Severe lower respiratory tract infection in infants and toddlers from a non-affluent population: viral etiology and co-detection as risk factors

Abstract Background Lower respiratory tract infection (LRTI) is a major cause of pediatric morbidity and mortality, especially among non-affluent communities. In this study we determine the impact of respiratory viruses and how viral co-detections/infections can affect clinical LRTI severity in children in a hospital setting. Methods Patients younger than 3 years of age admitted to a tertiary hospital in Brazil during the months of high prevalence of respiratory viruses had samples collected from nasopharyngeal aspiration. These samples were tested for 13 different respiratory viruses through real-time PCR (rt-PCR). Patients were followed during hospitalization, and clinical data and population characteristics were collected during that period and at discharge to evaluate severity markers, especially length of hospital stay and oxygen use. Univariate regression analyses identified potential risk factors and multivariate logistic regressions were used to determine the impact of specific viral detections as well as viral co-detections in relation to clinical outcomes. Results We analyzed 260 episodes of LRTI with a viral detection rate of 85% (n = 222). Co-detection was observed in 65% of all virus-positive episodes. The most prevalent virus was Respiratory Syncytial Virus (RSV) (54%), followed by Human Metapneumovirus (hMPV) (32%) and Human Rhinovirus (HRV) (21%). In the multivariate models, infants with co-detection of HRV + RSV stayed 4.5 extra days (p = 0.004), when compared to infants without the co-detection. The same trends were observed for the outcome of days of supplemental oxygen use. Conclusions Although RSV remains as the main cause of LRTI in infants our study indicates an increase in the length of hospital stay and oxygen use in infants with HRV detected by RT-PCR compared to those without HRV. Moreover, one can speculate that when HRV is detected simultaneously with RSV there is an additive effect that may be reflected in more severe clinical outcome. Also, our study identified a significant number of children infected by recently identified viruses, such as hMPV and Human Bocavirus (HBov), and this is a novel finding for poor communities from developing countries.

High specificity of line-immunoassay based algorithms for recent HIV-1 infection independent of viral subtype and stage of disease

Background Serologic testing algorithms for recent HIV seroconversion (STARHS) provide important information for HIV surveillance. We have shown that a patient's antibody reaction in a confirmatory line immunoassay (INNO-LIATM HIV I/II Score, Innogenetics) provides information on the duration of infection. Here, we sought to further investigate the diagnostic specificity of various Inno-Lia algorithms and to identify factors affecting it. Methods Plasma samples of 714 selected patients of the Swiss HIV Cohort Study infected for longer than 12 months and representing all viral clades and stages of chronic HIV-1 infection were tested blindly by Inno-Lia and classified as either incident (up to 12 m) or older infection by 24 different algorithms. Of the total, 524 patients received HAART, 308 had HIV-1 RNA below 50 copies/mL, and 620 were infected by a HIV-1 non-B clade. Using logistic regression analysis we evaluated factors that might affect the specificity of these algorithms. Results HIV-1 RNA <50 copies/mL was associated with significantly lower reactivity to all five HIV-1 antigens of the Inno-Lia and impaired specificity of most algorithms. Among 412 patients either untreated or with HIV-1 RNA ≥50 copies/mL despite HAART, the median specificity of the algorithms was 96.5% (range 92.0-100%). The only factor that significantly promoted false-incident results in this group was age, with false-incident results increasing by a few percent per additional year. HIV-1 clade, HIV-1 RNA, CD4 percentage, sex, disease stage, and testing modalities exhibited no significance. Results were similar among 190 untreated patients. Conclusions The specificity of most Inno-Lia algorithms was high and not affected by HIV-1 variability, advanced disease and other factors promoting false-recent results in other STARHS. Specificity should be good in any group of untreated HIV-1 patients.

Early antiretroviral therapy during primary HIV-1 infection results in a transient reduction of the viral setpoint upon treatment interruption

Long-term benefits of combination antiretroviral therapy (cART) initiation during primary HIV-1 infection are debated.

