Screening For Bacterial Contamination Research Articles

Safety Analysis of Extended Platelet Shelf-Life with Large-Volume Delayed Sampling on BACT/ALERT® VIRTUO® in Australia.

Transfusion-transmitted bacterial infection (TTBI) is the leading cause of transfusion-transmitted infections. Platelet components are more likely to be associated with bacterial contamination due to their storage requirements. Australian Red Cross Lifeblood introduced the bacterial contamination screening (BCS) of all platelet components in 2008. The process was recently updated with the use of BACT/ALERT® VIRTUO®, a large-volume delayed sampling (LVDS) protocol and extending platelet shelf-life to seven days. This article describes the results from the routine BCS of platelet components in Australia. Use of VIRTUO has resulted in lower false-positive rates, reducing wastage and improving platelet inventory. Our findings show that the combination of LVDS and VIRTUO improves the safety of platelet transfusions through earlier time to detection, especially for pathogenic bacterial species. Pathogenic bacteria grew within 24 h of incubation with a clear delineation between pathogenic and non-pathogenic species. The data show this protocol is very safe, with no TTBI cases during this time. There were no TTBI reports in recipients of platelet components that subsequently had a positive culture with Cutibacterium species, probably due to the low pathogenic potential of these organisms and slow replication in aerobic platelet bags. We conclude there is no advantage in incubating culture bottles beyond five days.

Proof of Concept for the Detection with Custom Printed Electrodes of Enterobactin as a Marker of Escherichia coli.

The rapid and decentralized detection of bacteria from biomedical, environmental, and food samples has the capacity to improve the conventional protocols and to change a predictable outcome. Identifying new markers and analysis methods represents an attractive strategy for the indirect but simpler and safer detection of pathogens that could replace existing methods. Enterobactin (Ent), a siderophore produced by Escherichia coli or other Gram-negative bacteria, was studied on different electrode materials to reveal its electrochemical fingerprint—very useful information towards the detection of the bacteria based on this analyte. The molecule was successfully identified in culture media samples and a future goal is the development of a rapid antibiogram. The presence of Ent was also assessed in wastewater and treated water samples collected from the municipal sewage treatment plant, groundwater, and tap water. Moreover, a custom configuration printed on a medical glove was employed to detect the target in the presence of another bacterial marker, namely pyocyanin (PyoC), that being a metabolite specific of another pathogen bacterium, namely Pseudomonas aeruginosa. Such new mobile and wearable platforms offer considerable promise for rapid low-cost on-site screening of bacterial contamination.

Bacterial contamination of neglected hospital surfaces and equipment in an Algerian hospital: an important source of potential infection

ABSTRACT Hospital surfaces are heavily contaminated with bacteria, which are a potential source of nosocomial infections. This study was undertaken to identify bacterial communities isolated from neglected hospital surfaces after cleaning routine in a Algerian public hospital. Screening of bacterial contamination in patient bed (PB), reception land-line phones (TF), door handles (DH) and medical equipment (ME) during five months generated 108 inocula. Isolates obtained were identified based on biochemical characteristics and confirmed by analysis of 16S rRNA sequences. Statistical analysis was performed to reveal possible relationship between bacterial diversity and swabbed surfaces. Our findings showed a high prevalence of bacteria in various hospital surfaces, reaching (65.25%), where a highest contaminated surface was the PB (47.22%) and a lowest contaminated was TF (5.55%). Gram negative bacteria were the dominant group (62.03%) mainly represented by Entrobacteriaceae (42.59%), whereas Staphylococcus aureus belonging to Gram positive was the main expanded pathogen with (15.74%).

A Step-by-Step Approach to Improve Clinical Translation of Liposome-Based Nanomaterials, a Focus on Innate Immune and Inflammatory Responses.

