Burkholderia pseudomallei
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Burkholderia pseudomallei levels in specimens and use of selective media. Burkholderia pseudomallei. Vanaporn Wuthiekanun Senior Microbiologist. Culture remains the diagnostic gold standard for meiloidosis although it has low sensitivity

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Vanaporn Wuthiekanun Senior Microbiologist

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Vanaporn wuthiekanun senior microbiologist

Burkholderia pseudomallei levels in specimens and use of selective media

Burkholderia pseudomallei

Vanaporn Wuthiekanun

Senior Microbiologist


Vanaporn wuthiekanun senior microbiologist

Culture remains the diagnostic gold standard for meiloidosis although it has low sensitivity

B. pseudomallei grows well on routine laboratory media, while selective media help with non-sterile specimens

Higher B. pseudomallei counts in clinical samples correlate with higher mortality rates.

Introduction

Limmathurotsakul et.al PLOS One 2010

Wuthiekanun et al.Am J. Trop. Med. Hyg 2007


Vanaporn wuthiekanun senior microbiologist

Melioidosis Research Programme

Clinical samples

  • Blood

  • Respiratory secretion

  • Urine

  • Pus and fluid

Spleenic pus from

Melioidosis patient


Vanaporn wuthiekanun senior microbiologist

Specimens from non-sterile sites can benefit from the use of selective media

A number of selective media have been developed to facilitate growth of B. pseudomallei from sites with mixed flora.

Ashdown agar (ASH)

B. pseudomallei selective agar (BPSA)

B. cepacia selective media (BCSA)

Pseudomonas cepacia agar (PCA)

Selective media

Ashdown et al.Pathology1979a, Howard et al. Clinical Micro 2003,

Peacock et al. Clinical Micro 2005, Glass et al.Am J. Trop. Med. Hyg 2009


Vanaporn wuthiekanun senior microbiologist

BCSA is an equivalent selective agar to ASH

The most sensitive medium for the growth of B. pseudomallei was ASH

PCA is a commercial selective agar that may be suitable for B. pseudomallei and B. mallei

Summary of selective media

Peacock et al. Clinical Micro 2005, Glass et al.Am J. Trop. Med. Hyg 2009


Ceftazidime resistant strains due to deletion of pbp3

Ceftazidime resistant strains due to deletion of PBP3

Etests Gram stain RTM3 Microscopy

Ceftazidime-sensitive initial isolate

Ceftazidime-resistant secondary isolate

Chantratita N et al. PNAS 2012


Vanaporn wuthiekanun senior microbiologist

Bacteraemia and outcome

100

1000

  • If a blood culture becomes positive within 24 h, the mortality rate has been found to be at 73.7% compared with a 40.9% mortality rate after 24 hours.

  • Quantitatively, if there are more than 100 colony forming units (CFU)/ml of B. pseudomalleiin blood, mortality rates reach 96% where <1 CFU/ml of B. pseudomallei has a relative mortality rate of 42%

MORTALITY

50

100

10

0

<1 1-100 >100

cfu/mL

1

B.pseudomallei cfu/mL

Walsh et.al Clin Infect Dis 1995

Wuthiekanun et.al Clin Infect Dis 2006


Vanaporn wuthiekanun senior microbiologist

  • Throat swab:1011 melioidosis, 3524 healthy (4,625)

  • Specificity 100% (no carrier)

  • Sensitivity 79% (sputum)

Value of throat swab in diagnosis of melioidosis

Wuthiekanun et al. J Clin Microbiol 2001


The role and significance of quantitative urine cultures in the diagnosis of melioidosis

The role and significance of quantitative urine cultures in the diagnosis of melioidosis

  • Mortality rose with the increasing urine bacterial quantity

  • Only 24% of patients with positive urine cultures had urinary symptoms

  • The presence of B. pseudomallei in urine is associated with a poorer prognosis

0

<103

103 -104

>105

Limmathurotsakul et al. J Clin Micro., 2005


Quantitative number of b pseudomallei in clinical specimens

Quantitative number of B. pseudomallei in clinical specimens

  • Data of positive samples:

    • total 376/730

    • blood 203/414 (49%)

    • urine 56/268 (21%)

    • respiratory secretions 94/120 (78%)

    • pus 23/28 (82%)

  • Blood samples had the lowest count from <0.1 to >100 CFU/ml

  • Pus and respiratory secretions had the highest median count

Wuthiekanun et al.Am J. Trop. Med. Hyg 2007


Summary

Summary

  • Pitfalls in identification can include technical unfamiliarity with the disease and organism

  • Specimens from non-sterile sites can benefit from the use of selective media

  • Routine use of culture, in addition with PCR and serological test, may lead to earlier diagnosis of melioidosis


Vanaporn wuthiekanun senior microbiologist

Collaborators and Contributors

MORU,

Faculty of Tropical Medicine,

Mahidol University

University of Cambridge, UK

Prof Sharon Peacock

Dr Narisara Chantratita

Dr Wirongrong Chierakul

Dr Direk Limmathurotsakul

Dr Rapeephan Rattanawongnara

Prof Nicholas White

Prof Nicholas Day

Sappasithiprasong Hospital

Prof Wipada Chaowagul

Dr Prapit Teparrukkul

Nitaya Teerawattanasuk

Medical staff and nurses

laboratory staff

Khon Kaen University

Dr Surasukdi Wongratanacheewin

Dr Rasana Wongratanacheewin

Menzies School of Health Reserach

Dr Bart J Currie

Dr. Allen Cheng


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