Abnormal increases in reactive oxygen species in dying insects infected with nematodes.

Stress enhances the concentration of reactive oxygen species (ROS) in animal plasma. Increased ROS alter various physiological functions, such as development and the immune response, but excessive increases could be harmful. In this study, we tested the hypothesis that abnormally increased plasma ROS levels are associated with animal death. Injection of the nematode Caenorhabditis elegans into insect larvae caused high mortality in Galleria mellonella, and the plasma ROS concentration was four times higher than M9 buffer-injected larvae. There was no difference in plasma antioxidant activity after nematode injection. However, coinjecting nematodes with an antioxidant (ascorbic acid or N-acetylcysteine) suppressed increases in ROS concentrations by the nematodes and increases in the number of nematodes in the larvae, which increased G. mellonella survival. These results suggest that the abnormal elevation of ROS associated with the stress caused by nematode propagation is lethal for G. mellonella.

Inactivation of a broad spectrum of viruses and parasites by photochemical treatment of plasma and platelets using amotosalen and ultraviolet A light.

BACKGROUND: The INTERCEPT Blood System pathogen reduction technology (PRT), which uses amotosalen and ultraviolet A light treatment (amotosalen/UV-PRT), inactivates pathogens in plasma and platelet components (PCs). This review summarizes data describing the inactivation efficacy of amotosalen/UVA-PRT for a broad spectrum of viruses and parasites. METHODS: Twenty-five enveloped viruses, six nonenveloped viruses (NEVs), and four parasites species were evaluated for sensitivity to amotosalen/UVA-PRT. Pathogens were spiked into plasma and PC at high titers. Samples were collected before and after PRT and assessed for infectivity with cell cultures or animal models. Log reduction factors (LRFs) were defined as the difference in infectious titers before and after amotosalen/UV-PRT. RESULTS: LRFs of ≥4.0 log were reported for 19 pathogens in plasma (range, ≥4.0 to ≥7.6), 28 pathogens in PC in platelet additive solution (PC-PAS; ≥4.1-≥7.8), and 14 pathogens in PC in 100% plasma (PC-100%; (≥4.3->8.4). Twenty-five enveloped viruses and two NEVs were sensitive to amotosalen/UV-PRT; LRF ranged from >2.9 to ≥7.6 in plasma, 2.4 or greater to greater than 6.9 in PC-PAS and >3.5 to >6.5 in PC-100%. Infectious titers for four parasites were reduced by >4.0 log in all PC and plasma (≥4.9 to >8.4). CONCLUSION: Amotosalen/UVA-PRT demonstrated effective infectious titer reduction for a broad spectrum of viruses and parasites. This confirms the capacity of this system to reduce the risk of viral and parasitic transfusion-transmitted infections by plasma and PCs in various geographies.

Plasma parasitemia as assessed by quantitative PCR in relation to clinical disease severity in African adults with falciparum malaria with and without HIV co-infection.

PURPOSE: When considering malaria disease severity, estimation of parasitemia in erythrocytes is important, but sometimes misleading, since the infected erythrocytes may be sequestered in peripheral capillaries. In African children and Asian adults with falciparum malaria, parasitemia as assessed by quantitative PCR (qPCR) in plasma seems to be a valuable indicator of disease severity, but data on African adults as well as the impact of co-infection with HIV is lacking. METHODS: In 131 patients with falciparum malaria in a public tertiary teaching hospital in Mozambique, plasma malaria parasitemia as assessed by qPCR, compared to qualitative malaria PCR in blood cell fraction, was related to malaria disease severity and HIV co-infection. RESULTS: Of the 131 patients with falciparum malaria, based on positive qualitative PCR in the blood cell fraction, 93 patients (72%) had positive malaria qPCR in plasma. Patients with severe malaria as defined by the WHO criteria had higher malaria quantitative plasma parasitemia (median 143 genomes/µL) compared to those with uncomplicated malaria (median 55 genomes/µL, p = 0.037) in univariate analysis, but this difference was attenuated after adjusting for age, sex and HIV co-infection (p = 0.055). A quarter of the patients with severe malaria had negative qPCR in plasma. CONCLUSIONS: This study of adult African in-patients with falciparum malaria with and without HIV co-infection, neither confirms nor rejects previous studies of malaria qPCR in plasma as an indicator of disease severity in patients with falciparum malaria. There is a need for further and larger studies to clarify if parasitemia as assessed malaria qPCR in plasma could be a surrogate marker of disease severity in falciparum malaria.

