobionate, caffeine, colestilan, caspofungin3141, capecitabine drug, captopril, preparationscarglumic acid, carbamazepine, carboplatin, cetirizine, carvedilol, cefadroxil, cephalexin, cefotaxime, cefradine, ceftazidime, ceftriaxone, cefuroxime, celecoxib, celiprolol, chlorambucil, chlorphenamine, colesevelam, ciprofloxacin, cimetidine, citalopram, cisplatin, clarithromycin, clindamycin,with clofarabine, clopidogreldiarrhoea, clozapine, codeine asphosphate, a colchicine, colestipol, colistimethate, corifollitropin alfa, crisantaspase, cytarabine, dacarbazine, daptomycin, darifenacin, decitabine, dantrolene, daunorubicin, deferasirox, deferiprone, desferrioxamine, desloratadine, dihydrocodeine, diazepam,known diclofenac, digoxin, unwanted diltiazem, docusate sodium, effect dipyridamole , disodium pamidronate, docetaxel, domperidone, donepezil, dorzolamide hydrochloride, doxazosin, doxorubicin, doxycycline, ethosuximide, esomeprazole, emtricitabine, enalapril, etanercept, entacapone, epirubicin, epoprostenol, etoposide, epoetin alfa, erdosteine, erythromycin, estradiol, ethambutol, ethosuximide, etodolac, ethinylestradiol, exemestane, fentanyl, fenofibrate, ferrous sulphate, ferrous fumarate, flecainide Electronic Medicines Compendium www.medicines.org.uk Managing diarrhoea in hospital is time consuming and inefficient • 20% of infection control team time is taken is spent on management of diarrhoea • Almost all patients (99%) with suspected infectious diarrhoea are isolated in a side room • Mean number of wards closed annually due to infectious diarrhoea = 12 per hospital • 25% of the time causative pathogens are identified after patient discharge Buchanan et al J Hosp Infect 2015 doi: 10.1016/j.hin.2014.12.021. ID 1 1 3 CCDA 18-24h CCDA 24h 2 Antimicrobial susceptibility testing 2-10h 6 4 Biochemical ID and agglutination tests Selenite-F 18- ABC5 18-24h ID 24h 2 Antimicrobial susceptibility testing 2-10h3 3.7h ID Biochemical ID and agglutination tests CT-SMAC7 18-24h 2 Antimicrobial susceptibility testing 2-10h ID Biochemical ID and agglutination tests XLD8 18-24h 2 Antimicrobial susceptibility testing 2-10h Gram stain, basic Selective culture Species level identification using Antimicrobial Sample biochemistry transportation +/- enrichment MALDI-TOF MS and biochemical susceptibility testing (Oxidase etc) identification (API 20E) Total time = 66.5 hours, range - 64 hours (negative) to 118 hours (positive) Conventional culture based detection of bacterial enteric pathogens Poor sensitivity of conventional and culture based tests compared with molecular assays Study Method n % positive % positive conventional molecular Amar. Eur J Clin Microbiol Infect Dis. In house 2205 19 42 2007 de Boer. J Clin Microbiol. 2010 In house 13,974 6.4 19.2 Wessels. Clin Microbiol Infect. 2014 Luminex xTAG GPP 393 11.2 22.1 Mengelle. Clin Microbiol Infect. 2013 Luminex xTAG GPP 440 32.7 40 McAuliffe. J Infect. 2013 Fast Track Diagnostics 530 18 30 Onori. Diagn Microbiol Infect Dis. SeeGene Seeplex ACE 245 66.5 76.3 2014 Cunningham. J Clin Microbiol. 2010 In house 392 26.5 96.7 Biswas. Eur J Clin Microbiol Infect BD Max Enteric Panel 434 86.1 94.4 Dis. 2014 Problems with current culture dependent diagnostic methods: • Inability to differentiate clinically between different cause of IG • Fragmented test request procedure • Inefficient laboratory workflow • Laborious workflow – slow, long TAT • Poor sensitivity – lack of clinical confidence Enterics panel evaluation: • RBioPharm RIDA Gene Bacterial Stool Panel and EHEC/EPEC Manual two-tube real time PCR on ABI7500 Separate automated extraction using Promega Maxwell instrument (magnetic beads) Salmonella, Camylobacter, Y. enterocolitica*, Shigella, E. coli • Fast Track Diagnostics Bacterial Gastroenteritis panel Manual two-tube real time PCR on ABI7500 Separate automated extraction using same extract as above Salmonella, Campylobacter, Y. enterocolitica*, Shigella, E. coli, C. difficile* • BD Max Gastroenteritis panel Fully automated reat time PCR with built in extraction Salmonella, Campylobacter, Shigella, E. coli • Conventional selective culture methods: CCDA – Campylobacter, XLD – Salmonella/Shigella CT-SMAC EC0157 Selenite F broth & ABC Harlequin – Salmonella *C. difficile and Y. enterocolitia not assessed in this evaluation Enterics panel evaluation: • 434 non-duplicate samples tested by all 4 methods: • 116 retrospective samples selected on the basis of culture positivity • Remaining 318 consecutive samples tested semi- prospectively Gold standard / True positive: • Either culture positive OR • 2/3 positive molecular tests Results 18 16 14 12 10 8 6 4 2 Campylobacter 0 Shigella Culture Salmonella Fast Track Diagnostics RIDA-Gene BD Max Relative performance characteristics of molecular panels and conventional culture combined samples (n=434) RIDA®GENE Bacterial Stool FTD® Bacterial BD MAX™ Enteric Bacterial Conventional culture Panel and EHEC/EPEC Gastroenteritis Panel Panel Sensitivity Specificity Sensitivity Specificity Sensitivity Specificity Sensitivity Specificity (95% CI) (95% CI) (95% CI) (95% CI) (95% CI) (95% CI) (95% CI) (95% CI) Campylobacter 91.1 98.8 80.2 99.4 92.1 100 91.1 100 (83.8 – 95.8) (97 – 99.7) (71.1 – 87.5) (97.8 – 99.9) (85 – 96.5) (98.9 – 100) (83.8 – 95.8) (98.9 – 100) Shigella 94.4 99.7 88.9 100 94.4 100 86.1 100 (81.3 – 100) (98.6 – 100) (73.9 – 96.9) (99.1 – 100) (81.3 – 99.3) (99.1 – 100) (70.5 – 95.3) (99.1 – 100) Salmonella 55 99 45 99.8 75 100 85 100 (31.5 – 76.9) (97.5 – 99.7) (23.1 – 68.5) (98.7 – 100) (50.9 – 91.3) (99.1 – 100) (62.1 – 96.8) (98.9 – 100) Shiga-toxin 0 99.5 100 99.8 100 99.5 100 100 producing E. (0 – 97.5) (98.3 – 99.9) (25 – 100) (98.7 – 100) 25 – 100) (98.3 – 99.9) (25 – 100) (99.2 – 100) coli RIDA®GENE bacterial Fast Track Diagnostics BD Max™ Enteric Conventional culture stool panel and bacterial bacterial panel Relative performance EHEC/EPEC gastroenteritis panel characteristics of Campylobacter, prevalence = 5.3% molecular panels and Sensitivity 100 88.2 100 52.9 conventional