Sofosbuvir Add-on to Ribavirin Treatment for Chronic Hepatitis E Virus Infection in Solid Organ Transplant Recipients Does Not Result in Sustained Virological Response.

Ribavirin is effective for treating immunocompromised patients with chronic hepatitis E virus infection. However, ribavirin treatment is not always successful. We describe 3 solid organ transplant recipients treated with sofosbuvir and ribavirin after failing ribavirin monotherapy. Complete elimination of hepatitis E virus could not be achieved.

Syndromic sample-to-result PCR testing for respiratory infections in adult patients.

BACKGROUND: Syndromic sample-to-result (SS2R) poly merase chain reaction (PCR) can rapidly identify causative pathogens of respiratory tract infections (RTI). We evaluated diagnostic accuracy and applicability of one of the current SS2R diagnostics, the FilmArray® Respiratory Viral Panel. METHODS: We performed a prospective study among adults presenting with symptoms of RTI at the Emergency Department of the University Medical Centre Utrecht (the Netherlands) during the 2016-2017 viral respiratory season. Clinical data were collected. We compared SS2R results on nasopharyngeal swabs to conventional real time PCR, calculated turnaround times (TAT) and explored implementation barriers using questionnaires. RESULTS: 62 Patients were included (64.5 yr [interquartile range (IQR) 44.3-75.0]). SS2R sensitivity was 82.9% [95% confidence interval (CI) 67.9-92.9] and specificity was 95.2% [95% CI 76.2-99.9] for detection of all present viruses (n = 60). Kappa agreement (0.73 [95% CI 0.56-0.90]) was good (p = 0.000). Median SS2R TAT was 2:06 hours [IQR 1:45-3:17] compared to 32:00 hours [IQR 26:50-40:42] of conventional PCR (n = 49, p = 0.000). Ease-of-use and fast TAT were unanimously reported as benefits, and low test capacity with a single SS2R system as drawback. CONCLUSION: SS2R testing for respiratory viruses offers a rapid and reliable diagnostic method which has great potential for more efficient and targeted management in adult patients with RTI.

The added value of hepatitis E diagnostics in determining causes of hepatitis in routine diagnostic settings in the Netherlands.

OBJECTIVES: Hepatitis E virus (HEV) genotype 3 is endemic in Europe and an underdiagnosed and emerging (public) health issue. In recent years commercial enzyme immunoassays (EIAs) that detect antibodies to HEV more adequately, became available. We investigated the added value of this HEV serology in the diagnostic work flow to detect viral causes of recent hepatitis. METHODS: During a 2-year period (May 2013 to May 2015), HEV serology was added to the hepatitis work flow, consisting of serological detection of hepatitis viruses A, B and C (HAV, HBV, HCV), Epstein-Barr virus (EBV) and cytomegalovirus (CMV). Samples positive for HEV IgM were also analysed using PCR to detect HEV RNA. If positive, HEV sequencing was performed for genotyping purposes. RESULTS: In 235 out of 2521 patients (9.3%), a viral cause for hepatitis was found. Recent HAV, HBV, HCV, EBV or CMV infections were serologically diagnosed in 3, 34, 10, 69 and 42 patients, respectively. Seventy-eight patients (3.1%) had a recent HEV infection. In 49 of them, sufficient HEV RNA was present for genotyping. All patients were infected with HEV genotype 3. CONCLUSIONS: In our region, an HEV infection is the most frequently diagnosed viral cause for recent hepatitis. These results indicate that, in a country where HEV is endemic, serological HEV diagnostics should be added to the standard work-up for viral hepatitis.

Influenza vaccination in healthcare workers; comparison of side effects and preferred route of administration of intradermal versus intramuscular administration.

OBJECTIVE: To explore the nature and severity of side effects and future preference of intradermal versus intramuscular influenza vaccination in healthcare workers. DESIGN: Prospective cohort study. SETTING: Two University Medical Centers in The Netherlands. PARTICIPANTS: Healthcare workers receiving an influenza vaccination. METHODS: Healthcare workers that were vaccinated during the influenza vaccination season of 2012-2013 were approached for participation in a questionnaire study. The questionnaire was divided into two parts. The first part had to be answered directly after vaccination and the second part two weeks after vaccination. The motivation for vaccine uptake, whether or not the HCWs had direct contact with patients and the prevalence and severity of local and systemic side effects of influenza vaccination were explored. In addition, it was assessed how participants experienced the vaccination and which type of administration they preferred for future vaccination. RESULTS: Side effects of vaccination were more prevalent in the intradermal group versus the intramuscular group (56% versus 26%, p<0.001). Local side effects were perceived as more severe in healthcare workers receiving the intradermal vaccine. Directly after vaccination, healthcare workers preferred the intradermal vaccination. Two weeks after vaccination both types of vaccine were equally appreciated. CONCLUSIONS: This study shows that there are significant differences in the nature and severity of side effects upon intramuscular and intradermal influenza vaccination. This difference did not result in a preference among the vaccinated subjects for one type of vaccine.

Human herpesvirus-6 DNAemia is a sign of impending primary CMV infection in CMV sero-discordant renal transplantations.