This study aims to provide guidelines to design and perform a robust and reliable physical-chemical characterization of liposome-based nanomaterials, and to support method development with a specific focus on their inflammation-inducing potential. Out of eight differently functionalized liposomes selected as “case-studies”, three passed the physical-chemical characterization (in terms of size-distribution, homogeneity and stability) and the screening for bacterial contamination (sterility and apyrogenicity). Although all three were non-cytotoxic when tested in vitro, they showed a different capacity to activate human blood cells. HSPC/CHOL-coated liposomes elicited the production of several inflammation-related cytokines, while DPPC/CHOL- or DSPC/CHOL-functionalized liposomes did not. This work underlines the need for accurate characterization at multiple levels and the use of reliable in vitro methods, in order to obtain a realistic assessment of liposome-induced human inflammatory response, as a fundamental requirement of nanosafety regulations.

Bacterial contamination screening and interpretation for biological laboratory environments

Advances in microbiome researches have led us to the realization that the compositions of bacterial communities of indoor environment is profoundly affected by the function of buildings, and they in turn may bring detrimental effects to the indoor environment and the occupants. These phenomena are common in biological product manufacturing, which might seriously compromise the analytical results. Thus, investigation is warranted for a deeper understanding of the potential impact of the indoor bacterial communities. Among these environments, the biological laboratories stand out because they are relatively clean and yet are highly susceptible to bacterial contaminants. However, few studies have examined the bacterial compositions of biological laboratories. In this study, we assessed the bacterial compositions of samples from the surfaces of various sites across different types of biological laboratories. We have qualitatively assessed these possible bacterial contaminants, and found distinct differences in their bacterial community composition. We also found that the types of laboratories had a larger influence than the sampling sites in shaping the bacterial community, in terms of both structure and richness. On the other hand, the public areas of the different types of laboratories shared very similar sets of bacteria. Tracing the main sources of these bacteria, we identified both environmental and human factors that are important factors in shaping the diversity and dynamics of these possible bacterial contaminations in biological laboratories. These possible bacterial contaminants that we have identified will be helpful for people who aim to eliminate them from samples.

Bacterial contamination screening of blood products

Bacterial contamination screening of blood products

Bacterial safety of blood components–a congress review of the ISBT transfusion‐transmitted infectious diseases working party, bacterial subgroup

The ISBT Working Party on Transfusion‐Transmitted Infectious Diseases, Bacterial Subgroup (TTID‐WP‐BS) held a meeting during the 2017 ISBT Congress covering topics on safety of platelet components (PC), which are highlighted in this review. The TTID‐WP‐BS along with the Paul Ehrlich Institute have pioneered the development of bacterial strain repositories for PC and red blood cells (RBC) to be used for the validation of detection and pathogen reduction (PR) technologies. Results of the development of the RBC repository strains indicated that six bacterial strains were selected for further testing in international studies, complementing the previous study developed for PC. The THERAFLEX UV‐Platelets PR technology has used the bacterial strain repository for PC to test the effectiveness of the system with data presented in this report. Other PR studies described herein include a report of inactivation of Serratia marcescens by the INTERCEPT Blood System for PC at the levels of the PR claim. Furthermore, a summary of current and future developments of the Mirasol PR system was presented with emphasis on PR in whole blood. Advances in PC screening for bacterial contamination were covered by a comparative study of the BacT/ALERT 3D and BacT/ALERT VIRTUO systems, demonstrating that the latter has a shorter time to detection. Finally, data on septic transfusion reactions involving PC produced in plasma and platelet additive solutions indicated different clinical outcomes and screening results. The topics covered during the TTID‐WP‐BS meeting generated insightful scientific discussions and advanced knowledge, one of the most relevant activities of the Bacterial Subgroup.

Detection of bacterial contamination in platelet concentrates from Brazilian donors by molecular amplification of the ribosomal 16S gene.

