BACKGROUND AND AIMS:Hepatitis delta virus (HDV) infection is associated with fast progression to liver cirrhosis and liver complications. Previous studies have, however, been mainly from tertiary care centers, with risk for referral bias toward patients with worse outcomes. Furthermore, the impact of HDV viremia per se on liver-related outcomes is not really known outside the human immunodeficiency virus co-infection setting. We have therefore evaluated the long-term impact of HDV viremia on liver-related outcomes in a nationwide cohort of patients with hepatitis B and D co-infection, cared for at secondary care centers in Sweden.APPROACH AND RESULTS:In total, 337 patients with anti-HDV positivity, including 233 patients with HDV RNA viremia and 91 without HDV viremia at baseline, were retrospectively studied, with a mean follow-up of 6.5 years (range, 0.5-33.1). The long-term risks for liver-related events (i.e., hepatocellular carcinoma [HCC], hepatic decompensation, or liver-related death/transplantation) were assessed, using Cox regression analysis. The risk for liver-related events and HCC was 3.8-fold and 2.6-fold higher, respectively, in patients with HDV viremia compared with those without viremia, although the latter was not statistically significant. Among patients with HDV viremia with no baseline cirrhosis, the cumulative risk of being free of liver cirrhosis or liver-related events was 81.9% and 64.0% after 5 and 10 years of follow-up, respectively. This corresponds to an incidence rate of 0.04 cases per person-year.CONCLUSIONS:HDV RNA viremia is associated with a 3.8-fold higher risk for liver-related outcomes. The prognosis was rather poor for patients with HDV viremia without cirrhosis at baseline, but it was nevertheless more benign than previous estimates from tertiary centers. Our findings may be of importance when making decisions about treatment and evaluating potential outcomes of upcoming antivirals against HDV.

Despite access to effective antiviral drugs and vaccines, hepatitis B virus (HBV) infection remains a major health issue worldwide. HBV is highly infectious and may cause chronic infection, progressive liver damage, hepatocellular cancer (HCC) and death. Early diagnosis, proper management and timing of treatment are crucial. The Swedish Reference group for Antiviral Treatment (RAV) here provides updated evidence-based guidelines for treatment and management of HBV infection which may be applicable also in other countries. Tenofovir alafenamide (TAF) has been introduced as a novel treatment option and new principles regarding indication and duration of treatment and characterization of hepatitis B have been gradually introduced which justifies an update of the previous guidelines from 2007. Updated guidelines on HCC surveillance in HBV-infected patients, treatment and prophylaxis for patients undergoing liver transplantation as well as management of pregnant women and children with HBV infection are also provided.

Background In the winter and spring of 2011 a large outbreak of cryptosporidiosis occurred in Skellefteå municipality, Sweden. This study summarizes the outbreak investigation in terms of outbreak size, duration, clinical characteristics, possible source(s) and the potential for earlier detection using calls to a health advice line. Methods The investigation included two epidemiological questionnaires and microbial analysis of samples from patients, water and other environmental sources. In addition, a retrospective study based on phone calls to a health advice line was performed by comparing patterns of phone calls between different water distribution areas. Results Our analyses showed that approximately 18,500 individuals were affected by a waterborne outbreak of cryptosporidiosis in Skellefteå in 2011. This makes it the second largest outbreak of cryptosporidiosis in Europe to date. Cryptosporidium hominis oocysts of subtype IbA10G2 were found in patient and sewage samples, but not in raw water or in drinking water, and the initial contamination source could not be determined. The outbreak went unnoticed to authorities for several months. The analysis of the calls to the health advice line provides strong indications early in the outbreak that it was linked to a particular water treatment plant. Conclusions We conclude that an earlier detection of the outbreak by linking calls to a health advice line to water distribution areas could have limited the outbreak substantially.

