Sepsis detectives

The start of a new year brought a flurry of papers on “laboratory diagnosis of sepsis”. Problems with the fundamentals of a laboratory (a geographic or abstract entity; not a person with diagnostic skills and insights) diagnosis aside, our colleagues have been busy examining the potential for molecular tests to guide early treatment of severe bacterial infection.

The core concept is to exploit the combination of speed, specificity and sensitivity of molecular biology to give a name to the biological causal agent much faster than the 12-24 hours it generally takes with culture-based methods that rely on the speed of bacterial replication. The idea is not new. It has been around pretty much as long as there have been molecular microbiologists standing by idle thermal cyclers. So what has stood in the way of progress for so long? The quick answer is the complexity of the task: septicaemia can be caused by a host of bacteria, and each requires their specific set of probes. True multiplex PCR-based methods are relatively new. The more targets, the more arduous the validation process. And the more costly the end product.

But there are other hurdles for the budding bacterial gene jockey. The first of these is that there is no true “gold standard” for septicaemia. Blood culture is prone to false positives when skin organisms contaminate the blood inoculated into culture vials, explaining why so many contain coagulase negative staphylococci, Corynebacterium and Propionibacterium species. There are also factors that reduce sensitivity, for example, variable but generally low quantities of blood use to inoculate blood culture bottles, variable showering of bacterial into the peripheral circulation, and prior use of antibiotics that may have already begun to inhibit or kill the very bacteria you’re trying to recover in the lab. Once we start using molecular methods, we stray into tiger country with potential inhibition by human DNA, or the anticoagulants used to stop the blood in the culture bottle from clotting. A tough job then.

So I applaud the huge effort the groups that have published have put in, and I welcome their optimism that we can do better. But as we read their reports, we need to ask the hard questions about how close their method is to offering a point-of-care support to the physicians who make those initial clinical decisions.

It strikes me that there are several key questions that any study of this kind needs to answer:

  1. Can the method offer a validated, definitive result less that four hours after the patients arrives at the hospital?
  2. Exactly how sensitive is it (i.e. how much blood do you need from a patient with septic shock to guarantee a positive result)?
  3. How wide a range of organism-specific targets are on offer and what percentage of septicæmias will this cover at the test site?
  4. Can the test procedure operate a round-the-clock service?
  5. How do the designers propose to integrate the method into both clinical laboratory practice and front line clinical medicine?

Two recent review articles provide a useful entry point to this complex area. They can be summarized by the current verdict:  “promising technology; needs to try harder”.


  1. Mancini N, Carletti S, Ghidoli N, Cichero P, Burioni R, Clementi M. The era of molecular and other non-culture-based methods in diagnosis of sepsis. Clin Microbiol Rev. 2010 Jan;23(1):235-51.
  2. Dark PM, Dean P, Warhurst G.. Bench -to-bedside review: the promise of rapid infection diagnosis during sepsis using polymerase chain reaction-based pathogen detection. Crit Care. 2009;13(4):217. Epub 2009 Jul 15.

Original papers

  1. Avolio M, Diamante P, Zamparo S, Modolo ML, Grosso S, Zigante P, Tosoni N, De Rosa R, Stano P, Camporese A. Molecular identification of bloodstream pathogens in patients presenting to the emergency room with suspected sepsis. Shock. 2010 Jan 19. [Epub ahead of print]
  2. Bloos F, Hinder F, Becker K, Sachse S, Dessap AM, Straube E, Cattoir V, Brun-Buisson C, Reinhart K, Peters G, Bauer M. A multicenter trial to compare blood culture with polymerase chain reaction in severe human sepsis. Intensive Care Med. 2010 Feb;36(2):241-7. Epub 2009 Nov 19.
  3. Lehmann LE, Hunfeld KP, Steinbrucker M, Brade V, Book M, Seifert H, Bingold T, Hoeft A, Wissing H, Stüber F. Improved detection of blood stream pathogens by real-time PCR in severe sepsis. Intensive Care Med. 2010 Jan;36(1):49-56. Epub 2009 Sep 15.
  4. Tissari P, Zumla A, Tarkka E, Mero S, Savolainen L, Vaara M, Aittakorpi A, Laakso S, Lindfors M, Piiparinen H, Mäki M, Carder C, Huggett J, Gant V.  Accurate and rapid identification of bacterial species from positive blood cultures with a DNA-based microarray platform: an observational study. Lancet. 2010 Jan 16;375(9710):224-230. Epub 2009 Dec 10.
  5. Tsalik EL, Jones D, Nicholson B, Waring L, Liesenfeld O, Park LP, Glickman SW, Caram LB, Langley RJ, van Velkinburgh JC, Cairns CB, Rivers EP, Otero RM, Kingsmore SF, Lalani T, Fowler VG, Woods CW. Multiplex PCR to diagnose bloodstream infections in patients admitted from the emergency department with sepsis. J Clin Microbiol. 2010 Jan;48(1):26-33. Epub 2009 Oct 21.