The aim of our work was to establish a semi-automated high-throughput DNA amplification method for the universal screening of bacteria in platelet concentrates (PCs). Among cases of transfusion transmission of infectious agents, bacterial contamination ranks first in the number of events, morbidity and mortality. Transmission occurs mainly by transfused PCs. Automated culture is adopted by some blood banks for screening of bacterial contamination, but this procedure is expensive and has a relatively long turnaround time. PCs were spiked with suspensions of five different bacterial species in a final concentration of 1 and 10 colony-forming units (CFU) per millilitre. After incubation, the presence of bacteria was investigated by real-time polymerase chain reaction (PCR) and by the Enhanced Bacterial Detection System (eBDS, Pall) assay as a reference method. Real-time PCR amplification was performed with a set of universal primers and probes targeting the 16S rRNA gene. Co-amplification of human mitochondrial DNA served as an internal control. Using the real-time PCR method, it was possible to detect the presence of all bacterial species tested with an initial concentration of 10CFU mL-1 24 h after contamination, except for Staphylococcus hominis. The PCR assay also detected, at 24 h, the presence of Serratia marcescens and Enterobacter cloacae with an initial concentration of 1CFU mL-1 . The real-time PCR assay may be a reliable alternative to conventional culture methods in the screening of bacterial contamination of PCs, enabling bacterial detection even with a low initial concentration of microorganisms.

Routine use of Day6 and Day7 platelets with rapid testing: two hospitals assess impact 1year after implementation.

In March 2016 the US Food and Drug Administration published a draft guidance to enhance the safety of platelets (PLTs) for transfusion. Options for hospital transfusion services include the use of rapid testing to extend apheresis PLT dating for up to 7 days. This report describes the impact of routine use of Day 6 and Day 7 PLTs at two hospital transfusion services 1 year after implementation of rapid testing for outdate extension. PLT transfusion and inventory data were obtained from two hospital-based transfusion services for 12months before and 12months after implementation of rapid testing to extend the outdate of apheresis PLTs to 7 days. The outdate rate decreased from 5% to 2% (p<0.0001) at Hospital 1 and 28% to 14% (p<0.001) at Hospital 2 after implementation of routine use of Day 6 and Day 7 PLTs. The proportion of apheresis PLT units that underwent secondary screening for bacterial contamination before transfusion in the postimplementation period increased from 33% to 54% at Hospital 1 and from 0% to 31% at Hospital 2. A significant decrease in outdate rate was observed after routine use of Day 6 and Day 7 PLTs. Use of rapid testing to extend PLT outdate also resulted in a larger proportion of PLTs that underwent secondary testing for bacterial contamination before transfusion. These observations demonstrate that use of rapid testing to extend apheresis PLT dating up to 7 days enhances the safety of PLTs for transfusion and decreases wastage of a limited resource.

Bacterial testing of platelets - has it prevented transfusion-transmitted bacterial infections in Australia?

Australia introduced bacterial contamination screening (BCS) for platelet components in April 2008. This study presents analysis performed to assess the efficacy of testing. Seven-day aerobic and anaerobic culture is performed using the BacT/ALERT 3D system. Following an initial machine positive (IMP) flag, all associated components are recalled, and/or clinicians treating already transfused patients are notified. IMPs are categorized as 'machine false positive', 'confirmed positive' or 'indeterminate' depending on culture results of initial and repeat samples. Between 2010 and 2012, 1·1% of platelet donations tested IMP; since 2013, this rate has fallen to 0·6% through improved instrument management, reducing false-positive IMPs but maintaining sensitivity for cultures yielding bacterial growth. On average, 66% of confirmed positive and indeterminate platelet units had been transfused at the time of detection. The majority (95%) of these grew Propionibacterium sp., a slow-growing organism that rarely causes sepsis in the transfusion setting. The incidence of reported transfuion-transmitted bacterial infection (TTBI) has fallen since the introduction of BCS, with a 4·2-fold [0·5, 28·2] lower rate from platelets. BCS has been successful in detecting platelet units containing pathogenic bacteria. The incidence of TTBI from platelets has fallen since the introduction of BCS, but the risk has not been eliminated due to rare false-negative results. In the absence of a pathogen inactivation system for red blood cells, BCS provides 'surrogate' testing of red blood cells from which platelets have been manufactured.

