Ultraviolet Inactivation of Chikungunya Virus.

OBJECTIVE: Chikungunya virus (CHIKV) is a rapidly emerging arbovirus causing millions of infections in more than 40 countries. CHIKV is typically a biosafety level 3 pathogen in many countries and handling of CHIKV requires a high standard of laboratory safety settings. Many studies require the whole virus to be handled in a biosafety level 2 setting. A potential solution for managing this problem is pathogen inactivation without affecting its antigenicity. In the present study, we attempted to inactivate CHIKV by ultraviolet (UV) irradiation. METHODS: Different UV doses were used to inactivate CHIKV. The replication status of the inactivated virus was verified in cell lines. Western blot, electron microscopy, and immune fluorescence assay were used, respectively, to view the antigenicity, structural integrity, and entry of the virus into cell lines. RESULTS: The inactivation was complete when a UV dose of 0.09 J/cm2 for 3 × 30 s was used and no change in antigenicity and integrity was observed. CONCLUSIONS: The study concludes that the UV-inactivated virus is antigenically stable and could be used in biosafety level 2 settings for different experiments.

Co-protoporphyrin IX and Sn-protoporphyrin IX inactivate Zika, Chikungunya and other arboviruses by targeting the viral envelope.

The global situation of diseases transmitted by arthropod-borne viruses such as Dengue (DENV), Yellow Fever (YFV), Chikungunya (CHIKV) and Zika (ZIKV) viruses is alarming and treatment of human infection by these arboviruses faces several challenges. The discovery of broad-spectrum antiviral molecules, able to inactivate different groups of viruses, is an interesting approach. The viral envelope is a common structure among arboviruses, being a potential target for antivirals. Porphyrins are amphipathic molecules able to interact with membranes and absorb light, being widely used in photodynamic therapy. Previously, we showed that heme, Co-protoporphyrin IX (CoPPIX) and Sn-protoporphyrin IX (SnPPIX) directly inactivate DENV and YFV infectious particles. Here we demonstrate that the antiviral activity of these porphyrins can be broadened to CHIKV, ZIKV, Mayaro virus, Sindbis virus and Vesicular Stomatitis virus. Porphyrin treatment causes viral envelope protein loss, affecting viral morphology, adsorption and entry into target cells. Also, light-stimulation enhanced the SnPPIX activity against all tested arboviruses. In summary, CoPPIX and SnPPIX were shown to be efficient broad-spectrum compounds to inactivate medically and veterinary important viruses.

Deinococcus Mn2+-peptide complex: A novel approach to alphavirus vaccine development.

Over the last ten years, Chikungunya virus (CHIKV), an Old World alphavirus has caused numerous outbreaks in Asian and European countries and the Americas, making it an emerging pathogen of great global health importance. Venezuelan equine encephalitis virus (VEEV), a New World alphavirus, on the other hand, has been developed as a bioweapon in the past due to its ease of preparation, aerosol dispersion and high lethality in aerosolized form. Currently, there are no FDA approved vaccines against these viruses. In this study, we used a novel approach to develop inactivated vaccines for VEEV and CHIKV by applying gamma-radiation together with a synthetic Mn-decapeptide-phosphate complex (MnDpPi), based on manganous-peptide-orthophosphate antioxidants accumulated in the extremely radiation-resistant bacterium Deinococcus radiodurans. Classical gamma-irradiated vaccine development approaches are limited by immunogenicity-loss due to oxidative damage to the surface proteins at the high doses of radiation required for complete virus-inactivation. However, addition of MnDpPi during irradiation process selectively protects proteins, but not the nucleic acids, from the radiation-induced oxidative damage, as required for safe and efficacious vaccine development. Previously, this approach was used to develop a bacterial vaccine. In the present study, we show that this approach can successfully be applied to protecting mice against viral infections. Irradiation of VEEV and CHIKV in the presence of MnDpPi resulted in substantial epitope preservation even at supra-lethal doses of gamma-rays (50,000Gy). Irradiated viruses were found to be completely inactivated and safe in vivo (neonatal mice). Upon immunization, VEEV inactivated in the presence of MnDpPi resulted in drastically improved protective efficacy. Thus, the MnDpPi-based gamma-inactivation approach described here can readily be applied to developing vaccines against any pathogen of interest in a fast and cost-effective manner.

Use of hydrogen peroxide vapour & plasma irradiation in combination for quick decontamination of closed chambers.

