[논문]Affinity Apheresis for Treatment of Bacteremia Caused by Staphylococcus aureus and/or Methicillin-Resistant S. aureus (MRSA)

Mattsby-Baltzer, Inger; Bergstrom, Tomas; Mccrea, Keith; Ward, Robert; Msc, Lars Adolfsson; Larm, Olle

doi:10.4014/jmb.1102.02016

Affinity Apheresis for Treatment of Bacteremia Caused by Staphylococcus aureus and/or Methicillin-Resistant S. aureus (MRSA) 원문보기

Journal of microbiology and biotechnology, v.21 no.6, 2011년, pp.659 - 664

Mattsby-Baltzer, Inger (Department of Infectious Diseases) , Bergstrom, Tomas (Department of Infectious Diseases) , Mccrea, Keith (Emergence LLC) , Ward, Robert (Emergence LLC) , Msc, Lars Adolfsson (ExThera AB, Karolinska Science Park) , Larm, Olle (ExThera AB, Karolinska Science Park)

Abstract ▼ AI-Helper

Staphylococcus aureus (SA) bacteremia is associated with high mortality, and often results in metastatic infections. The methicillin-resistant SA (MRSA) is an urgent health care issue, as nosocomial infections with these bacteria represent limited treatment alternatives. Samples of whole blood containing challenge inoculums of SA and MRSA strains were passed through columns packed with surfaceheparinized polyethylene beads. The bound bacteria were eluted and quantitatively determined by culturing and by real-time PCR. Significant amounts of both SA and MRSA adhered to the heparinized beads (more than 65% of inoculated bacteria). After rinsing with buffer at high ionic strength, viable bacteria or bacterial DNA were eluted from the columns, indicating that the binding was specific. The conclusions that can be made from these experiments are that, as earlier reported in the literature, the high affinity of SA to heparin is retained in whole blood, and MRSA in whole blood binds to heparin with similar or higher affinity than SA. It should be possible to lower the amount of SA and/or MRSA from the blood of infected patients to levels that could be taken care of by the immune system. In previous studies, we have shown that passing blood from septic patients over beads coated with end-point-attached, biologically active heparin is a useful technique for regulating the levels of heparinbinding cytokine. These findings in combination with the present findings indicate the possibility of creating an apheresis technology for treatment of sepsis caused by SA and/or MRSA.

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가설 설정

We hypothesize that when infected blood is passed through a device, consisting of a column packed with beads coated with biologically active heparin (i.e., end-point attached), many bacteria including SA would be removed. Our aim was to investigate to what extent SA and MRSA are removed from whole blood after passage over surface-heparinized polyethylene beads.

제안 방법

Comparing the recoveries for each staphylococcal strain in void, wash, and eluate samples, based on percentages of the total numbers of CFU or bDNA in corresponding nonpassed blood controls, the bacteria bound more strongly to the Hep columns compared with nonHep columns regarding the content in the void samples (Fig. 2). However, owing to the variability, no statistically significant difference was achieved.
, end-point attached), many bacteria including SA would be removed. Our aim was to investigate to what extent SA and MRSA are removed from whole blood after passage over surface-heparinized polyethylene beads.
The authors thank Dr. George Pitarra at Emergence Inc., for financial support and valuable discussions, and Kerstin Florén, Sahlgren’s hospital, Gothenburg, for assistance with the real-time PCR analyses.

대상 데이터

aureus (CCUG1800T, not methcillin resistant) was used as a control. Five MRSA strains were obtained from the Culture Collection, University of Gothenburg (CCUG). Three of those strains were PVL-positive and typed as USA300 (CCUG: 51485, 53251, and 54860), whereas the other two strains were typed as USA100 (CCUG: 46314, 55517).
Polyethylene (PE) beads, with an average diameter of 0.3 mm (lot no. 180153), were supplied by the Polymer Technology Group (Berkeley, USA) and the columns (Polyprep, 0.8×4 cm) were obtained from BIORAD (USA).

데이터처리

Means are indicated. Paired t-test was used for the statistics.

이론/모형

Real-time PCR based on SyberGreen fluorescent dye, which selectively binds to double-stranded DNA templates, was performed with the Rotorgene method (Corbett Rotor-Gene 3000, QIAGEN AB, Sollentuna, Sweden). The analysis was performed according to the manufacturer´s manual.

성능/효과

It should be mentioned that the percentages of mean total recoveries of the Hep columns were 60% for CFU, and 82% for bDNA. The corresponding figures for nonHep columns were 50% and 85%, respectively.
The capacity of bacteria to bind to HS proteoglycans could be regarded as a virulence factor [9], and two heparin-binding proteins have been isolated from SA [15]. The two MRSA strains that showed the lowest eluate values (bDNA and CFU) of the Hep column (Fig. 2), and which also corresponded to the lowest void values, indicated that their binding to heparin was particularly strong, and therefore less easily reverted by the salt solution. Different isolates of MRSA may vary in their level of expression of HS-binding proteins.

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