Survey for bacterial testing of platelet components in Latin America

IntroductionBacterial contamination of platelet concentrates (PCs) poses the highest post‐transfusion infectious risk in developed countries. However, there is not extensive information about similar strategies implemented in Latin America.AimTo assess the status of platelet screening for bacterial contamination in Latin American blood banks.MethodsAn online‐based survey with 10 comprehensive questions was sent to 46 blood banks in five countries: Argentina, Brazil, Colombia, Honduras and Mexico. The centres were asked about PC production and shelf life, and strategies to improve PC safety. Centres were also surveyed regarding postsampling quarantine, results interpretation and haemovigilance data.ResultsTen centres(21·8%) in four countries answered the survey. Annual PC production ranges from 600 to 19 200. All sites store PCs for 5 days, and eight sites use leucocyte reduction. Five centres(50%) produce apheresis and platelet‐rich‐plasma PCs, one centre(10%) produces buffy coat (BC) PCs, one centre(10%) produces apheresis PCs, and three centres(30%) produce apheresis and BC PCs. Nine sites reported using established donor skin donor disinfection protocols, eight employ first aliquot diversion, and two visually inspect PCs. Eight sites screen PCs for bacterial contamination with culture methods: three sites(37·5%) test 1% of outdated PCs and five sites(62·5%) test 100% of PCs 20‐24 h after blood collection. Of these five sites, two quarantine PCs for 24 h after sampling. Three sites have haemovigilance data, which are unavailable to the public. No sites have implemented pathogen reduction technologies.ConclusionsScreening practices and mitigation strategies vary considerably between sites highlighting the need for standardisation of procedures in Latin America.

The infectious disease blood safety risk of Australian hemochromatosis donations.

It has been suggested that blood donors with hereditary hemochromatosis may pose an increased infectious disease risk and adversely affect recipient outcomes. This study compares the infectious disease risk of whole blood (WB) donors enrolled as therapeutic (T) donors to voluntary WB donors to evaluate the safety of blood products provided by the T donors. This was a retrospective cohort study of all WB donations at the Australian Red Cross Blood Service who donated between January 1, 2011, and December 31, 2013, comparing a yearly mean of 11,789 T donors with 107,773 total donations and a yearly mean of 468,889 voluntary WB donors with 2,584,705 total donations. We compared postdonation notification of infectious illnesses, bacterial contamination screening results, and positive tests for blood borne viruses in T and WB donors. Rates of transfusion-transmissible infections in donations destined for component manufacture were significantly lower in therapeutic donations compared to voluntary donations (8.4 vs. 21.6 per 100,000 donations). Bacterial contamination (43.0 vs. 45.9 per 100,000 donations) and postdonation illness reporting (136.2 vs. 110.8 per 100,000 donations) were similar in both cohorts. The Australian therapeutic venisection program enables T donors to provide a safe and acceptable source of donated WB that has a low infectious disease risk profile.

An Internal Reference Control Duplex Real-Time Polymerase Chain Reaction Assay for Detecting Bacterial Contamination in Blood Products.

Real-time polymerase chain reaction (RT-PCR) enables effective and sensitive screening for infectious risk in the field of blood safety. However, when using RT-PCR to detect bacterial contamination, several intractable points must be considered, one of which is the lack of appropriate quality control. In this study, we developed a simplified RT-PCR assay in which the same primer set and two distinct probes were used to detect both, an internal reference control and the target in a reaction. The copy number of the internal reference control represents the positive detection limit of the assay; therefore, when the threshold-cycle value of the target is less than or equal to that of the internal reference control, the result obtained for the target can be considered to be a true positive. When human gDNA was spiked with Escherichia coli gDNA and the detection limit for the internal reference control was set to five copies, the measured detection limit for E. coli gDNA was two copies. The internal reference control duplex RT-PCR assay showed high efficiency (0.91–1.02), high linearity (R2 > 0.99), and good reproducibility in intra- and inter-assay comparisons. Lastly, when human platelet-rich plasma samples were spiked with E. coli or other bacterial species, all species were detected efficiently, and the results of a two-sample pooled t test showed that the limit of detection for E. coli was 1 cfu/mL. Here, we present a synthetic internal reference control molecule and a new statistical method for improving the reliability of RT-PCR assays when screening for bacterial contamination in blood products.