Metagenomics in pooled plasma, with identification of potential emerging infectious pathogens.

BACKGROUND: Many emerging infectious pathogens are well known for existing in healthy blood donors and could be transmitted via blood transfusion or plasma derivatives usage. Therefore, there is an urgent need to discover the pathogens in qualified blood donation to avoid potential threats to blood safety. STUDY DESIGN AND METHODS: The objective of this study was to investigate the microbiome that existed in pooled plasma from different manufacturers in Chengdu and Guiyang. Random polymerase chain reaction, large-scale clone sequencing, and bioinformatics were used to investigate the metagenomics and microbiome structure of pooled plasma. Among detected microbiomes, potential pathogens were subsequently identified. RESULT: After host DNA cleaning, 551 clones were classified as bacteria; 88 clones were classified as viruses, and four clones were considered to be parasites, respectively. Thirteen kinds of bacteria and two kinds of parasites that might potentially threaten blood safety were identified along with six kinds of nonpathogenic viruses. The infection status of one identified pathogen Coxiella burnetii was evaluated in 1638 plasma samples. The reactive rate of immunoglobulin (Ig)G1 was 1.10% (18/1638), the reactive rate of IgG2 was 0.85% (14/1638), and the reactive rate of IgM was 0.98% (16/1638). CONCLUSION: Some pathogens that were already considered as threats to blood safety were discovered in those pooled plasma such as C. burnetii, Orientia tsutsugamushi, and Plasmodium sp. As a result, we should initiate some specific tests in the endemic area on plasma donors to enhance the blood safety in China.

Protein expression profiling in haemocytes and plasma of the Manila clam Ruditapes philippinarum in response to infection with Perkinsus olseni.

The protein expression profiling in clam haemocytes and plasma in response to Perkinsus olseni was addressed. Adult Manila clams from a P. olseni-free bed were experimentally challenged with parasite zoospores to analyse immune response. In another experiment, the effects of longer term infection were assessed in adult clams collected from a P. olseni-affected bed, by comparing moderate to very heavily infected clams with non-infected ones. Haemocyte and plasma proteins were separated by two-dimensional electrophoresis; spot patterns were qualitatively compared between treatments within each experiment and the spots indicating differential protein expression associated with P. olseni challenge or with field infection were processed for protein identification. Fifteen clam proteins (four in haemocytes and eleven in plasma) of which expression was markedly affected by P. olseni were identified. Some of the identified proteins have a well-known role in clam immune response against the parasite, such as lysozyme and lectins. Rho GTPase-activating protein 6 could be a marker of resistance against P. olseni, which should be further studied.

Multiplex detection and identification of viral, bacterial, and protozoan pathogens in human blood and plasma using a high-density resequencing pathogen microarray platform.

BACKGROUND: The implementation of nucleic acid-based tests for blood donor screening has improved the safety of the blood supply; however, the increasing number of emerging pathogen tests is burdensome. Development of multiplex testing platforms that allow simultaneous screening for different pathogens is a potential solution. STUDY DESIGN AND METHODS: The TessArray resequencing microarray is a platform that allows multiplex detection and identification of 97 different blood-borne pathogens in one single test. The objective was to evaluate the lowest concentration detected in blood or plasma, species discrimination, and applicability of the TessArray microarray platform for testing blood donors. Human blood or plasma spiked with selected pathogens (10,000, 1000, or 100 cells or copies/mL), including three viral, four bacterial, and four protozoan pathogens were each tested on this platform. The nucleic acids were extracted, amplified using multiplexed sets of pooled specific primers, fragmented, labeled, and hybridized to a microarray. Finally, the detected sequences were identified using an automated genomic database alignment algorithm. RESULTS: The performance of this platform demonstrated detection for spiked bacterial and protozoan pathogens of 100 cells/mL and viral pathogens as low as 100 copies/mL. Coded specimens, including spiked and negative controls, were identified correctly for blood specimens (31/32, 97%) and for plasma specimens (20/22, 91%) demonstrating the effectiveness of the platform. CONCLUSION: These results indicated that the TessArray microarray platform could be employed for multiplex detection and identification, with a high level of discriminatory power for numerous blood-borne pathogen targets with potential for use in blood safety.