BACKGROUND: Human herpesvirus-6 (HHV-6) frequently reactivates in immunocompromized individuals. Most commonly HHV-6 DNA is detected without organ localized disease. This HHV-6 DNAemia usually occurs in patients who also have CMV reactivations. The question if reactivation of one virus causes reactivation of the other, or whether both viruses reactivate independently, cannot be answered in populations with high seroprevalence for both viruses. Our study is the first in which 35 patients have been included who were CMV seronegative prior to transplantation. OBJECTIVE: We investigated the occurrence of HHV-6 reactivations in relation to the CMV-serostatus of renal transplantation donor and recipient. STUDY DESIGN: 9 consecutive patients receiving a renal transplantation were included. All available stored whole blood samples were tested for HHV-6 DNA by quantitative PCR. Details including CMV serostatus of donor and recipient were recorded. RESULTS: CMV-seropositive recipients have a 68% chance of developing HHV-6 DNAemia if the kidney came from a CMV-seropositive donor, compared to 26% if the kidney came from a CMV-seronegative donor. CMV-seronegative recipients, who are bound to undergo primary CMV infections following transplantation with a renal graft from a CMV-seropositive donor, have 88% chance of developing HHV-6 DNAemia. If they receive a graft from a CMV-seronegative donor the chance of developing HHV-6 DNAemia is 22%. CONCLUSION: Receiving a renal transplant from a CMV-seropositive donor increases the chance of developing HHV-6 DNAemia. HHV-6 DNAemia is a sign of impending primary CMV infections following sero-discordant renal transplantations.

Silent hepatitis E virus infection in Dutch blood donors, 2011 to 2012.

In Europe, the dynamics of endemic hepatitis E virus (HEV) infection remain enigmatic. We studied the presence of silent HEV infection among Dutch blood donors. Using donations collected throughout the Netherlands in 2011 and 2012, 40,176 donations were tested for HEV RNA in 459 pools of 48 or 480 donations. Deconstruction of the reactive pools identified 13 viraemic donors. In addition, 5,239 donors were tested for presence of anti-HEV IgG and IgM and for HEV RNA when IgM-positive. Of the 5,239 donations, 1,401 (27%) tested repeat-positive for HEV IgG, of which 49 (3.5%) also tested positive for anti-HEV IgM. Four of the HEV IgM-positive donors tested positive for HEV RNA. HEV IgG seroprevalence ranged from 13% among donors younger than 30 years to 43% in donors older than 60 years. The finding of 17 HEV RNA-positive donations among 45,415 donations corresponds to one HEV-positive blood donation per day in the Netherlands. For 16 of the 17 HEV RNA-positive donors, genotyping succeeded, revealing HEV genotype 3, which is circulating among Dutch pigs. Apparently, silent HEV infection is common in the Netherlands, which possibly applies to larger parts of Europe.

The significance of rhinovirus detection in hospitalized children: clinical, epidemiological and virological features.

Recent developments in molecular diagnostic tools have led to the easy and rapid detection of a large number of rhinovirus (HRV) strains. However, the lack of clinical and epidemiological data hampers the interpretation of these diagnostic findings. From October 2009 to January 2011, we conducted a prospective study in hospitalized children from whom samples were taken for the detection of respiratory viruses. Clinical, epidemiological and microbiological data from 644 patients with 904 disease episodes were collected. When HRV tested positive, strains were further characterized by sequencing the VP4/VP2 region of the HRV genome. HRV was the single respiratory virus detected in 254 disease episodes (28%). Overall, 99 different serotypes were detected (47% HRV-A, 12% HRV-B, 39% HRV-C). Patients with HRV had more underlying pulmonary illness compared with patients with no virus (p 0.01), or patients with another respiratory virus besides HRV (p 0.007). Furthermore, cough, shortness of breath and a need for oxygen were significantly more present in patients with HRV infection. Particularly, patients with HRV-B required extra oxygen. No respiratory symptom, except for oxygen need, was predictive of the presence of HRV. In 22% of HRV-positive disease episodes, HRV infection was hospital acquired. Phylogenetic analysis revealed several clusters of HRV; in more than 25% of these clusters epidemiological information was suggestive of transmission within specific wards. In conclusion, the detection of HRV may help in explaining respiratory illness, particular in patients with pulmonary co-morbidities. Identifying HRV provides opportunities for timely implementation of infection control measures to prevent intra-hospital transmission.

Influenza in the immediate post-pandemic era: a comparison with seasonal and pandemic influenza in hospitalized patients.

