Co-Culture of Acinetobacter calcoaceticus-baumannii complex and Staphylococcus saprophyticus Supports Simple Point Contamination Model in Recent Cases of Transfusion-Related Sepsis

2020 ◽  
Vol 154 (Supplement_1) ◽  
pp. S14-S14
Author(s):  
Christopher Kerantzas ◽  
Jacob Merwede ◽  
Edward Snyder ◽  
Jeanne Hendrickson ◽  
Christopher Tormey ◽  
...  

Abstract The CDC recently reported a series of four septic transfusion reactions across three states resulting from contamination of apheresis platelet products. The apheresis platelets were contaminated with strains of both Acinetobacter calcoaceticus-baumannii complex (ACBC) and Staphylococcus saprophyticus (Ss). Two of the reported septic transfusion reactions occurred at our institution. The CDC investigation showed that isolates of each species were genetically related and suggested a point source of contamination. However, the contamination of blood products with ACBC is rare and the co-occurrence of these two species in all four cases was unusual. We hypothesized that there was an augmentative interaction between the clinical isolates of ACBC and Ss from these cases that contributed to their repeated co-occurrence. To test this hypothesis, we compared the growth characteristics of ACBC and Ss when cultured together versus independently. We used isolates from the contaminated platelets for our studies and performed experiments using both solid and liquid growth media. Experiments on solid media assessed density of growth and macroscopic morphology after cross-streaking on Columbia Blood Agar (CBA) and Luria-Bertani (LB) agar. Experiments in liquid media assessed growth by CFU counts in LB broth, platelet-poor plasma, and apheresis platelets. Growth in apheresis platelets was performed using standard, room temperature blood bank platelet storage conditions. Results of these experiments showed a higher CFU concentration of Ss when co-cultured with ACBC in LB broth only after several days, as compared to Ss alone under the same conditions. Otherwise, there was no evidence of augmented growth by either CFU concentration or growth rate in LB broth, plasma, or platelets at other time points. Similarly, there was no evidence of augmented growth by colony density or morphology when cross-streaking strains on either CBA or LB agar. As a result, we conclude that the co-occurrence of these two species in platelets is likely a coincidence of the point contamination suggested by the CDC investigation and not the result of growth augmentation between the two species.

2020 ◽  
Vol 0 (0) ◽  
Author(s):  
Gianmatteo Vit ◽  
Harald Klüter ◽  
Patrick Wuchter

AbstractPlatelet transfusion is a topic of common interest for many specialists involved in patient care, from laboratory staff to clinical physicians. Various aspects make this type of transfusion different from those of other blood components. In this review, the challenges in platelet transfusion practice that are relevant for laboratory colleagues will be discussed, highlighting how the biochemical and structural characteristics of these blood elements directly affect their function and consequently the clinical outcome. More than 1,300 platelet concentrates are transfused in Germany every day, and several types are offered by their respective manufacturers. We describe the technological advances in platelet concentrate production, with a focus on how the storage conditions of platelets can be improved. Laboratory quality assessment procedures for a safe transfusion are discussed in detail. For this purpose, we will refer to the Hemotherapy Directives (Richtlinie Hämotherapie) of the German Medical Association.


Transfusion ◽  
1998 ◽  
Vol 38 (3) ◽  
pp. 229-235 ◽  
Author(s):  
CB Sarkodee-Adoo ◽  
JM Kendall ◽  
R Sridhara ◽  
EJ Lee ◽  
CA Schiffer

1990 ◽  
Vol 68 (1) ◽  
pp. 117-122 ◽  
Author(s):  
Julie Lacasse ◽  
Rosalind S. Labow ◽  
Morris Kates ◽  
George A. Adams

Human platelets are routinely stored for 5 days prior to transfusion, but they deteriorate during storage. Since very little information is available concerning the effect of storage on platelet phospholipid metabolism, the biosynthesis and remodelling of platelet phospholipids were studied. Platelets were incubated separately with [14C]glycerol, [14C]arachidonic acid, or a mixture of [14C]glycerol and [3H]arachidonic acid, and stored in a platelet storage medium at 22 °C. Maximum glycerol uptake (20%) was attained after 6 h. [14C]Glycerol was incorporated into phosphatidylcholine, phosphatidylethanolamine, and phosphatidylinositol, and to a much lesser extent phosphatidylserine, under storage conditions for 5 days. The distribution of the initial arachidonic acid uptake was not as would be expected based on the molar composition of endogenous phospholipids. The arachidonic acid (75%) which was taken up within 10 min of incubation distributed 55% into the phosphatidylcholine and only 14% into the phosphatidylethanolamine; the molar composition is actually 18% phosphatidylcholine and 47% phosphatidylethanolamine. During storage, there was a continuous transfer of the radiolabeled arachidonic acid from phosphatidylcholine to phosphatidylethanolamine until, after 5 days, the distribution of arachidonic acid was identical to the endogenous distribution. In contrast, no change in the glycerol incorporation pattern was detected during storage. This suggested that the mechanism for arachidonic acid redistribution was not through exchange of polar head groups, but through acyl transfer of arachidonic acid from phosphatidylcholine to phosphatidylethanolamine.Key words: human, platelet, storage, arachidonate, phospholipids.