A high-efficiency superhydrophobic plasma separator.

To meet stringent limit-of-detection specifications for low abundance target molecules, a relatively large volume of plasma is needed for many blood-based clinical diagnostics. Conventional centrifugation methods for plasma separation are not suitable for on-site testing or bedside diagnostics. Here, we report a simple, yet high-efficiency, clamshell-style, superhydrophobic plasma separator that is capable of separating a relatively large volume of plasma from several hundred microliters of whole blood (finger-prick blood volume). The plasma separator consists of a superhydrophobic top cover with a separation membrane and a superhydrophobic bottom substrate. Unlike previously reported membrane-based plasma separators, the separation membrane in our device is positioned at the top of the sandwiched whole blood film to increase the membrane separation capacity and plasma yield. In addition, the device's superhydrophobic characteristics (i) facilitates the formation of well-defined, contracted, thin blood film with a high contact angle; (ii) minimizes biomolecular adhesion to surfaces; (iii) increases blood clotting time; and (iv) reduces blood cell hemolysis. The device demonstrated a "blood in-plasma out" capability, consistently extracting 65 ± 21.5 µL of plasma from 200 µL of whole blood in less than 10 min without electrical power. The device was used to separate plasma from Schistosoma mansoni genomic DNA-spiked whole blood with a recovery efficiency of >84.5 ± 25.8%. The S. mansoni genomic DNA in the separated plasma was successfully tested on our custom-made microfluidic chip by using loop mediated isothermal amplification (LAMP) method.

The prevalence of toxoplasmosis in Imam Reza Hospital blood bank samples, Tehran, Iran.

BACKGROUND: The prevalence of toxoplasma gondii (T.g) infection in blood donors has been poorly studied. The aim of this study was to assess the prevalence of acute and chronic toxoplasmosis in blood products. METHODS: A total of 223 blood products (101 fresh frozen plasma (FFP) and 122 packed cells (PC)) in Imam Reza hospital blood bank, Tehran, Iran were tested for specific T.g antibodies (IgG and IgM) by ELISA method. Positive IgG anti-T.g samples were further tested for IgM anti-T.g. A positive IgG test with the negative and positive IgM test was interpreted as a chronic and acute toxoplasmosis respectively. RESULTS: Of 223 samples 38.6% and 0.45% were positive for IgG anti-T.g and IgM anti-T.g levels respectively. Therefore, one and 85 samples were involved acute and chronic toxoplasmosis respectively. Twenty-six of fresh frozen plasma samples were positive for IgG anti-T.g and one of them was positive for IgM anti-T.g. Sixty packed cell samples were positive for IgG anti-T.g. CONCLUSIONS: Our study showed that there were chronic and acute toxoplasmosis in blood products and the prevalence of toxoplasmosis especially chronic form was high. Therefore screening of blood for T.g antibodies may be considered.

Performance of parasitological and molecular techniques for the diagnosis and surveillance of gambiense sleeping sickness.

