1921
Volume 94, Issue 4
  • ISSN: 0002-9637
  • E-ISSN: 1476-1645

Abstract

Abstract

Leptospirosis is an important zoonosis worldwide, with infections occurring after exposure to contaminated water. Despite being a global problem, laboratory diagnosis remains difficult with culture results taking up to 3 months, serology being retrospective by nature, and polymerase chain reaction showing limited sensitivity. Leptospira have been shown to survive and multiply in blood culture media, and we hypothesized that extracting DNA from incubated blood culture fluid (BCF), followed by quantitative real-time polymerase chain reaction (qPCR) could improve the accuracy and speed of leptospira diagnosis. We assessed this retrospectively, using preincubated BCF of spp. positive ( = 109) and negative ( = 63) febrile patients in Vientiane, Lao PDR. The final method showed promising sensitivities of 66% (95% confidence interval [CI]: 55–76) and 59% (95% CI: 49–68) compared with direct or direct and indirect testing combined, as the respective reference standards (specificities > 95%). Despite these promising diagnostic parameters, a subsequent prospective evaluation in a Lao hospital population ( = 352) showed that the sensitivity was very low (∼30%) compared with qPCR on venous blood samples. The disappointingly low sensitivity does suggest that venous blood samples are preferable for the clinical microbiology laboratory, although BCF might be an alternative if leptospirosis is only suspected postadmission after antibiotics have been used.

[open-access] This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

Loading

Article metrics loading...

/content/journals/10.4269/ajtmh.15-0674
2016-04-06
2017-11-22
Loading full text...

Full text loading...

/deliver/fulltext/14761645/94/4/736.html?itemId=/content/journals/10.4269/ajtmh.15-0674&mimeType=html&fmt=ahah