Daclatasvir (DCV) is a potent, pangenotypic nonstructural protein 5A inhibitor with demonstrated antiviral efficacy when combined with sofosbuvir (SOF) or simeprevir (SMV) with or without ribavirin (RBV) in patients with chronic hepatitis C virus (HCV) infection. Herein, we report efficacy and safety data for DCV‐based all‐oral antiviral therapy in liver transplantation (LT) recipients with severe recurrent HCV. DCV at 60 mg/day was administered for up to 24 weeks as part of a compassionate use protocol. The study included 97 LT recipients with a mean age of 59.3 ± 8.2 years; 93% had genotype 1 HCV and 31% had biopsy‐proven cirrhosis between the time of LT and the initiation of DCV. The mean Model for End‐Stage Liver Disease (MELD) score was 13.0 ± 6.0, and the proportion with Child‐Turcotte‐Pugh (CTP) A/B/C was 51%/31%/12%, respectively. Mean HCV RNA at DCV initiation was 14.3 × 6 log 10 IU/mL, and 37% had severe cholestatic HCV infection. Antiviral regimens were selected by the local investigator and included DCV+SOF (n = 77), DCV+SMV (n = 18), and DCV+SMV+SOF (n = 2); 35% overall received RBV. At the end of treatment (EOT) and 12 weeks after EOT, 88 (91%) and 84 (87%) patients, respectively, were HCV RNA negative or had levels <43 IU/mL. CTP and MELD scores significantly improved between DCV‐based treatment initiation and last contact. Three virological breakthroughs and 2 relapses occurred in patients treated with DCV+SMV with or without RBV. None of the 8 patient deaths (6 during and 2 after therapy) were attributed to therapy. In conclusion, DCV‐based all‐oral antiviral therapy was well tolerated and resulted in a high sustained virological response in LT recipients with severe recurrent HCV infection. Most treated patients experienced stabilization or improvement in their clinical status. Liver Transplantation 22 446‐458 2016 AASLD

In this pilot study (RibaC), 58 hepatitis C virus (HCV) genotype 1 infected treatment-naïve patients were randomized to (i) 2 weeks ribavirin double dosing concomitant with pegylated interferon-α (pegIFN-α), (ii) 4 weeks ribavirin mono-therapy prior to adding pegIFN-α, or (iii) standard-of-care (SOC) ribavirin dosing concurrent with pegIFN-α. Four weeks of ribavirin mono-therapy resulted in a mean 0.46 log10 IU/mL HCV RNA reduction differentially regulated across IL28B genotypes (0.89 vs. 0.21 log10 IU/mL for CC and CT/TT respectively; P = 0.006), increased likelihood of undetectable HCV RNA week 4 after initiating pegIFN-α and thus shortened treatment duration (P<0.05), and decreased median IP-10 concentration from 550 to 345 pg/mL (P<0.001). Both experimental strategies impacted on ribavirin concentrations, and high levels were achieved after one week of double dosing. However, by day 14, double dosing entailed a greater hemoglobin decline as compared to SOC (2.2 vs. 1.4 g/dL; P = 0.03). Conclusion: Ribavirin down-regulates IP-10, and may have an antiviral effect differently regulated across IL28B genotypes.

IntroductionNephropatia epidemica (NE), a relatively mild form of hemorrhagic fever with renal syndrome caused by the Puumala virus (PUUV), is endemic in northern Sweden. We aim to study the risk factors associated with NE in this region.MethodsWe conducted a matched case–control study between June 2011 and July 2012. We compared confirmed NE cases with randomly selected controls, matched by age, sex, and place of infection or residence. We analyzed the association between NE and several occupational, environmental, and behavioral exposures using conditional logistic regression.ResultsWe included in the final analysis 114 cases and 300 controls, forming 246 case–control pairs. Living in a house with an open space beneath, making house repairs, living less than 50 m from the forest, seeing rodents, and smoking were significantly associated with NE.ConclusionOur results could orient public health policies targeting these risk factors and subsequently reduce the NE burden in the region.

Phosphoglucomutase 3 (PGM3) is an enzyme converting N-acetyl-glucosamine-6-phosphate to N-acetyl-glucosamine-1-phosphate, a precursor important for glycosylation. Mutations in the PGM3 gene have recently been identified as the cause of novel primary immunodeficiency with a hyper-IgE like syndrome. Here we report the occurrence of a homozygous mutation in the PGM3 gene in a family with immunodeficient children, described already in 1976. DNA from two of the immunodeficient siblings was sequenced and shown to encode the same homozygous missense mutation, causing a destabilized protein with reduced enzymatic capacity. Affected individuals were highly prone to infections, but lack the developmental defects in the nervous and skeletal systems, reported in other families. Moreover, normal IgE levels were found. Thus, belonging to the expanding group of congenital glycosylation defects, PGM3 deficiency is characterized by immunodeficiency, with or without increased IgE levels, and with variable forms of developmental defects affecting other organ systems.