Constrained pattern of viral evolution in acute and early HCV infection limits viral plasticity

Cellular immune responses during acute Hepatitis C virus (HCV) and HIV infection are a known correlate of infection outcome. Viral adaptation to these responses via mutation(s) within CD8+ T-cell epitopes allows these viruses to subvert host immune control. This study examined HCV evolution in 21 HCV genotype 1-infected subjects to characterise the level of viral adaptation during acute and early HCV infection. Of the total mutations observed 25% were within described CD8+ T-cell epitopes or at viral adaptation sites. Most mutations were maintained into the chronic phase of HCV infection (75%). The lack of reversion of adaptations and high proportion of silent substitutions suggests that HCV has structural and functional limitations that constrain evolution. These results were compared to the pattern of viral evolution observed in 98 subjects during a similar phase in HIV infection from a previous study. In contrast to HCV, evolution during acute HIV infection is marked by high levels of amino acid change relative to silent substitutions, including a higher proportion of adaptations, likely reflecting strong and continued CD8+ T-cell pressure combined with greater plasticity of the virus. Understanding viral escape dynamics for these two viruses is important for effective T cell vaccine design.

Ambiguous nucleotide calls from population-based sequencing of HIV-1 are a marker for viral diversity and the age of infection

The time passed since the infection of a human immunodeficiency virus (HIV)-infected individual (the age of infection) is an important but often only poorly known quantity. We assessed whether the fraction of ambiguous nucleotides obtained from bulk sequencing as done for genotypic resistance testing can serve as a proxy of this parameter.

"Self" and "nonself" manipulation of interferon defense during persistent infection: bovine viral diarrhea virus resists alpha/beta interferon without blocking antiviral activity against unrelated viruses replicating in its host cells

Bovine viral diarrhea virus (BVDV), together with Classical swine fever virus (CSFV) and Border disease virus (BDV) of sheep, belongs to the genus Pestivirus of the Flaviviridae. BVDV is either cytopathic (cp) or noncytopathic (ncp), as defined by its effect on cultured cells. Infection of pregnant animals with the ncp biotype may lead to the birth of persistently infected calves that are immunotolerant to the infecting viral strain. In addition to evading the adaptive immune system, BVDV evades key mechanisms of innate immunity. Previously, we showed that ncp BVDV inhibits the induction of apoptosis and alpha/beta interferon (IFN-alpha/beta) synthesis by double-stranded RNA (dsRNA). Here, we report that (i) both ncp and cp BVDV block the induction by dsRNA of the Mx protein (which can also be induced in the absence of IFN signaling); (ii) neither biotype blocks the activity of IFN; and (iii) once infection is established, BVDV is largely resistant to the activity of IFN-alpha/beta but (iv) does not interfere with the establishment of an antiviral state induced by IFN-alpha/beta against unrelated viruses. The results of our study suggest that, in persistent infection, BVDV is able to evade a central element of innate immunity directed against itself without generally compromising its activity against unrelated viruses ("nonself") that may replicate in cells infected with ncp BVDV. This highly selective "self" and "nonself" model of evasion of the interferon defense system may be a key element in the success of persistent infection in addition to immunotolerance initiated by the early time point of fetal infection.

Evidence of viral adaptation to HLA class I-restricted immune pressure in chronic hepatitis C virus infection

Cellular immune responses are an important correlate of hepatitis C virus (HCV) infection outcome. These responses are governed by the host's human leukocyte antigen (HLA) type, and HLA-restricted viral escape mutants are a critical aspect of this host-virus interaction. We examined the driving forces of HCV evolution by characterizing the in vivo selective pressure(s) exerted on single amino acid residues within nonstructural protein 3 (NS3) by the HLA types present in two host populations. Associations between polymorphisms within NS3 and HLA class I alleles were assessed in 118 individuals from Western Australia and Switzerland with chronic hepatitis C infection, of whom 82 (69%) were coinfected with human immunodeficiency virus. The levels and locations of amino acid polymorphisms exhibited within NS3 were remarkably similar between the two cohorts and revealed regions under functional constraint and selective pressures. We identified specific HCV mutations within and flanking published epitopes with the correct HLA restriction and predicted escaped amino acid. Additional HLA-restricted mutations were identified that mark putative epitopes targeted by cell-mediated immune responses. This analysis of host-virus interaction reveals evidence of HCV adaptation to HLA class I-restricted immune pressure and identifies in vivo targets of cellular immune responses at the population level.