References

  1. Pappas G, Papadimitriou P, Siozopoulou V, Christou L, Akritidis N, , 2008. The globalization of leptospirosis: worldwide incidence trends. Int J Infect Dis 12: 351357.[Crossref]
  2. Lau CL, Smythe LD, Craig SB, Weinstein P, , 2010. Climate change, flooding, urbanisation and leptospirosis: fuelling the fire? Trans R Soc Trop Med Hyg 104: 631638.[Crossref]
  3. Forbes AE, Zochowski WJ, Dubrey SW, Sivaprakasam V, , 2012. Leptospirosis and Weil's disease in the UK. QJM 105: 11511162.[Crossref]
  4. Taylor AJ, Paris DH, Newton PN, , 2015. A systematic review of the mortality from untreated leptospirosis. PLoS Negl Trop Dis 9: e0003866.[Crossref]
  5. Dittrich S, Rattanavong S, Lee SJ, Panyanivong P, Craig SB, Tulsiani SM, Blacksell SD, Dance DA, Dubot-Peres A, Sengduangphachanh A, Phoumin P, Paris DH, Newton PN, , 2015. Orientia, Rickettsia, and Leptospira pathogens as causes of CNS infections in Laos: a prospective study. Lancet Glob Health 3: e104e112.[Crossref]
  6. Goris MG, Leeflang MM, Loden M, Wagenaar JF, Klatser PR, Hartskeerl RA, Boer KR, , 2013. Prospective evaluation of three rapid diagnostic tests for diagnosis of human leptospirosis. PLoS Negl Trop Dis 7: e2290.[Crossref]
  7. Levett PN, , 2001. Leptospirosis. Clin Microbiol Rev 14: 296326.[Crossref]
  8. Picardeau M, Bertherat E, Jancloes M, Skouloudis AN, Durski K, Hartskeerl RA, , 2014. Rapid tests for diagnosis of leptospirosis: current tools and emerging technologies. Diagn Microbiol Infect Dis 78: 18.[Crossref]
  9. Thaipadunpanit J, Chierakul W, Wuthiekanun V, Limmathurotsakul D, Amornchai P, Boonslip S, Smythe LD, Limpaiboon R, Hoffmaster AR, Day NP, Peacock SJ, , 2011. Diagnostic accuracy of real-time PCR assays targeting 16S rRNA and lipL32 genes for human leptospirosis in Thailand: a case-control study. PLoS One 6: e16236.[Crossref]
  10. Wuthiekanun V, Chierakul W, Limmathurotsakul D, Smythe LD, Symonds ML, Dohnt MF, Slack AT, Limpaiboon R, Suputtamongkol Y, White NJ, Day NP, Peacock SJ, , 2007. Optimization of culture of Leptospira from humans with leptospirosis. J Clin Microbiol 45: 13631365.[Crossref]
  11. Agampodi SB, Matthias MA, Moreno AC, Vinetz JM, , 2012. Utility of quantitative polymerase chain reaction in leptospirosis diagnosis: association of level of leptospiremia and clinical manifestations in Sri Lanka. Clin Infect Dis 54: 12491255.[Crossref]
  12. Boonsilp S, Thaipadungpanit J, Amornchai P, Wuthiekanun V, Chierakul W, Limmathurotsakul D, Day NP, Peacock SJ, , 2011. Molecular detection and speciation of pathogenic Leptospira spp. in blood from patients with culture-negative leptospirosis. BMC Infect Dis 11: 338.[Crossref]
  13. Griffith ME, Horvath LL, Mika WV, Hawley JS, Moon JE, Hospenthal DR, Murray CK, , 2006. Viability of Leptospira in BacT/ALERT MB media. Diagn Microbiol Infect Dis 54: 263266.[Crossref]
  14. Villumsen S, Pedersen R, Krogfelt KA, Jensen JS, , 2010. Expanding the diagnostic use of PCR in leptospirosis: improved method for DNA extraction from blood cultures. PLoS One 5: e12095.[Crossref]
  15. Mayxay M, Castonguay-Vanier J, Chansamouth V, Dubot-Pérès A, Paris DH, Phetsouvanh R, Tangkhabuanbutra J, Douangdala P, Inthalath S, Souvannasing P, Slesak G, Tongyoo N, Chanthongthip A, Panyanouvong P, Sibounheuang B, Phommasone K, Dohnt M, Phonekeo D, Hongvanthong B, Xayadeth S, Ketmayoon P, Blacksell SD, Moore CE, Craig SB, Burns M-A, von Sonnenburg F, Corwin A, de Lamballerie X, González IJ, Christophel EM, Cawthorne A, Bell D, Newton PN, , 2013. Causes of non-malarial fever in Laos: a prospective study. Lancet Glob Health 1: e46e54.[Crossref]
  16. Phetsouvanh R, Phongmany S, Soukaloun D, Rasachak B, Soukhaseum V, Soukhaseum S, Frichithavong K, Khounnorath S, Pengdee B, Phiasakha K, Chu V, Luangxay K, Rattanavong S, Sisouk K, Keolouangkot V, Mayxay M, Ramsay A, Blacksell SD, Campbell J, Martinez-Aussel B, Heuanvongsy M, Bounxouei B, Thammavong C, Syhavong B, Strobel M, Peacock SJ, White NJ, Newton PN, , 2006. Causes of community-acquired bacteremia and patterns of antimicrobial resistance in Vientiane, Laos. Am J Trop Med Hyg 75: 978985.
  17. Nic Fhogartaigh C, Dance DA, Davong V, Tann P, Phetsouvanh R, Turner P, Dittrich S, Newton PN, , 2015. A novel technique for detecting antibiotic-resistant typhoid from rapid diagnostic tests. J Clin Microbiol 53: 17581760.[Crossref]
  18. Bustin SA, Benes V, Garson JA, Hellemans J, Huggett J, Kubista M, Mueller R, Nolan T, Pfaffl MW, Shipley GL, Vandesompele J, Wittwer CT, , 2009. The MIQE guidelines: minimum information for publication of quantitative real-time PCR experiments. Clin Chem 55: 611622.[Crossref]
  19. Greiner M, Pfeiffer D, Smith RD, , 2000. Principles and practical application of the receiver-operating characteristic analysis for diagnostic tests. Prev Vet Med 45: 2341.[Crossref]
  20. Jiang J, Chan TC, Temenak JJ, Dasch GA, Ching WM, Richards AL, , 2004. Development of a quantitative real-time polymerase chain reaction assay specific for Orientia tsutsugamushi . Am J Trop Med Hyg 70: 351356.
  21. Waggoner JJ, Balassiano I, Abeynayake J, Sahoo MK, Mohamed-Hadley A, Liu Y, Vital-Brazil JM, Pinsky BA, , 2014. Sensitive real-time PCR detection of pathogenic Leptospira spp. and a comparison of nucleic acid amplification methods for the diagnosis of leptospirosis. PLoS One 9: e112356.[Crossref]
  22. Noda AA, Rodriguez I, Rodriguez Y, Govin A, Fernandez C, Obregon AM, , 2014. High sensitive PCR method for detection of pathogenic Leptospira spp. in paraffin-embedded tissues. Rev Inst Med Trop Sao Paulo 56: 411415.[Crossref]
  23. Villumsen S, Pedersen R, Borre MB, Ahrens P, Jensen JS, Krogfelt KA, , 2012. Novel TaqMan(R) PCR for detection of Leptospira species in urine and blood: pit-falls of in silico validation. J Microbiol Methods 91: 184190.[Crossref]
  24. Waggoner JJ, Balassiano I, Mohamed-Hadley A, Vital-Brazil JM, Sahoo MK, Pinsky BA, , 2015. Reverse-transcriptase PCR detection of leptospira: absence of agreement with single-specimen microscopic agglutination testing. PLoS One 10: e0132988.[Crossref]
  25. Limmathurotsakul D, Turner EL, Wuthiekanun V, Thaipadungpanit J, Suputtamongkol Y, Chierakul W, Smythe LD, Day NP, Cooper B, Peacock SJ, , 2012. Fool's gold: why imperfect reference tests are undermining the evaluation of novel diagnostics: a reevaluation of 5 diagnostic tests for leptospirosis. Clin Infect Dis 55: 322331.[Crossref]
  26. Ganoza CA, Matthias MA, Saito M, Cespedes M, Gotuzzo E, Vinetz JM, , 2010. Asymptomatic renal colonization of humans in the Peruvian Amazon by Leptospira . PLoS Negl Trop Dis 4: e612.[Crossref]
http://instance.metastore.ingenta.com/content/journals/10.4269/ajtmh.15-0674
Loading
/content/journals/10.4269/ajtmh.15-0674
Loading

Data & Media loading...

  • Received : 14 Sep 2015
  • Accepted : 28 Dec 2015

Most Cited This Month

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error