In 2010–2011, two large waterborne outbreaks caused by Cryptosporidium hominis affected two cities in Sweden, Östersund and Skellefteå. We investigated potential post-infection health consequences in people who had reported symptoms compatible with cryptosporidiosis during the outbreaks using questionnaires.

For the purpose of developing a national system of outbreak surveillance, we compared local outbreak signals in three sources of syndromic data: telephone triage of acute gastroenteritis, web queries about symptoms of gastrointestinal illness, and OTC pharmacy sales of anti-diarrhea medication. The sensitivity and specificity were highest for telephone triage data. It provided the most promising source of syndromic data for surveillance of point-source outbreaks. Currently, a project has been initialized to develop and implement a national system in Sweden for daily syndromic surveillance based on 1177 Health Care Direct, supporting regional and local outbreak detection and investigation.

Francisella tularensis, the causative agent of tularemia, is a potent pathogen in humans and other mammals. The ulceroglandular form of the disease is the most common expression in humans with a clinical picture characterized by a skin ulcer, enlarged regional lymph nodes and fever. Despite being a preferred route of infection, the skin also affords an effective defense barrier against F. tularensis. Doses required to induce infection by intradermal inoculation are several logs higher than those needed for infection by other routes. In the present thesis, the requirements for the local and systemic host defense to intradermal infection with F. tularensis was studied in experimental mouse models. Naive mice and mice immunized by previous infection were challenged, mostly with the live vaccine strain F. tularensis LVS but also with a clinical isolate of F. tularensis. In naive mice, intradermal inoculation of F. tularensis LVS resulted in a rapid increase of bacterial numbers during the first few days in the skin, lymph nodes, spleen and liver, followed by a decrease and eradication of the bacteria within two weeks of inoculation. Immune mice controlled the infection at the site of infection and very few bacteria spread to internal organs. When immunohistochemical staining of skin specimens was performed during the first 3 days, naive mice showed a weak or barely discernible local expression of TNF-α, IL-12 and IFN-γ. In immune mice, the expression of all three cytokines was strongly enhanced, TNF-α and IL-12 within 24 h and IFN-γ within 72 h of inoculation. To investigate the role of T cells in the defense against intradermal infection with F. tularensis LVS, naive and immune T-cell knockout mice (e.g., αβ TCR-/-, γδ TCR-/-, αβγδ TCR-/-) were used. Naive mice lacking the αβ TCR had persistently high bacterial numbers in all organs and died at 4 weeks. Mice lacking the γδ TCR, on the other hand, controlled the infection as effectively as did wild-type mice. To enable αβ TCR-/- and αβγδ TCR-/- mice to survive, antibiotic treatment was given from day 10 to 20 of infection. When intradermally challenged 2 weeks later, these animals were found to control a secondary infection, resulting in decreasing viable counts in skin and lymph nodes and prevention of spread to liver and spleen. The results indicated the presence of a T-cell independent mechanism of resistance and analyses of serum showed high levels of F. tularensis-specific IgM, findings suggesting a role for antibodies in the protection against cutaneous tularemia. To study the effect of F. tularensis-specific antibodies on host resistance, we adoptively transferred immune serum to B-cell-deficient mice. After receiving immune serum, both naive and immunized mice became capable of surviving an otherwise lethal dose of F. tularensis LVS. Moreover, transfer of immune serum to wild type mice, afforded significant protection to a lethal dose of a wild-type strain of F. tularensis subsp. holarctica, as disclosed by reduced bacterial counts in spleen and liver. Finally, we studied the effect of immune serum on the local expression of proinflammatory cytokines and neutrophils in response to an intradermal injection of F. tularensis LVS. As compared to normal serum, transfer of immune serum resulted in increased expression of TNF-α, IL-12 and neutrophils. These findings afford a possible explanation for the effect of specific antibodies in the local host protection in the skin against tularemia.