Immunohistochemical diagnosis of persistent infection with Bovine Viral Diarrhea Virus (BVDV) on skin biopsies

Detection of persistent infection with BovineViral Diarrhea Virus (BVDV) is essential for both epidemiological and clinical reasons. In addition to the classical virological methods such as virus isolation in tissue culture, ELISA and RT-PCR, immunohistochemistry of skin biopsies has become a useful and reliable tool. Assuming that the presence of BVDV antigen in skin structures is restricted to persistent infection, this method could differentiate from transient infection. In order to answer this question, 6 calves were experimentally infected orally with a non-cytopathic genotype 1 BVDV strain belonging to the subtype k.The calves developed fever, mucopurulent nasal discharge, coughing and leucopenia with relative lymphopenia. Immunohistochemistry of skin biopsies taken daily up to day 13-post infection did not reveal any evidence of BVDV infection. BVDV was, however, isolated from blood samples on cell cultures. Anti-NS3-antibody-ELISA and serum neutralization tests showed that all six calves seroconverted. We conclude that in acute BVDV infections, with genotype 1 and the subtypes found in Switzerland (b, e, h and k) viral antigen is not found in epidermal structures of the skin. In contrast, persistently infected animals test positive for BVD viral antigen by immunohistochemistry of the skin.

Comparison of five diagnostic methods for detecting bovine viral diarrhea virus infection in calves

Five diagnostic techniques performed on skin biopsies (shoulder region) and/or serum were compared for detection of bovine viral diarrhea virus infection in 224 calves 0-3 months of age, 23 calves older than 3 months but younger than 7 months, and 11 cattle older than 7 months. The diagnostic methods used were immunohistochemistry (IHC), 2 commercial antigen ELISAs, 1 commercial antibody ELISA, and real-time RT-PCR. Results of 249 out of 258 skin and serum samples were identical and correlated within the 3 antigen detection methods and the real-time RT-PCR used. Twenty-six of these 249 samples were BVDV-positive with all antigen detection methods and the real-time RT-PCR. Nine out of 258 samples yielding discordant results were additionally examined by RT-PCR, RT-PCR Reamplification (ReA), and antigen ELISA I on serum and by immunohistochemistry on formalin fixed and paraffin-embedded skin biopsies. Virus isolation and genotyping was performed as well on these discordant samples. In 3 cases, transiently infected animals were identified. Two samples positive by real-time RT-PCR were interpreted as false positive and were ascribed to cross-contamination. The antigen ELISA II failed to detect 2 BVDV-positive calves due to the presence of maternal antibodies; the cause of 2 false-positive cases in this ELISA remained undetermined. Only persistently infected animals were identified in skin samples by IHC or antigen ELISA I. The 3 antigen detection methods and the real-time RT-PCR used in parallel had a high correlation rate (96.5%) and similar sensitivity and specificity values.

Bovine viral diarrhea virus in free-ranging wild ruminants in Switzerland: low prevalence of infection despite regular interactions with domestic livestock