Transfusion ◽  
1997 ◽  
Vol 37 (5) ◽  
pp. 476-481 ◽  
Author(s):  
J Wildt-Eggen ◽  
JG Schrijver ◽  
HJ Bouter-Valk ◽  
R Fijnheer ◽  
M Bins ◽  
...  

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 946-946 ◽  
Author(s):  
Sherrill J. Slichter ◽  
Doug Bolgiano ◽  
Jill Corson ◽  
Mary Kay Jones ◽  
Todd Christoffel ◽  
...  

Abstract Background: With the introduction of bacterial testing, extended storage of platelets is now possible as long as platelet viability is maintained. Although apheresis platelets have recently been licensed for seven days of storage, platelet concentrates are still only licensed for five days. We performed studies to determine the post-storage viability of platelets stored as concentrates. As yet, there are no established criteria for platelet viability that must be met at the end of storage. However, two different criteria for evaluating post-storage platelet viability have been suggested: compare the stored platelet results with each donor’s fresh platelet results; or establish a fixed platelet standard with an expected mean value along with the lower limit of the 95% confidence interval (lower 95% CI) of the mean. Methods: Forty-four normal volunteers donated a unit of whole blood, the whole blood was centrifuged to give platelet-rich plasma (PRP), and the PRP was then centrifuged to produce a platelet concentrate. The platelet concentrates were re-suspended and stored in either 100% plasma or in 20% plasma and 80% Plasmalyte (a platelet storage solution) for five to eight days in Terumo Teruflex bags. At the end of storage, a sample of blood was obtained from each volunteer to prepare fresh platelets to compare the results of a fresh platelet transfusion with the transfusion of the same volunteer’s stored platelets. The fresh and stored platelets were alternately radiolabeled with either 51Cr or 111In, re-injected into their volunteer donor, and serial blood samples were drawn from the volunteer after transfusion of the fresh and stored platelets to determine platelet recoveries and survivals. Results: Results are shown in Table 1. Conclusions: After seven days of storage in plasma, the platelets could qualify for standards of: 74% and 53% as a simple ratio of fresh to stored platelet recoveries and survivals, respectively; according to Dumont’s method of analysis (Transfusion, in press), achievable stored platelet recoveries would be 63% of fresh and survivals would be 40% of fresh; and as a fixed standard, platelet recoveries averaged 45% and survivals 4.4 days with lower 95% CI’s of 35% and 3.3 days, respectively. After eight days of storage in plasma, platelet recoveries and survivals gave unacceptable results. Instead of increasing or at least maintaining post-storage platelet viability, Plasmalyte markedly reduced platelet viability compared to plasma stored platelets at each storage interval. In Vivo Radiolabeled Autologous Platelet Recoveries and Survivals STORAGE CONDITIONS PLATELET RECOVERIES (%) PLATELET SURVIVALS (Days) Solution Time (Days) N Fresh Stored Fresh Stored Data are given as the average ±1 S.D. Plasma 5 10 62±12 55±11 7.7±1.5 6.1±1.0 Plasma 6 10 61±7 46±12 8.2±1.5 5.1±1.6 Plasmalyte 6 3 47±30 29±16 5.1±3.4 1.8±0.2 Plasma 7 10 61±11 45±14 8.3±1.0 4.4±1.5 Plasmalyte 7 5 62±17 30±12 7.4±0.6 2.2±1.1 Plasma 8 3 67±12 31±8 8.3±0.6 2.9±0.8 Plasmalyte 8 3 70±3 21±8 8.2±0.8 0.7±0.1


Author(s):  
Rebecca Brouwers ◽  
Hugh Vass ◽  
Angela Dawson ◽  
Tracey Squires ◽  
Sharareh Tavaddod ◽  
...  

AbstractLaboratory assays such as MIC tests assume that antibiotic molecules are stable in the chosen growth medium - but rapid degradation has been observed for antibiotics including β-lactams under some conditions in aqueous solution. Degradation rates in bacterial growth medium are less well known. Here, we develop a ‘delay time bioassay’ that provides a simple way to estimate antibiotic stability in bacterial growth media. We use the bioassay to measure degradation half-lives of the β-lactam antibiotics mecillinam, aztreonam and cefotaxime in widely-used bacterial growth media based on MOPS and Luria-Bertani (LB) broth. We find that mecillinam degradation can occur rapidly, with a half-life as short as 2 hours in MOPS medium at 37°C and pH 7.4, and 4-5 hours in LB, but that adjusting the pH and temperature can increase its stability to a half-life around 6 hours without excessively perturbing growth. Aztreonam and cefotaxime were found to have half-lives longer than 6 hours in MOPS medium at 37°C and pH 7.4, but still shorter than the timescale of a typical minimum inhibitory concentration (MIC) assay. Taken together, our results suggest that care is needed in interpreting MIC tests and other laboratory growth assays for β-lactam antibiotics, since there may be significant degradation of the antibiotic during the assay.