OBJECTIVES: Recently, improvements have been made to diagnostics for gambiense sleeping sickness control but their performance remains poorly documented and may depend on specimen processing prior to examination. In a prospective study in the Democratic Republic of the Congo, we compared the diagnostic performance of several parasite detection techniques, immune trypanolysis and of m18S PCR on whole blood stored in a stabilisation buffer or dried on filter paper. METHODS: Individuals with CATT whole blood (WB) titer ≥1∶4 or with clinical signs indicative for sleeping sickness were examined for presence of trypanosomes in lymph node aspirate (LNA) and/or in blood. Blood was examined with Capillary Centrifugation Technique (CTC), mini-Anion Exchange Centrifugation Technique (mAECT) and mAECT on buffy coat (BC). PCR was performed on whole blood (i) stored in guanidine hydrochloride EDTA (GE) stabilisation buffer and (ii) dried on filter paper, and repeatability and reproducibility were assessed. Immune trypanolysis (TL) was performed on plasma. RESULTS: A total of 237 persons were included. Among 143 parasitologically confirmed cases, 85.3% had a CATT-WB titre of ≥1/8, 39.2% were positive in LNA, 47.5% in CTC, 80.4% in mAECT-WB, 90.9% in mAECT-BC, 95.1% in TL and up to 89.5% in PCR on GE-stabilised blood. PCR on GE-stabilised blood showed highest repeatability (87.8%) and inter-laboratory reproducibility (86.9%). Of the 94 non-confirmed suspects, respectively 39.4% and 23.4% were TL or PCR positive. Suboptimal specificity of PCR and TL was also suggested by latent class analysis. CONCLUSION: The combination of LNA examination with mAECT-BC offered excellent diagnostic sensitivity. For PCR, storage of blood in stabilisation buffer is to be preferred over filter paper. TL as well as PCR are useful for remote diagnosis but are not more sensitive than mAECT-BC. For TL and PCR, the specificity, and thus usefulness for management of non-confirmed suspects remain to be determined.

Inactivation of Plasmodium spp. in plasma and platelet concentrates using riboflavin and ultraviolet light.

BACKGROUND: Photochemical treatment of blood products could help prevent transfusion-transmitted malaria and reduce the need for donor deferrals. In this study we evaluated the effectiveness of riboflavin and ultraviolet (UV) light against both Plasmodium falciparum, which causes the most severe form of human malaria, and Plasmodium yoelii, an in vivo murine model for malaria. STUDY DESIGN AND METHODS: Plasma and platelet (PLT) concentrates were inoculated with either P. falciparum- or P. yoelii-infected red blood cells (RBCs). Aliquots from each unit were collected after inoculation, after addition of riboflavin, and after treatment. In vitro P. falciparum growth was assessed using thin blood films of duplicate samples at 24, 48, 72, and 96 hours. P. yoelii parasitemia was followed in mice for 14 days postinoculation. RESULTS: In the in vitro studies, the mean P. falciparum parasitemia increased 12- to 19-fold in pretreatment samples, both before and after addition of riboflavin, after 96-hour culture. Few parasites were observed in Mirasol-treated units at 24 hours; those that were observed were degenerating. Through the remainder of the 96-hour culture period, cultures of treated samples were negative. In the in vivo study, mouse plasma containing P. yoelii-infected RBCs had a mean starting titer of 4.6 log mouse infectious dose 50%/mL. No infectious parasite was detected in treated samples. CONCLUSION: Treatment with riboflavin and UV light was effective at reducing viable P. falciparum in both PLT and plasma products by at least 3.2 logs. Additionally, an at least 4.4-log reduction was observed with P. yoelii.

The application of photosensitisers to tropical pathogens in the blood supply.

The onset of the HIV pandemic led both to significant alterations in blood collection and screening practice and to the development of more sophisticated methods of inactivation of infectious agents from the blood supply. Photodynamic (i.e. light activated) pathogen inactivation is one such method currently in limited use in various European states. The approach is based on the generation of a burst of reactive oxygen and nitrogen species, resulting in the activation of several cell death mechanisms. However, its application to tropical pathogens is perhaps less appreciated, despite the fact that the efficacies of photoantimicrobial agents such as methylene blue were originally reported following screening against organisms such as Trypanosoma cruzi and viruses such as those responsible for dengue and yellow fever. Since the objective of pathogen inactivation is to remove both established and emerging infective agents, it is necessary for photoantimicrobial agents to be broad-spectrum in activity. While this is demonstrable in plasma and platelet fractions, the application to red blood cells is currently under investigation.

[Photochemical inactivation of pathogens in platelets and plasma: five years of clinical use in routine and hemovigilance. Towards a change of paradigm in transfusion safety]. / Inactivation photochimique des pathogènes des plaquettes et du plasma: cinq ans d'utilisation clinique de routine et d'hémovigilance. Vers un changement de paradigme de la sécurité en transfusion.