BACKGROUND & OBJECTIVES: Various conventional methods such as gaseous, vapour and misting systems, fogging, manual spray and wipe techniques employing a number of chemical agents are used for decontamination of enclosed spaces. Among all these methods, use of aerosolized formaldehyde is the most preferred method due to cost-effectiveness and practical aspects. However, being extremely corrosive in nature generating very irritating fumes and difficulty in maintaining a high level of gas concentration, many laboratories prefer the vaporization of hydrogen peroxide (H 2 O 2 ) as an alternative. We present here the results of using H 2 O 2 vapour in combination with plasma irradiation for quick decontamination of closed chambers. METHODS: The present study describes a decontamination method, using plasma irradiation in combination with H 2 O 2 (5%). Effect of plasma irradiation and H 2 O 2 on the viability of bacterial spores (Bacillus subtilis), Chikungunya and Kyasanur Forest Disease viruses was assessed. RESULTS: Data suggest that with the combination of H 2 O 2 vapour and plasma irradiation, within short time (three minutes), decontamination of surfaces and space volume could be achieved. Although it showed damage of spores present on the strips, it did not show any penetration power. INTERPRETATION & CONCLUSIONS: The results were encouraging, and this method was found to be efficient for achieving surface sterilization in a short time. This application may be useful in laboratories and industries particularly, those working on clean facility concept following good laboratory and manufacturing practices.

Dengue and chikungunya viruses in plasma are effectively inactivated after treatment with methylene blue and visible light.

BACKGROUND: Arboviruses, such as dengue viruses (DENV) and chikungunya virus (CHIKV), pose a risk to the safe transfusion of blood components, including plasma. Pathogen inactivation is an approach to manage this transfusion transmission risk, with a number of techniques being used worldwide for the treatment of plasma. In this study, the efficacy of the THERAFLEX MB-Plasma system to inactivate all DENV serotypes (DENV-1, DENV-2, DENV-3, DENV-4) or CHIKV in plasma, using methylene blue and light illumination at 630 nm, was investigated. STUDY DESIGN AND METHODS: Pooled plasma units were spiked with DENV-1, DENV-2, DENV-3 DENV-4, or CHIKV and treated with the THERAFLEX MB-Plasma system at four light illumination doses: 20, 40, 60, and 120 (standard dose) J/cm(2) . Pre- and posttreatment samples were collected and viral infectivity was determined. The reduction in viral infectivity was calculated for each dose. RESULTS: Treatment of plasma with the THERAFLEX MB-Plasma system resulted in at least a 4.46-log reduction in all DENV serotypes and CHIKV infectious virus. The residual infectivity for each was at the detection limit of the assay used at 60 J/cm(2) , with dose dependency also observed. CONCLUSIONS: Our study demonstrated the THERAFLEX MB-Plasma system can reduce the infectivity of all DENV serotypes and CHIKV spiked into plasma to the detection limit of the assay used at half of the standard illumination dose. This suggests this system has the capacity to be an effective option for managing the risk of DENV or CHIKV transfusion transmission in plasma.

Inactivation of dengue, chikungunya, and Ross River viruses in platelet concentrates after treatment with ultraviolet C light.

BACKGROUND: Arboviruses, including dengue (DENV 1-4), chikungunya (CHIKV), and Ross River (RRV), are emerging viruses that are a risk for transfusion safety globally. An approach for managing this risk is pathogen inactivation, such as the THERAFLEX UV-Platelets system. We investigated the ability of this system to inactivate the above mentioned arboviruses. STUDY DESIGN AND METHODS: DENV 1-4, CHIKV, or RRV were spiked into buffy coat (BC)-derived platelet (PLT) concentrates in additive solution and treated with the THERAFLEX UV-Platelets system at the following doses: 0.05, 0.1, 0.15, and 0.2 J/cm(2) (standard dose). Pre- and posttreatment samples were taken for each dose, and the level of viral infectivity was determined. RESULTS: At the standard ultraviolet C (UVC) dose (0.2 J/cm(2) ), viral inactivation of at least 4.43, 6.34, and 5.13 log or more, was observed for DENV 1-4, CHIKV, and RRV, respectively. A dose dependency in viral inactivation was observed with increasing UVC doses. CONCLUSIONS: Our study has shown that DENV, CHIKV, and RRV, spiked into BC-derived PLT concentrates, were inactivated by the THERAFLEX UV-Platelets system to the limit of detection of our assay, suggesting that this system could contribute to the safety of PLT concentrates with respect to these emerging arboviruses.

Photochemical inactivation of chikungunya virus in human apheresis platelet components by amotosalen and UVA light.