ABSTRACT: BACKGROUND: In the frame of an eradication program for bovine viral diarrhea (BVD) in Swiss livestock, the question was raised whether free-ranging wildlife could threaten the success of this sanitary measure. Therefore, we conducted serological and virological investigations on BVD virus (BVDV) infections in the four indigenous wild ruminant species (roe deer, red deer, Alpine chamois and Alpine ibex) from 2009 to 2011, and gathered information on interactions between wild and domestic ruminants in an alpine environment by questionnaire survey. RESULTS: Thirty-two sera out of 1'877 (1.7%, 95% confidence interval [CI] 1.2-2.4) were seropositive for BVDV, and a BVDV1 sub genotype h virus was found in a seropositive chamois (0.05%, 95% CI 0.001-0.3). The seropositive animals originated from sub-alpine or alpine regions and significantly more seropositive red deer, chamois and ibex than roe deer were found. There were no statistically significant differences between sampling units, age classes, genders, and sampling years. The obtained prevalences were significantly lower than those documented in livestock, and most positive wild ruminants were found in proximity of domestic outbreaks. Additionally, BVDV seroprevalence in ibex was significantly lower than previously reported from Switzerland. The survey on interspecific interactions revealed that interactions expected to allow BVDV transmission, from physical contacts to non-simultaneous use of the same areas, regularly occur on pastures among all investigated ruminant species. Interactions involving cervids were more often observed with cattle than with small ruminants, chamois were observed with all three domestic species, and ibex interacted mostly with small ruminants. Interactions related to the use of anthropogenic food sources were frequently observed, especially between red deer and cattle in wintertime. CONCLUSIONS: To our knowledge, this is the first report of BVDV RNA isolated from an Alpine chamois. Nevertheless, our results suggest that BVDV infections are only sporadic in Swiss wild ruminants, despite regular occurrence of interactions with potentially infected livestock. Overall, serological, virological and ethological data indicate that wildlife is currently an incidental spill-over host and not a reservoir for BVDV in Switzerland.

Impact of viral load and the duration of primary infection on HIV transmission

Abstract Objectives: HIV 'treatment as prevention' (TasP) describes early treatment of HIV-infected patients intended to reduce viral load and transmission. Crucial assumptions for estimating TasP's effectiveness are the underlying estimates of transmission risk. We aimed to determine transmission risk during primary infection, and of the relation of HIV transmission risk to viral load. Design: A systematic review and meta-analysis. Methods: We searched PubMed and Embase databases for studies that established a relationship between viral load and transmission risk, or primary infection and transmission risk, in serodiscordant couples. We analysed assumptions about the relationship between viral load and transmission risk, and between duration of primary infection and transmission risk. Results: We found 36 eligible articles, based on six different study populations. Studies consistently found that larger viral loads lead to higher HIV transmission rates, but assumptions about the shape of this increase varied from exponential increase to saturation. The assumed duration of primary infection ranged from 1.5 to 12 months; for each additional month, the log10 transmission rate ratio between primary and asymptomatic infection decreased by 0.40. Conclusion: Assumptions and estimates of the relationship between viral load and transmission risk, and the relationship between primary infection and transmission risk, vary substantially and predictions of TasP's effectiveness should take this uncertainty into account.

Additive effects of HLA alleles and innate immune genes determine viral outcome in HCV infection

BACKGROUND Chronic HCV infection is a leading cause of liver-related morbidity globally. The innate and adaptive immune responses are thought to be important in determining viral outcomes. Polymorphisms associated with the IFNL3 (IL28B) gene are strongly associated with spontaneous clearance and treatment outcomes. OBJECTIVE This study investigates the importance of HLA genes in the context of genetic variation associated with the innate immune genes IFNL3 and KIR2DS3. DESIGN We assess the collective influence of HLA and innate immune genes on viral outcomes in an Irish cohort of women (n=319) who had been infected from a single source as well as a more heterogeneous cohort (Swiss Cohort, n=461). In the Irish cohort, a number of HLA alleles are associated with different outcomes, and the impact of IFNL3-linked polymorphisms is profound. RESULTS Logistic regression was performed on data from the Irish cohort, and indicates that the HLA-A*03 (OR 0.36 (0.15 to 0.89), p=0.027) -B*27 (OR 0.12 (0.03 to 0.45), p=<0.001), -DRB1*01:01 (OR 0.2 (0.07 to 0.61), p=0.005), -DRB1*04:01 (OR 0.31 (0.12 to 0.85, p=0.02) and the CC IFNL3 rs12979860 genotypes (OR 0.1 (0.04 to 0.23), p<0.001) are significantly associated with viral clearance. Furthermore, DQB1*02:01 (OR 4.2 (2.04 to 8.66), p=0.008), KIR2DS3 (OR 4.36 (1.62 to 11.74), p=0.004) and the rs12979860 IFNL3 'T' allele are associated with chronic infection. This study finds no interactive effect between IFNL3 and these Class I and II alleles in relation to viral clearance. There is a clear additive effect, however. Data from the Swiss cohort also confirms independent and additive effects of HLA Class I, II and IFNL3 genes in their prediction of viral outcome. CONCLUSIONS This data supports a critical role for the adaptive immune response in the control of HCV in concert with the innate immune response.