The transfusion of labile blood products is vital and essential for patients in absence of alternative treatment. Patients and doctors have always feared transfusion-transmitted infections by blood, blood components and blood-derived drugs. Photochemical inactivation of platelet concentrates and plasma, using a technique associating amotosalen and UVA, has been used for five years in a French region for the whole population and a large spectrum of patients, with efficacy and safety. It would seem wise to introduce labile blood products, submitted to pathogen inactivation by a technique already approved by a regulatory agency and not to wait for a perfect system including red blood cells concentrates. Universal implementation of pathogen inactivation in labile blood products is a major and key step to improve safety against infection in transfusion.

Augmented plasma microparticles during acute Plasmodium vivax infection.

BACKGROUND: In the last few years, the study of microparticles (MPs)--submicron vesicles released from cells upon activation or apoptosis--has gained growing interest in the field of inflammation and in infectious diseases. Their role in the human malaria parasite Plasmodium vivax remains unexplored. Because acute vivax malaria has been related to pro-inflammatory responses, the main hypothesis investigated in this study was that Plasmodium vivax infection is associated with elevated levels of circulating MPs, which may play a role during acute disease in non-immune patients. METHODS: Plasma MPs were analysed among thirty-seven uncomplicated P. vivax infections from an area of unstable malaria transmission in the Brazilian Amazon. The MP phenotype was analysed by flow cytometry using the classical MP marker, annexin, and fluorochrome-labeled monoclonal antibodies against specific cell surface markers. The frequencies of plasma MPs in P. vivax patients (n=37) were further compared to malaria-unexposed controls (n=15) and ovarian carcinoma patients (n=12), a known MPs-inducing disease non-related to malaria. RESULTS: The frequencies of plasma circulating MPs were markedly increased in P. vivax patients, as compared to healthy age-matched malaria-unexposed controls. Although platelets, erythrocytes and leukocytes were the main cellular sources of MPs during vivax malaria, platelet derived-MPs (PMPs) increased in a linear fashion with the presence of fever at the time of blood collection (ß=0.06, p<0.0001) and length of acute symptoms (ß=0.36, p<0.0001). Finally, the results suggest that plasma levels of PMPs diminish as patient experience more episodes of clinical malaria (ß=0.07, p<0.003). CONCLUSIONS: Abundant circulating MPs are present during acute P. vivax infection, and platelet derived-MPs may play a role on the acute inflammatory symptoms of malaria vivax.

Transmission of Trypanosoma cruzi by heart transplantation.

BACKGROUND: Trypanosoma cruzi infection (i.e., Chagas disease) is an unusual complication that can occur after solid-organ transplantation and that can result in severe illness or death. In 2006, there were 2 heart transplant recipients in Los Angeles, California, reported to have acute trypanosomiasis during the same month. We conducted an investigation to determine the source of these infections. METHODS: We reviewed the medical, organ procurement, and donor transfusion and transplantation records of these 2 heart transplant recipients. The 2 heart transplant recipients were interviewed regarding any kind of natural exposure and were screened for parasites by obtaining blood and other tissue samples for buffy coat, culture, and polymerase chain reaction. Serum samples from the heart transplant recipients, organ donors, and blood donors were tested for T. cruzi antibodies by use of immunofluorescence assay and radioimmunoprecipitation assay. Tissue samples from the organ donors were examined by use of polymerase chain reaction and immunohistochemical staining. Other recipients of organs from the same donors were monitored for T. cruzi infection by use of polymerase chain reaction and immunofluorescence assay. RESULTS: Both heart transplant recipients had no apparent risk factors for preexisting T. cruzi infection. Both were seronegative but tested positive for the parasite, indicating recent infection. Both recipients died despite medical treatment. The organ donors tested positive for T. cruzi antibodies by use of radioimmunoprecipitation assay; the blood donors were seronegative. Six other patients had received a liver or kidney from these organ donors. None showed evidence of T. cruzi infection. CONCLUSIONS: To our knowledge, this is the first report of T. cruzi transmission associated with heart transplantation. Clinicians and public health authorities should be aware that manifestations of Chagas disease can occur after transplantation, requiring rapid evaluation, diagnosis, and treatment.

Diagnosing schistosomiasis by detection of cell-free parasite DNA in human plasma.

