The peptidoglycan and biofilm matrix of Staphylococcus epidermidis undergo structural changes when exposed to human platelets.

Staphylococcus epidermidis is a bacterium frequently isolated from contaminated platelet concentrates (PCs), a blood product used to treat bleeding disorders in transfusion patients. PCs offer an accidental niche for colonization of S. epidermidis by forming biofilms and thus avoiding clearance by immune factors present in this milieu. Using biochemical and microscopy techniques, we investigated the structural changes of the peptidoglycan (PG) and the biofilm matrix of S. epidermidis biofilms formed in whole-blood derived PCs compared to biofilms grown in glucose-supplemented trypticase soy broth (TSBg). Both, the PG and the biofilm matrix are primary mechanisms of defense against environmental stress. Here we show that in PCs, the S. epidermidis biofilm matrix is mainly of a proteinaceous nature with extracellular DNA, in contrast to the predominant polysaccharide nature of the biofilm matrix formed in TSBg cultures. PG profile studies demonstrated that the PG of biofilm cells remodels during PC storage displaying fewer muropeptides variants than those observed in TSBg. The PG muropeptides contain two chemical modifications (amidation and O-acetylation) previously associated with resistance to antimicrobial agents by other staphylococci. Our study highlights two key structural features of S. epidermidis that are remodeled when exposed to human platelets and could be used as targets to reduce septic transfusions events.

Septic transfusion case caused by a platelet pool with visible clotting due to contamination with Staphylococcus aureus.

BACKGROUND: Contamination of platelet concentrates (PCs) with Staphylococcus aureus is one of the most significant ongoing transfusion safety risks in developed countries. CASE REPORT: This report describes a transfusion reaction in an elderly patient diagnosed with acute myeloid leukemia, transfused with a 4-day-old buffy coat PC through a central venous catheter. The transfusion was interrupted when a large fibrous clot in the PC obstructed infusion pump flow. Shortly afterward, a red blood cell (RBC) unit transfusion started. After septic symptoms were developed, the RBC transfusion was also interrupted. While the RBC unit tested negative for bacterial contamination, the PC and the patient samples were found to be contaminated with a S. aureus strain that exhibited the same phenotypic and genome sequencing profiles. The isolated S. aureus forms biofilms and produces the superantigen enterotoxin-like U, which was detected in a sample of the transfused PCs. The patient received posttransfusion antibiotic treatment and had her original central line removed and replaced. DISCUSSION: As the implicated PC had been tested for bacterial contamination during routine screening yielding negative results, this is a false-negative transfusion sepsis case. Using a point-of-care test could have prevented the transfusion reaction. This report highlights the increasing incidence of S. aureus as a major PC contaminant with grave clinical implications. Importantly, S. aureus is able to interact with platelet components resulting in visible changes in PCs. CONCLUSION: Visual inspection of blood components before transfusion is an essential safety practice to interdict the transfusion of bacterially contaminated units.

Biofilm-forming skin microflora bacteria are resistant to the bactericidal action of disinfectants used during blood donation.

BACKGROUND: A one-step skin disinfection method containing 2% chlorhexidine-gluconate (CHG) and 70% isopropyl alcohol (IPA) is currently used by blood suppliers worldwide. Reports of bacterially contaminated platelet concentrates (PCs) indicate that skin disinfection is not fully effective. Approximately 20% of skin microflora exist as surface-attached aggregates (biofilms), known for displaying increased resistance to disinfectants. This study was aimed at determining whether skin microflora biofilm-positive Staphylococcus epidermidis and Staphylococcus capitis are resistant to CHG and/or IPA. STUDY DESIGN AND METHODS: Free-floating cells and mono or dual (1 : 1 ratio) biofilms of S. epidermidis and S. capitis were exposed to CHG, IPA, or CHG/IPA for 30 seconds, simulating skin disinfection practices. Residual viable cells were quantified by colony counting. Morphology of disinfectant-treated S. epidermidis biofilms was examined by scanning electron microscopy. Treated S. epidermidis and S. capitis biofilms were inoculated into PCs and bacterial concentrations were determined on Days 0 and 5 of storage. RESULTS: Treatment of staphylococcal biofilm cells with all disinfectants caused cell damage and significant reduction in viability, with CHG/IPA being the most effective. However, biofilms were significantly more resistant to treatment than free-floating cells. Disinfectant-treated S. epidermidis proliferated better in PCs than S. capitis, especially when grown as monospecies biofilms. CONCLUSION: Although CHG/IPA is effective in reducing the viability of S. epidermidis and S. capitis biofilms, these organisms are not completely eliminated. Furthermore, disinfectant-treated staphylococcal biofilms multiply well in PCs. These results demonstrate that the biofilm-forming capability of the skin microflora reduces the bactericidal efficiency of blood donor skin disinfectants.

Effect of platelet additive solution on bacterial dynamics and their influence on platelet quality in stored platelet concentrates.