Coherent Fluctuation Nephelometry: A Rapid Method for Urine Screening for Bacterial Contamination.

Express-test by the method of coherent fluctuation nephelometry for urine contamination was carried out on two prototype instruments with standard polystyrene photometric cuvettes. We analyzed 209 and 119 urine samples. Due to high sensitivity of the method, up to 50% negative samples were detected within 10 min by initial opacity and 90% negative samples were detected during 3.5 h by registration of the bacterial growth curves.

Platelet concentrates: reducing the risk of transfusion-transmitted bacterial infections

Platelet concentrates: reducing the risk of transfusion-transmitted bacterial infections Dirk de Korte,1 Jan H Marcelis2 1Department of Product and Process Development, Sanquin Blood Bank, Amsterdam, 2Department of Microbiology, St Elisabeth Hospital, Tilburg, the Netherlands Abstract: The introduction of a combination of interventions during collection of whole-blood or platelet concentrates has been successful in lowering the degree of bacterial contamination in the final product, the platelet concentrate, by 50%&ndash;75%. These interventions were improved donor questionnaires, best-practice skin disinfection, and diversion of first blood volume. These interventions have reduced the number of bacteria present in the platelet concentrates. In combination with screening for bacterial contamination of platelet concentrates with a culture method, the degree of transfusion-transmitted bacterial infection has been reduced significantly. Due to the very low initial bacteria counts upon collection of the products, the need for improved sensitivity of early screenings tests or highly selective point-of-issue tests remains. The latter should be rapid and easy to perform. An alternative approach might be the implementation of pathogen-inactivation methods for cellular blood products to reduce the amount of pathogens. However, these methods are costly, and so far not proved to be cost-effective, especially in countries with an already-low incidence of transfusion-transmitted infections by viruses, parasites, or bacteria. Keywords: blood products, bacterial contamination, screening, point of issue, pathogen inactivation

Current status of bacterial detection in blood components – successes and challenges

Background Blood safety and the prevention of transfusion‐transmitted pathogens are among the principal aims of transfusion medicine. Based on the many improvements over the last decade in donor selection programs, the implementation of nucleic acid testing (NAT) and the introduction of third‐ and fourth‐generation serology assays into blood donor screening, the residual risk of transfusion‐transmitted viruses has been reduced to a minimum, and the quality and safety of blood components are at their highest level ever. In contrast, the residual risk of bacterial transmissions, especially in platelet concentrates, is an on‐going challenge in transfusion medicine. To improve blood safety regarding bacterial contaminations, many countries have implemented bacterial culture methods, in combination with the negative‐to‐date concept. This strategy has improved blood safety and reduced transfusion‐transmitted fatalities due to bacterial infections. Nevertheless, false‐negative screening results have been reported in some cases. The current manuscript summarises the clinical and experimental data based on culture methods and rapid bacterial detection systems.Methods Many countries have implemented bacterial culture methods (BacT/ALERT, BACTEC or PAL eBDS), in combination with the negative‐to‐date concept. Differences between countries have been recognised regarding testing with aerobic bottles alone or with aerobic and anaerobic bottles and regarding sample input volumes. Therefore, international comparisons for the prevalence and incidence of contamination rates are different between countries. Within the last few years, a couple of rapid bacterial detection methods have been developed in experimental spiking studies. The most promising methods are FACS systems, such as Bactiflow, impedance measurements, pan generic detection, and automated generic 16s nucleic amplification testing (NAT).Results Culture methods, in combination with the negative‐to‐date concept, have been implemented in many countries as a ‘gold standard’ to improve blood safety. Analytical sensitivity could be increased by using aerobic and anaerobic bottles, as well as by using an input volume of 8–10 ml per bottle. Countries using both screening bottle types have detected Probionibacterium acnes in the majority of cases. False‐negative screening results with fatal outcomes have been prevented. In contrast, rapid bacterial detection methods have been investigated in spiking studies. Clinical data are available in Germany for Bactiflow and 16s NAT. The analytical sensitivity of rapid bacterial detection methods is 1–2 log phases less than for culture methods.Conclusions In addition to diversion of the first 30–40 ml, reduction of platelet shelf life and the implementation of pathogen‐reduction methods, blood donor screening for bacterial contamination in platelets has been able to improve blood safety and to prevent fatal septic reactions. Different rapid bacterial detection methods are currently feasible for routine screening and to enable new opportunities. The negative‐to‐date concept, with the residual risk of sampling errors, might be replaced by the rapid release of untested platelets on day 1 and day 2 after donation, in combination with a late sampling‐screening model on day 3 after donation with rapid bacterial detection systems. Clinical data for the newly developed rapid detection methods are eagerly awaited.