INTRODUCTION: Schistosomiasis (bilharzia), one of the most relevant parasitoses of humans, is confirmed by microscopic detection of eggs in stool, urine, or organ biopsies. The sensitivity of these procedures is variable due to fluctuation of egg shedding. Serological tests on the other hand do not distinguish between active and past disease. In patients with acute disease (Katayama syndrome), both serology and direct detection may produce false negative results. To overcome these obstacles, we developed a novel diagnostic strategy, following the rationale that Schistosoma DNA may be liberated as a result of parasite turnover and reach the blood. Cell-free parasite DNA (CFPD) was detected in plasma by PCR. METHODOLOGY/PRINCIPAL FINDINGS: Real-time PCR with internal control was developed and optimized for detection of CFPD in human plasma. Distribution was studied in a mouse model for Schistosoma replication and elimination, as well as in human patients seen before and after treatment. CFPD was detectable in mouse plasma, and its concentration correlated with the course of anti-Schistosoma treatment. Humans with chronic disease and eggs in stool or urine (n = 14) showed a 100% rate of CFPD detection. CFPD was also detected in all (n = 8) patients with Katayama syndrome. Patients in whom no viable eggs could be detected and who had been treated for schistomiasis in the past (n = 30) showed lower detection rates (33.3%) and significantly lower CFPD concentrations. The duration from treatment to total elimination of CFPD from plasma was projected to exceed one year. CONCLUSIONS/SIGNIFICANCE: PCR for detection of CFPD in human plasma may provide a new laboratory tool for diagnosing schistosomiasis in all phases of clinical disease, including the capacity to rule out Katayama syndrome and active disease. Further studies are needed to confirm the clinical usefulness of CFPD quantification in therapy monitoring.

Modulatory effects of hard clam (Mercenaria mercenaria) tissue extracts on the in vitro growth of its pathogen QPX.

Quahog parasite unknown (QPX) is a fatal protistan parasite affecting cultured and wild hard clams Mercenaria mercenaria along the northeastern coasts of the USA and maritime Canada. Field investigations and laboratory transmission studies revealed some variations in the susceptibility of different hard clam stocks to QPX infection. In this study, we used in vitro QPX cultures to investigate the effect of plasma and tissue extracts from two different clam stocks on parasite survival and growth. Results demonstrated the presence of factors in clams that significantly modulate QPX growth. Extracts from gills and mantle tissues as well as plasma inhibited in vitro QPX growth, whereas foot and adductor muscle extracts enhanced parasite growth. Investigations of anti-QPX activities in plasma from two clam stocks displaying different susceptibility toward QPX disease in vivo demonstrated higher inhibition of QPX growth by plasma from New York (resistant) clams compared to Florida (susceptible) clams. Some clams appeared to be deficient in inhibitory factors, suggesting that such animals may become more easily infected by the parasite.

Proteomic analysis of parasitized Plutella xylostella larvae plasma.

Insects use their innate immunity to defend themselves against foreign invaders, such as microorganisms, nematodes and parasites. Cotesia plutellae, an endoparasitoid wasp that parasitizes the diamondback moth Plutella xylostella, uses several strategies to attack the host immune system, such as injection of viruses, venom, and serosal membrane-derived cells denoted teratocytes. However, the proteome profiles related to these immune deficiency systems have yet to be clearly defined. In this study, we investigate differences in protein expression patterns in parasitized P. xylostella larvae, with a view to identifying parasitism-specific factors. Using 2D polyacrylamide gel electrophoresis, proteins in the host plasma were assessed every 48 h after parasitism by C. plutellae. A large number of protein spots (350 in total) were detected, and approximately 50 spots were differentially expressed in the parasitized P. xylostella larvae every 48 h. In total, 26 potential candidates, including P. xylostella Serpin 2 (pxSerpin 2), translationally controlled tumor protein, signal transduction histidine kinase, apolipophorin-III, and fatty-acid binding protein were identified through quadrupole time-of-flight tandem mass spectrometry and sequence homology analysis. These proteins were classified into the following functional groups: immunity, signaling, lipid metabolism, energy metabolism, amino acid/nucleotide metabolism, and others. The pxSerpin 2 gene was cloned, and its expression profile investigated during the course of parasitism. Real-time PCR analysis of pxSerpin 2 revealed a poor correlation between the mRNA level and protein abundance. Our results clearly suggest that parasitism-specific proteins participate in suppression of the host immune response.