Reactive nitrogen species (RNS) and reactive oxygen species (ROS) are important mediators of the bactericidal host response. We investigated the contribution of these two mediators to the control of infection with the facultative intracellular bacterium Francisella tularensis. When intradermally infected with the live vaccine strain F. tularensis LVS, mice deficient in production of RNS (iNOS(-/-) mice) or in production of ROS by the phagocyte oxidase (p47(phox-/-) mice) showed compromised resistance to infection. The 50% lethal dose (LD50) for iNOS(-/-) mice was <20 CFU, and the LD50 for p47(phox-/-) mice was 4,400 CFU, compared to an LD50 of >500,000 CFU for wild-type mice. The iNOS(-/-) mice survived for 26.4 +/- 1.8 days, and the p47(phox-/-) mice survived for 10.1 +/- 1.3 days. During the course of infection, the serum levels of gamma interferon (IFN-gamma) and interleukin-6 were higher in iNOS(-/-) and p47(phox-/-) mice than in wild-type mice. Histological examination of livers of iNOS(-/-) mice revealed severe liver pathology. Splenocytes obtained 5 weeks after primary infection from antibiotic-treated iNOS(-/-) mice showed an in vitro recall response that was similar in magnitude and greater secretion of IFN-gamma compared to cells obtained from wild-type mice. In summary, mice lacking expression of RNS or ROS showed extreme susceptibility to infection with F. tularensis LVS. The roles of RNS and ROS seemed to be distinct since mice deficient in production of ROS showed dissemination of infection and died during the early phase of infection, whereas RNS deficiency led to severe liver pathology and a contracted course of infection.

This study demonstrates that passive transfer of Francisella tularensis-specific antibodies before experimental cutaneous infection with the live vaccine strain of F. tularensis has profound effects. Recipient mice showed stronger staining for TNF-alpha and IL12, and larger numbers of neutrophils in skin samples after infection than control mice.

T cells are crucial to the control and eradication of the facultative intracellular bacterium Francisella tularensis. A contributory role of humoral antibodies in the host defence remains to be assessed. We used B-cell-deficient mice to study the possible contribution of antibodies to the defence against the live vaccine strain (LVS) or a clinical isolate of F. tularensis, both belonging to the subspecies holarctica (type B). When B-cell-deficient (Igμ−/−) mice of the C57BL/10 background were administered immune serum one day before intradermal injection of LVS, they developed lower bacterial numbers in skin, liver, and spleen than did mice receiving normal serum, and survived a challenge inoculum that was lethal for mice given normal serum. Administration of immune serum to C57BL/10 mice afforded protection also against infection with the clinical isolate of F. tularensis subsp. holarctica. Five days after intradermal inoculation of bacteria of the isolate, animals receiving immune serum showed 4log10 lower bacterial counts in liver and spleen than mice administered normal serum. In mice primed by LVS infection, T-cell immunity and host protection were strong and only a marginal contribution of immune serum against a secondary intradermal infection was demonstrated. Together, these findings show that specific antibodies contribute to the host defence of mice against F. tularensis subsp. holarctica.

Francisella tularensis LVS is an effective live vaccine strain used for cutaneous vaccination against tularemia in man. In mice, injection of LVS causes invasive disease and subsequent development of immunity that is characterized by effective control of otherwise lethal doses of the organism. In the present investigation, it is shown that LVS-immune mice controlled an intradermal infection much more effectively than did naive mice; bacterial counts in skin samples were 1.5 to 2.0 log10 lower 24 h after injection and 6 log10 lower 72 h after injection in immune mice. Moreover, in contrast to naive mice, no bacteria were demonstrated in samples from livers and spleens of immune mice. By immunohistochemistry, skin samples from immune mice showed an intense staining for interleukin-12 (IL-12) and a moderate staining for tumor necrosis factor alpha (TNF-alpha) at 24 h postinoculation, after which staining for both cytokines faded. In naive mice, the staining for IL-12 was weak at all time points and no staining for TNF-alpha was observed. No staining for gamma interferon (IFN-gamma) was observed in any group before 72 h. At that time point, skin samples from immune mice showed moderate staining and skin samples from naive mice showed weak staining. Reverse transcriptase PCR showed an induction of mRNA of the three cytokines in the skin within the first day after injection. A quantitative analysis demonstrated higher IFN-gamma and TNF-alpha mRNA levels in immune mice at 24 h postinoculation. In conclusion, immunization with F. tularensis LVS conferred a capability to respond to cutaneous reinfection, with rapid local expression of IL-12, TNF-alpha, and IFN-gamma, and this expression was paralleled by containment and mitigation of the infection. The cytokine response may be part of a local barrier function of the skin, important to host protection against tularemia.