Chikungunya virus (CHIKV) is a mosquito-borne alphavirus that recently re-emerged in Africa and rapidly spread into countries of the Indian Ocean basin and South-East Asia. The mean viremic blood donation risk for CHIKV on La Réunion reached 1.5% at the height of the 2005-2006 outbreaks, highlighting the need for development of safety measures to prevent transfusion-transmitted infections. We describe successful inactivation of CHIKV in human platelets and plasma using photochemical treatment with amotosalen and long wavelength UVA illumination. Platelet components in additive solution and plasma units were inoculated with two different strains of high titer CHIKV stock (6.0-8.0 logs/mL), and then treated with amotosalen and exposure to 1.0-3.0 J/cm² UVA. Based on in vitro assays of infectious virus pre- and post-treatment to identify endpoint dilutions where virus was not detectable, mean viral titers could effectively be reduced by > 6.4 ± 0.6 log10 TCID50/mL in platelets and ≥ 7.6 ± 1.4 logs in plasma, indicating this treatment has the capacity to prevent CHIKV transmission in human blood components collected from infected donors in or traveling from areas of CHIKV transmission.

Photochemical inactivation of chikungunya virus in plasma and platelets using the Mirasol pathogen reduction technology system.

BACKGROUND: Chikungunya virus (CHIKV) is a reemerging mosquito-borne virus that has been responsible for a number of large-scale epidemics as well as imported cases covering a wide geographical range. As a blood-borne virus capable of mounting a high-titer viremia in infected humans, CHIKV was included on a list of risk agents for transfusion and organ transplant by the AABB Transfusion-Transmitted Diseases Committee. Therefore, we evaluated the ability of the Mirasol pathogen reduction technology (PRT) system (CaridianBCT Biotechnologies) to inactivate live virus in contaminated plasma and platelet (PLT) samples. STUDY DESIGN AND METHODS: Plasma, PLTs, and phosphate-buffered saline controls were spiked with CHIKV and treated with riboflavin and varying doses of ultraviolet (UV) light using the Mirasol PRT system. Samples were tested before and after treatment for cytotoxicity, interference, and virus titer by titration and quantitative real-time reverse transcription-polymerase chain reaction. RESULTS: A significant reduction in CHIKV titer of greater than 99% was recorded after treatment of plasma or PLTs with the Mirasol PRT system, and the titer reduction was directly proportional to the UV dose delivered to the samples. No cytotoxicity of interference was observed in any sample at any treatment dose. CONCLUSION: These data indicate that the Mirasol PRT system efficiently inactivated live CHIKV in plasma and PLTs and could therefore potentially be used to prevent CHIKV transmission through the blood supply.

Transfusion of platelet components prepared with photochemical pathogen inactivation treatment during a Chikungunya virus epidemic in Ile de La Réunion.

BACKGROUND: During the Chikungunya virus (CHIKV) epidemic on Ile de La Réunion, France, more than 30% of 750,000 inhabitants were infected. Local blood donation was suspended to prevent transfusion-transmitted infection (TT-CHIKV). To sustain the availability of platelet (PLT) components, the Etablissement Français du Sang implemented universal pathogen inactivation (INTERCEPT, Cerus Europe BV) of PLT components (CPAs). The study assessed the safety of PLT components treated with pathogen inactivation transfused in routine clinical practice. STUDY DESIGN AND METHODS: This was a retrospective observational study using patient medical records and the AFSSAPS hemovigilance database (eFIT) to identify TT-CHIKV and adverse events (AEs) classified as acute transfusion reactions (ATRs) to PLT components prepared with pathogen inactivation. RESULTS: During 1 year, 1950 INTERCEPT-CPAs were transfused to 335 adult, 51 pediatric, and 41 infant patients. Nineteen AEs were observed in 15 patients and 10 were classified as ATRs. Eight ATRs occurred in 6 pediatric hematology-oncology patients. No ATRs were observed in infants. The most frequently reported signs and symptoms were Grade 1 urticaria, itching, chills, fever, and anxiety. No cases of transfusion-related acute lung injury, TT-sepsis, or TT-CHIKV were detected. CONCLUSIONS: INTERCEPT-CPAs were well tolerated in a broad range of patients, including infants. ATR incidence was low and when present ATRs were of mild severity.

Immunogenicity of purified, inactivated chikungunya virus in monkeys.

Chikungunya (CHIK) virus, harvested from infected BHK-21 cell cultures and highly purified by a method combining zinc acetate precipitation, Sephadex-Sepharose column chromatography, and sucrose density gradient centrifugation, was subjected to ultraviolet (UV) irradiation and treatment with formalin. Inactivation of the virus by UV light was apparently a first-order reaction. The virus treated with 0.05% formalin at 4 degrees C was inactivated completely after 7 weeks, but when treated with 0.025% formalin retained its plaque infectivity at least 17 weeks. Both UV- and formalin-inactivated viruses induced production of anti-CHIK neutralizing antibodies in Japanese monkeys with no preimmune antibodies. Judging by the titres of antibodies produced, the immunogenic effect of the UV-irradiated virus was superior to that of the formalin-treated virus.