Photochemical inactivation with amotosalen and long-wavelength ultraviolet light of Plasmodium and Babesia in platelet and plasma components.

BACKGROUND: Transfusion-transmitted cases of malaria and babesiosis have been well documented. Current efforts to screen out contaminated blood products result in component wastage due to the lack of specific detection methods while donor deferral does not always guarantee safe blood products. This study evaluated the efficacy of a photochemical treatment (PCT) method with amotosalen and long-wavelength ultraviolet light (UVA) to inactivate these agents in red blood cells (RBCs) contaminating platelet (PLT) and plasma components. STUDY DESIGN AND METHODS: Plasmodium falciparum- and Babesia microti-contaminated RBCs seeded into PLT and plasma components were treated with 150 micromol per L amotosalen and 3 J per cm2 UVA. The viability of both pathogens before and after treatment was measured with infectivity assays. Treatment with 150 micromol per L amotosalen and 1 J per cm2 UVA was used to assess the robustness of the PCT system. RESULTS: No viable B. microti was detected in PLTs or plasma after treatment with 150 mol per L amotosalen and 3 J per cm2 UVA, demonstrating a mean inactivation of greater than 5.3 log in PLTs and greater than 5.3 log in plasma. After the same treatment, viable P. falciparum was either absent or below the limit of quantification in three of four replicate experiments both in PLTs and in plasma demonstrating a mean inactivation of at least 6.0 log in PLTs and at least 6.9 log in plasma. Reducing UVA dose to 1 J per cm2 did not significantly affect the level of inactivation. CONCLUSION: P. falciparum and B. microti were highly sensitive to inactivation by PCT. Pathogen inactivation approaches could reduce the risk of transfusion-transmitted parasitic infections and avoid unnecessary donor exclusions.

Pathogen inactivation of Trypanosoma cruzi in plasma and platelet concentrates using riboflavin and ultraviolet light.

BACKGROUND: Emigration of people infected with Trypanosoma cruzi to non-endemic areas has resulted in transfusion transmission to immunocompromised recipients. We studied the feasibility of inactivating T. cruzi using a new technology which utilizes riboflavin as a photosensitizer in combination with UV light, Mirasol PRT. METHODS: One billion T. cruzi organisms and 30 mL of 500 microM riboflavin were added to each of six units of human plasma and six units of platelets. To determine the level of detection of organism, a sample of each unit was cultured in tenfold serial dilutions beginning with 100 billion/250 mL as the starting culture. After 30 min, each unit was illuminated with 5.9 J/cm(2) of UV light (6.24 J/mL). The units were then cultured again in tenfold serial dilutions post-treatment. RESULTS: A 6 log reduction of pathogens was demonstrated in 5 of 6 units of plasma, and a 7 log reduction of pathogens was demonstrated in one unit. A 6 log reduction of pathogens was demonstrated in 3 of 6 units of platelets, a 7 log reduction was demonstrated in 2 of 6 units of platelets, and an 8 log reduction of pathogens was demonstrated in 1 of 6 units. CONCLUSIONS: Mirasol PRT treatment demonstrated an ability to inactivate 5-7 logs of T. cruzi in plasma and platelets.

The efficacy of photochemical treatment with methylene blue and light for the reduction of Trypanosoma cruzi in infected plasma.

BACKGROUND AND OBJECTIVES: Chagas disease is a transfusion-transmitted infection. This study evaluates the efficacy of a methylene blue (MB) and light system for reducing the viability of Trypanosoma cruzi in plasma. MATERIALS AND METHODS Trypanosoma cruzi strains were spiked in plasma pools. Treatment arms included combined filtration, MB, light and freezing. Post-treatment parasite viability was assayed through in vitro cultures and in vivo inoculation in inducible nitric oxide synthase- and interferon-gamma-receptor-deficient mice. RESULTS: The filtration, MB and light combined treatment showed a log reduction of > 3.4 in in vitro cultures, and log reductions that ranged from > 4.9 to > 5.8 in deficient mice inoculated with different T. cruzi strains. CONCLUSION: The treatment of plasma units with the MB and light system reduces the T. cruzi burden and could be useful in preventing transfusion-transmitted Chagas disease.