BACKGROUND: Comparative data on severity and treatment of seasonal, pandemic and post-pandemic influenza virus infections are scarce. OBJECTIVES: To systematically analyze characteristics of hospitalized patients with influenza in the post-pandemic period compared to seasonal and pandemic influenza. STUDY DESIGN: Clinical and virological data of patients hospitalized in a tertiary referral hospital with post-pandemic influenza (2010-2011) were compared with those during seasonal influenza epidemics (2007-2009) and the influenza A(H1N1)pdm09 pandemic (2009-2010). RESULTS: 82 patients were admitted during the post-pandemic period, compared to 85 during the pandemic and 60 during seasonal influenza epidemics. No differences were observed in the occurrence of complicated illness and the need for intensive care. However, radiographic pneumonia was significantly more often diagnosed in patients with influenza A(H1N1)pdm09 compared to patients with seasonal influenza A (25% versus 71% in pandemic, p=0.004, and 55% in post-pandemic, p=0.047). Oseltamivir was more frequently prescribed in post-pandemic and pandemic patients compared to previous influenza seasons (48.9% resp. 76.5% versus 6.5%, p<0.0001). During the post-pandemic period, patients with influenza B were significantly less often treated with oseltamivir compared to patients with influenza A (27.0% versus 48.9%, p=0.043), although the course of illness in patients with influenza B was comparable with influenza A. No upsurge of oseltamivir resistance was observed. CONCLUSIONS: In our center, severity of illness was comparable for all influenza seasons, although more radiographic pneumonia was diagnosed in patients with influenza A(H1N1)pdm09. Despite the increased use of oseltamivir, no increase in oseltamivir resistance was detected.

Evaluation of the antiviral response to zanamivir administered intravenously for treatment of critically ill patients with pandemic influenza A (H1N1) infection.

A retrospective nationwide study on the use of intravenous (IV) zanamivir in patients receiving intensive care who were pretreated with oseltamivir in the Netherlands was performed. In 6 of 13 patients with a sustained reduction of the viral load, the median time to start IV zanamivir was 9 days (range, 4-11 days) compared with 14 days (range, 6-21 days) in 7 patients without viral load reduction (P = .052). Viral load response did not influence mortality. We conclude that IV zanamivir as late add-on therapy has limited effectiveness. The effect of an immediate start with IV zanamivir monotherapy or in combination with other drugs need to be evaluated.

A comparative study on the immunotherapeutic efficacy of recombinant Semliki Forest virus and adenovirus vector systems in a murine model for cervical cancer.

Currently, various therapeutic strategies are being explored as a potential means to immunize against metastatic malignant cells or even primary tumours. Using recombinant viral vectors systems or protein-based immunization approaches, we are developing immunotherapeutic strategies against cervical cancer or premalignant cervical disease, as induced by high-risk type human papillomaviruses (HPVs). We previously demonstrated that immunization of mice with recombinant replication-defective Semliki Forest virus (rSFV) encoding a fusion protein of HPV16 E6 and -E7 (SFV-eE6,7) induces strong cytotoxic T-lymphocyte (CTL) activity and eradication of established HPV-transformed tumours. In this study, we compared the antitumour efficacy of SFV-eE6,7 with that of a recombinant adenovirus (rAd) type 5 vector, expressing the same antigen construct (Ad-eE6,7). Prime-boosting with SFV-eE6,7 resulted in higher precursor CTL frequencies and CTL activity compared to prime-boosting with Ad-eE6,7 and also in murine tumour treatment experiments SFV-eE6,7 was more effective than Ad-eE6,7. To elicit a therapeutic effect with Ad-eE6,7, 100/1000-fold higher doses were needed compared to SFV-eE6,7. In vivo T-cell depletion experiments demonstrated that these differences could not be explained by the induction of a different type of effector cells, since CD8+ T cells were the main effector cells involved in the protection against tumour growth in both rSFV- and rAd-immunized mice. Also comparable amounts of in vivo transgene expression were found upon immunization with rSFV and rAd encoding the reportor gene luciferase. However, anti-vector responses induced by a single injection with rAd resulted in a more than 3-log decrease in luciferase expression after a second injection of rAd. With rSFV, transgene expression was inhibited by only one to two orders of magnitude in preinjected mice. As an antigen-specific booster immunization strongly increases the level of the CTL response and is essential for efficient induction of immunological memory, it is likely that (part of) the difference in efficacy between rSFV and rAd type 5 can be ascribed to a diminished efficacy of the booster immunization in the case of rAd due to anti-vector antibody responses.

Induction of human papilloma virus E6/E7-specific cytotoxic T-lymphocyte activity in immune-tolerant, E6/E7-transgenic mice.

Despite promising preclinical results of various therapeutic anticancer immunization strategies, these approaches may not be effective enough to eradicate tumors in cancer patients. While most animal models are based on fast-growing transplantable tumors, malignancies in, for example, cervical cancer patients in general develop much more slowly, which may lead to immune suppression and/or immune tolerance. As a consequence, the immunomodulating signal of any therapeutic immunization regimen should be sufficiently potent to overcome this immunocompromised condition. In previous studies, we demonstrated that an experimental vaccine against human papillomavirus (HPV)-induced cervical cancer, based on Semliki Forest virus (SFV), induces robust HPV-specific cellular immune responses in mice. Now we studied whether this strategy is potent enough to also prime a cellular immune response in immune-tolerant HPV transgenic mice, in which CTL activity cannot be induced using protein or DNA vaccines. We demonstrate that, depending on the route of immunization, SFV-expressing HPV16 E6 and E7 indeed has the capacity to induce HPV16 E7-specific cytotoxic T cells in HPV-transgenic mice.