PLoS ONE ◽  
2021 ◽  
Vol 16 (11) ◽  
pp. e0258934
Author(s):  
Nico Ortlieb ◽  
Elke Klenk ◽  
Andreas Kulik ◽  
Timo Horst Johannes Niedermeyer

Natural products are an important source of lead compounds for the development of drug substances. Actinomycetes have been valuable especially for the discovery of antibiotics. Increasing occurrence of antibiotic resistance among bacterial pathogens has revived the interest in actinomycete natural product research. Actinobacteria produce a different set of natural products when cultivated on solid growth media compared with submersed culture. Bioactivity assays involving solid media (e.g. agar-plug assays) require manual manipulation of the strains and agar plugs. This is less convenient for the screening of larger strain collections of several hundred or thousand strains. Thus, the aim of this study was to develop a 96-well microplate-based system suitable for the screening of actinomycete strain collections in agar-plug assays. We developed a medium-throughput cultivation and agar-plug assay workflow that allows the convenient inoculation of solid agar plugs with actinomycete spore suspensions from a strain collection, and the transfer of the agar plugs to petri dishes to conduct agar-plug bioactivity assays. The development steps as well as the challenges that were overcome during the development (e.g. system sterility, handling of the agar plugs) are described. We present the results from one exemplary screening campaign targeted to identify compounds inhibiting Agr-based quorum sensing where the workflow was used successfully. We present a novel and convenient workflow to combine agar diffusion assays with microtiter-plate-based cultivation systems in which strains can grow on a solid surface. This workflow facilitates and speeds up the initial medium throughput screening of natural product-producing actinomycete strain collections against monitor strains in agar-plug assays.


2014 ◽  
Author(s):  
Jessica Polka ◽  
Pamela A Silver

Bacterial from several taxa, including Kurthia zopfii, Myxococcus xanthus, and Bacillus mycoides, have been reported to align growth of their colonies to small features on the surface of solid media, including anisotropies created by compression. While the function of this phenomenon is unclear, it may help organisms navigate on solid phases, such as soil. The origin of this behavior is also unknown: it may be biological (that is, dependent on components that sense the environment and regulate growth accordingly) or merely physical. Here we show that B. subtilis, an organism which typically does not respond to media compression, can be induced to do so with two simple and synergistically perturbations: a mutation that maintains cells in the swarming (chained) state, and the addition of EDTA to the growth media, which further increases chain length. EDTA apparently increases cell length by inducing defects in cell separation, as the treatment has only marginal effects on the length of individual cells. These results lead us to three conclusions. First, the wealth of genetic tools available to B. subtilis will provide a genetically tractable chassis for engineering compression sensitive organisms in the future. Second, the sensitivity of colony morphology to media compression in Bacillus is a physical rather than biological phenomenon dependent on a simple physical property of rod-shaped cells. And third, colony morphology under compression holds promise as a rapid, simple, and low-cost way to screen for changes in the length of rod-shaped cells or chains thereof.


2019 ◽  
Vol 63 (4) ◽  
Author(s):  
René T. Rozenbaum ◽  
Henny C. van der Mei ◽  
Willem Woudstra ◽  
Ed D. de Jong ◽  
Henk J. Busscher ◽  
...  

ABSTRACTPseudomonas aeruginosacolonizes the sputum of most adult cystic fibrosis patients, forming difficult-to-eradicate biofilms in which bacteria are protected in their self-produced extracellular polymeric substance (EPS) matrices. EPS provide biofilms with viscoelastic properties, causing time-dependent relaxation after stress-induced deformation, according to multiple characteristic time constants. These time constants reflect different biofilm (matrix) components. Since the viscoelasticity of biofilms has been related to antimicrobial penetration but not yet bacterial killing, this study aims to relate killing ofP. aeruginosa, in its biofilm mode of growth, by three antimicrobials to biofilm viscoelasticity.P. aeruginosabiofilms were grown for 18 h in a constant-depth film fermenter, with mucin-containing artificial sputum medium (ASM+), artificial sputum medium without mucin (ASM−), or Luria-Bertani (LB) broth; this yielded 100-μm-thick biofilms that differed in their amounts of matrix environmental DNA (eDNA) and polysaccharides. Low-load compression testing, followed by three-element Maxwell analyses, showed that the fastest relaxation component, associated with unbound water, was most important in LB-medium-grown biofilms. Slower components due to water with dissolved polysaccharides, insoluble polysaccharides, and eDNA were most important in the relaxation of ASM+-grown biofilms. ASM−-grown biofilms showed intermediate stress relaxation.P. aeruginosain LB-medium-grown biofilms was killed most by exposure to tobramycin, colistin, or an antimicrobial peptide, while ASM+provided the most protective matrix, with less water and most insoluble polysaccharides and eDNA. In conclusion, stress relaxation ofP. aeruginosabiofilms grown in different media revealed differences in matrix composition that, within the constraints of the antimicrobials and growth media applied, correlated with the matrix protection offered against different antimicrobials.


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