Evaluation of an Automated Urine Flow Cytometer for Screening of Bacterial Contamination in Platelet Concentrates

Evaluation of an Automated Urine Flow Cytometer for Screening of Bacterial Contamination in Platelet Concentrates

Performance and suitability of polymerase chain reaction for early detection of bacteria in platelet concentrates

In this study the applicability of a 16S rRNA real-time reverse transcriptase polymerase chain reaction (RT-PCR) and a Staphylococcus genus-specific PCR for screening of bacterial contamination in platelet concentrates (PCs) was determined. A total of 336 sample bags, from PCs that were routinely tested in the BacT/ALERT (bioMérieux), were collected and frozen until testing by the PCR assays. Based on the BacT/ALERT results, 107 PCs were positive and 229 were negative for bacterial growth. The analytical sensitivity of the 16S rRNA real-time RT-PCR ranged from 5 to 40 colony-forming units (CFUs)/mL. The PCR detected five positive samples, four of which were also positive in the BacT/ALERT. The sensitivity of the test was 3.8%, and the specificity was 99.5%. The analytical sensitivity of the Staphylococcus genus-specific PCR ranged from 5 to 15 CFUs/mL. Thirty-nine units that were BacT/ALERT positive for staphylococci were tested with this PCR. Six samples were positive with the PCR, five of which were also BacT/ALERT positive. The sensitivity of the Staphylococcus genus-specific PCR was 12.8%, and the specificity was 98.8%. Despite the rapid availability of results compared to the BacT/ALERT, the analytical sensitivity of a generic or specific PCR assay is not high enough to be an alternative for the BacT/ALERT when PCs are screened on the day of production.

10 Years Experience with Bacterial Screening of Platelet Concentrates in the Netherlands

Background: Contamination of platelets with bacteria is the major microbiological risk of blood transfusion. Screening for bacterial contamination can reduce the frequency of bacterial transmission considerably. In the present paper, the results of 10-year screening in the Netherlands are described. Methods: All platelet concentrates were cultured with the BacT/Alert culturing system with large volume (7.5 ml) cultures in either an aerobic or an anaerobic bottle. Products were released on a ‘negative-to-date’ basis. Results: After introduction of the diversion of the first milliliters of collected blood, the number of positive screening cultures decreased significantly from 0.85% to 0.37%. The frequency of transfusion-transmitted bacterial infections by platelet concentrates is currently less than 1 per 2 years in the Netherlands. Conclusion: Over a period of 10 years the bacterial screening system for platelet concentrates proved to result in a safe system with respect to microbiological infection as a result of platelet transfusions.

Bacterial Pre-Release Testing of Platelets – the Australian Red Cross Blood Service Clinical Experience

The risk of bacterial transmission by platelet transfusion has been recognised internationally as the leading residual infections transfusion risk in the last decade. We describe the clinical and logistical aspects of bacterial contamination screening of platelets introduced in Australia in early 2008. Sampling occurs at 24 h, and platelets are released to hospitals ‘negative to date’. Bacterial screening detection of initial machine-positive (IMP) and all follow-up results are notified to transfusing laboratories. Results of screening between 2008 and 2010 found a significant rate of IMP samples (1.06%) with a true- positive/indeterminate rate of 0.18%. Components were already transfused in 32.5% of cases at time of initial notification. Confirmed cases of septic transfusion reactions have declined significantly since the introduction of pre- release platelet screening, reflecting an important additional improvement in transfusion safety in Australia.