BACKGROUND: Platelet additive solutions (PASs) are an alternative to plasma for the storage of platelet concentrates (PCs). However, little is known about the effect of PAS on the growth dynamics of contaminant bacteria. Conversely, there have been no studies on the influence of bacteria on platelet (PLT) quality indicators when suspended in PAS. STUDY DESIGN AND METHODS: Eight buffy coats were pooled, split, and processed into PCs suspended in either plasma or PAS (SSP+, MacoPharma). PCs were inoculated with 10 and 100 colony-forming units (CFUs)/bag of either Serratia liquefaciens or Staphylococcus epidermidis. Bacterial growth was measured over 5 days by colony counts and bacterial biofilm formation was assayed by scanning electron microscopy and crystal violet staining. Concurrently, PLT markers were measured by an assay panel and flow cytometry. RESULTS: S. liquefaciens exhibited an apparent slower doubling time in plasma-suspended PCs (plasma-PCs). Biofilm formation by S. liquefaciens and S. epidermidis was significantly greater in PCs stored in plasma than in PAS. Although S. liquefaciens altered several PLT quality markers by Days 3 to 4 postinoculation in both PAS- and plasma-PCs, S. epidermidis contamination did not produce measurable PLT changes. CONCLUSIONS: S. liquefaciens can be detected more quickly in PAS-suspended PCs (PAS-PCs) than in plasma-PCs by colony counting. Furthermore, reduced biofilm formation by S. liquefaciens and S. epidermidis during storage in PAS-PCs increases bacteria availability for sampling detection. Culture-based detection remains the earliest indicator of bacterial presence in PAS-PCs, while changes of PLT quality can herald S. liquefaciens contamination when in excess of 10(8) CFUs/mL.

Staphylococcus epidermidis forms biofilms under simulated platelet storage conditions.

BACKGROUND: Staphylococcus epidermidis grows slowly in platelet (PLT) preparations compared to other bacteria, presenting the possibility of missed detection by routine screening. S. epidermidis is a leading cause of nosocomial sepsis, with virulence residing in its ability to establish chronic infections through production of slime layers, or biofilms, on biomedical devices. This study aims to establish biofilm formation (BF) as a mode of growth by S. epidermidis in PLT preparations. STUDY DESIGN AND METHODS: Biofilm-positive (BFpos) and -negative (BFneg) S. epidermidis strains were grown in whole blood-derived PLTs (WBDPs) and in glucose-rich medium (TSBg). An assay for BF was adapted for cultures grown in WBDPs or filtered WBDPs in polystyrene culture plates. Bacterial attachment to polyvinylchloride PLT bags and PLTs was examined by scanning electron microscopy. RESULTS: Both strains display similar growth profiles in WBDPs and TSBg. Unexpectedly, evidence of BF was observed on PLT bags and on PLTs directly, not only by the BFpos strain but also by the BFneg strain. The BFpos strain displayed greater plastic adherence than the BFneg strain in WBDPs (p < 0.05). BF by the BFneg strain was approximately 10-fold greater in WBDPs compared to TSBg (p < 0.05), likely by use of PLTs as a scaffold. Furthermore, BF by S. epidermidis was significantly decreased when PLT concentration was reduced 1000-fold. CONCLUSIONS: S. epidermidis forms biofilms on PLT aggregates and on PLT bags under PLT storage conditions. Our results demonstrate that the PLT storage environment can promote a BF growth mechanism for contaminant bacteria.

Factors Influencing the Survival and Growth of Listeria monocytogenes on the Surface of Canadian Retail Wieners.

A study of survival and growth of Listeria monocytogenes on Canadian retail wieners was undertaken to assess the potential hazard of this product. All-beef, poultry or beef/pork wieners from a total of six processing plants were surface-inoculated with a three-strain cocktail of L. monocytogenes , and stored under vacuum at 5°C for up to 28 days. Of a total of 61 samples tested, 40 (65.6%) supported growth of the pathogen. A model was derived for 35 samples with stepwise multiple regression analysis, which used initial pH (pH0), initial lactic acid bacteria counts (LAB0), and LAB after 14 days (LAB14) to explain 48.1% of the variation in the L monocytogenes counts after 14 days. The observed times to 1-log increase in counts for 25 samples in which growth occurred were compared to times predicted by the U. S. Dept. of Agriculture (USDA) Pathogen Modelling Program. The latter model, based on storage temperature, and initial pH, nitrite and salt concentrations, explained only 12.4% of the variation in the observed times. Scanning electron microscopy showed that the wiener surface was composed of a dense layer of coagulated protein. This may alter the ability of L. monocytogenes to develop relative to growth conditions in the meat formulation. These results suggest that retail wieners may support growth of L monocytogenes , which could present a health hazard to target groups of consumers if subjected to post-processing contamination. They also suggest that further work is needed to derive adequate models to allow an accurate prediction of the growth of L. monocytogenes on ready-to-eat meat products.