µGnome, 31st January, 2010

“swine flu” or pneumonia?

Ingram PR, Inglis T, Moxon D, Speers D. Procalcitonin and C-reactive protein in severe 2009 H1N1 influenza infection. Intensive Care Med. 2010 Jan 13. [Epub ahead of print]

When we were faced with an influx of ICU patients with H1N1/09 influenza, we wanted a more rapid method of distinguishing bacterial pneumonia from influenza to help with antibiotic decisions. We had expected the procalcitonin would be the most effective decision support but it turned out that C-reactive protein was a bit better as a single test, and the most effective was a combination of both when ruling out serious bacterial infection.

Pro-calcitonin has become a useful test in ICU, supporting decisions regarding the progression of bacterial infection and the need for antibiotic interventions (1,2). It will be interesting to see how the combination of CRP and PCT fares during the next influenza surge.

  1. Tsangaris I, Plachouras D, Kavatha D, Gourgoulis GM, Tsantes A, Kopterides P, Tsaknis G, Dimopoulou I, Orfanos S, Giamarellos-Bourboulis E, Giamarellou H, Armaganidis A. Diagnostic and prognostic value of procalcitonin among febrile critically ill patients with prolonged ICU stay. Crit Care. 2009;13(2):R38.
  2. Charles PE, Tinel C, Barbar S, Aho S, Prin S, Doise JM, Olsson NO, Blettery B, Quenot JP.Procalcitonin kinetics within the first days of sepsis: relationship with the appropriateness of antibiotic therapy and the outcome. BMC Infect Dis. 2009 Dec 22;9:213.


Malaria in gorillas?

You heard about the origins of malaria in chimps? Well know, there is evidence it’s in gorillas as well. Admittedly, this is just a whiff of a trace in gorilla droppings, but it does have the smell of Plasmodium falciparum.

It remains to be seen whether this is a one-off or part of a bigger picture. In future, scouts are going to need a thermal cycler.


Life in liquids

Our most recent scientific paper highlights the survival ability of the bacteria that cause melioidosis (1).

Medical microbiology places its emphasis on the behaviour of medically-important microorganisms in conditions in and around the human body. In the clinical lab we force bacteria to grow as rapidly as we can in nutrient-rich cultures at 37’C. We only rarely use a wider range of other physical or chemical conditions and place the emphasis on environmental survival. However some bacteria are natural survivors. Burkholderia pseudomallei has been known survive for years in sterile, distilled water (2).

In our recent paper we took a look at what happened to B. pseudomallei in different types of water: distilled, tropical rain water and sea water across. We also looked at temperature range and pH, exploring physical and chemical conditions relevant to the Western Australian melioidosis outbreak in late 1997 (3). These bacteria are remarkably tough, adapting to acid pH, high temperatures and the osmotic stress of distilled water. But a part of that adaptation appears to be a change in shape. We observed bizarre forms that don’t fit neatly with the conventional description of B. pseudomallei as a Gram negative bacillus.

We also saw a modest increase in numbers while surviving in tropical rain water. B. pseudomallei caught up in clouds and deposited at a distance; something we believe may have explain the apparent movement of the WA outbreak strain NCTC 13177 500km to the East over an eight year period(4).

  1. Robertson J, Levy A, Sagripanti JL, Inglis TJ.  The survival of Burkholderia pseudomallei in liquid media.  Am J Trop Med Hyg. 2010 Jan;82(1):88-94.
  2. Wuthiekanun V, Smith MD, White NJ. Survival of Burkholderia pseudomallei in the absence of nutrients. Trans R Soc Trop Med Hyg. 1995 Sep-Oct;89(5):491.
  3. Inglis TJ, Garrow SC, Adams C, Henderson M, Mayo M. Dry-season outbreak of melioidosis in Western Australia. Lancet. 1998 Nov 14;352(9140):1600.
  4. Inglis TJ, Levy A, Merritt AJ, Hodge M, McDonald R, Woods DE. Melioidosis risk in a tropical industrial environment.  Am J Trop Med Hyg. 2009 Jan;80(1):78-84.


μGnome, 18-JAN-10.

New home for the μGnome

In the interests of  a better future for the μGnation, we’ve moved home.

The removers came this weekend and did a beautiful job migrating the μhome to Priobe Net, where we will put down roots and establish a thriving colony close to our friends, the Priobiologists.

We expect this will result in a rapid proliferation of posts on μGnostics, μGnomics and other μGnecessaries. The μGnation is excited.

μGnome, 17-JAN-10