Tularemia is a disease caused by the facultative intracellular bacterium Francisella tularensis. Here we demonstrate that during the first weeks of infection, a significant increase in levels of Vgamma9 Vdelta2 cells occurred in peripheral blood: in 13 patients analyzed 7 to 18 days after the onset of disease, these lymphocytes represented, on average, 30.5% of CD3+ cells and nearly 100% of gammadelta+ T cells. By contrast, after vaccination with the live vaccine strain (LVS) of F. tularensis, only a minor increase occurred. Eleven days after vaccination, gammadelta T cells represented an average of 6.7% and Vgamma9 Vdelta2 cells represented an average of 5.3% of T cells, as in control subjects. Since derivatives of nonpeptidic pyrophosphorylated molecules, referred to as phosphoantigens, are powerful stimuli for Vgamma9 Vdelta2 cells, this observation prompted an investigation of phosphoantigens in F. tularensis strains. The F. tularensis phosphoantigens triggered in vitro a proliferative response of human Vgamma9 Vdelta2 peripheral blood leukocytes as well as a cytotoxic response and tumor necrosis factor release from a Vgamma9 Vdelta2 T-cell clone. Quantitatively similar phosphoantigenic activity was detected in acellular extracts from two clinical isolates (FSC171 and Schu) and from LVS. Taken together, the chemical nature of the stimulus from the clinical isolates and the significant increase in levels of Vgamma9 Vdelta2 cells in peripheral blood of tularemia patients indicate that phosphoantigens produced by virulent strains of F. tularensis trigger in vivo expansion of gammadelta T cells in tularemia.

ABSTRACTTularemia is a disease caused by the facultative intracellular bacteriumFrancisella tularensis. Here we demonstrate that during the first weeks of infection, a significant increase in levels of Vγ9Vδ2 cells occurred in peripheral blood: in 13 patients analyzed 7 to 18 days after the onset of disease, these lymphocytes represented, on average, 30.5% of CD3+cells and nearly 100% of γδ+T cells. By contrast, after vaccination with the live vaccine strain (LVS) ofF. tularensis, only a minor increase occurred. Eleven days after vaccination, γδ T cells represented an average of 6.7% and Vγ9Vδ2 cells represented an average of 5.3% of T cells, as in control subjects. Since derivatives of nonpeptidic pyrophosphorylated molecules, referred to as phosphoantigens, are powerful stimuli for Vγ9Vδ2 cells, this observation prompted an investigation of phosphoantigens inF. tularensisstrains. TheF. tularensisphosphoantigens triggered in vitro a proliferative response of human Vγ9Vδ2 peripheral blood leukocytes as well as a cytotoxic response and tumor necrosis factor release from a Vγ9Vδ2 T-cell clone. Quantitatively similar phosphoantigenic activity was detected in acellular extracts from two clinical isolates (FSC171 and Schu) and from LVS. Taken together, the chemical nature of the stimulus from the clinical isolates and the significant increase in levels of Vγ9Vδ2 cells in peripheral blood of tularemia patients indicate that phosphoantigens produced by virulent strains ofF. tularensistrigger in vivo expansion of γδ T cells in tularemia.

The role of human fetal thymocytes in the regulation of IgG and IgM production was examined. In pokeweed mitogen (PWM)-driven cocultures between thymocytes and normal adult peripheral blood mononuclear cells (PBMs) significantly enhanced secretion of both IgM (350% expected; P < 0.02) and IgG (450% expected; P < 0.001) was observed. In contrast, adult PBMs or adult T cells neither suppressed nor enhanced IgM or IgG production in coculture with allogeneic adult PBMs. Enhanced immunoglobulin secretion was not found when fetal thymocytes were cocultured with the T-cell-depleted fraction of adult PBMs. These results suggest that human fetal thymocytes (as early as 12 weeks gestational age) can provide helper cell function for both IgM and IgG secretion in PWM-driven cell cultures; however, a more mature T-cell effector is required for expression of this helper function.

Total B-cell numbers, serum immunoglobulin levels, autoantibody formation, and suppressor/helper cell function were evaluated in a group of well-nourished healthy aged individuals (65–103 years). Total B-cell numbers in the elderly were similar to those in young controls. Mean IgG levels were significantly higher (P < 0.05) and mean IgM levels were significantly lower (P < 0.01) in the aged, while IgA levels were comparable to those of young controls. Autoantibodies were found in 28%. Using peripheral blood lymphocyte coculture techniques, we found normal helper/suppressor cell function in this elderly population. As a further investigation of suppressor cell function, we demonstrated that concanavalin A preincubation of peripheral blood lymphocytes from the elderly and from young controls generated equally effective suppressor cells for IgG secretion into coculture supernatants. It is possible that defects in macrophage/phagocytic cell function could lead to the abnormalities in humoral immune responses seen in the elderly.