Model for ranking freshwater fish farms according to their risk of infection and illustration for viral haemorrhagic septicaemia

We developed a model to calculate a quantitative risk score for individual aquaculture sites. The score indicates the risk of the site being infected with a specific fish pathogen (viral haemorrhagic septicaemia virus (VHSV); infectious haematopoietic necrosis virus, Koi herpes virus), and is intended to be used for risk ranking sites to support surveillance for demonstration of zone or member state freedom from these pathogens. The inputs to the model include a range of quantitative and qualitative estimates of risk factors organised into five risk themes (1) Live fish and egg movements; (2) Exposure via water; (3) On-site processing; (4) Short-distance mechanical transmission; (5) Distance-independent mechanical transmission. The calculated risk score for an individual aquaculture site is a value between zero and one and is intended to indicate the risk of a site relative to the risk of other sites (thereby allowing ranking). The model was applied to evaluate 76 rainbow trout farms in 3 countries (42 from England, 32 from Italy and 2 from Switzerland) with the aim to establish their risk of being infected with VHSV. Risk scores for farms in England and Italy showed great variation, clearly enabling ranking. Scores ranged from 0.002 to 0.254 (mean score 0.080) in England and 0.011 to 0.778 (mean of 0.130) for Italy, reflecting the diversity of infection status of farms in these countries. Requirements for broader application of the model are discussed. Cost efficient farm data collection is important to realise the benefits from a risk-based approach.

Hepatitis B Infection, Viral Load and Resistance in HIV-Infected Patients in Mozambique and Zambia.

BACKGROUND Few data on the virological determinants of hepatitis B virus (HBV) infection are available from southern Africa. METHODS We enrolled consecutive HIV-infected adult patients initiating antiretroviral therapy (ART) at two urban clinics in Zambia and four rural clinics in Northern Mozambique between May 2013 and August 2014. HBsAg screening was performed using the Determine® rapid test. Quantitative real-time PCR and HBV sequencing were performed in HBsAg-positive patients. Risk factors for HBV infection were evaluated using Chi-square and Mann-Whitney tests and associations between baseline characteristics and high level HBV replication explored in multivariable logistic regression. RESULTS Seventy-eight of 1,032 participants in Mozambique (7.6%, 95% confidence interval [CI]: 6.1-9.3) and 90 of 797 in Zambia (11.3%, 95% CI: 9.3-13.4) were HBsAg-positive. HBsAg-positive individuals were less likely to be female compared to HBsAg-negative ones (52.3% vs. 66.1%, p<0.001). Among 156 (92.9%) HBsAg-positive patients with an available measurement, median HBV viral load was 13,645 IU/mL (interquartile range: 192-8,617,488 IU/mL) and 77 (49.4%) had high values (>20,000 UI/mL). HBsAg-positive individuals had higher levels of ALT and AST compared to HBsAg-negative ones (both p<0.001). In multivariable analyses, male sex (adjusted odds ratio: 2.59, 95% CI: 1.22-5.53) and CD4 cell count below 200/μl (2.58, 1.20-5.54) were associated with high HBV DNA. HBV genotypes A1 (58.8%) and E (38.2%) were most prevalent. Four patients had probable resistance to lamivudine and/or entecavir. CONCLUSION One half of HBsAg-positive patients demonstrated high HBV viremia, supporting the early initiation of tenofovir-containing ART in HIV/HBV-coinfected adults.

Cytotoxic T lymphocytes and viral turnover in HIV type 1 infection

To understand the role of the immune system in limiting HIV type 1 replication, it is critical to know to what extent the rapid turnover of productively infected cells is caused by viral cytopathicity or by immune-mediated lysis. We show that uncultured peripheral blood mononuclear cells of many patients contain cytotoxic T lymphocytes (CTL) that lyse target cells—at plausible peripheral blood mononuclear cell-to-target ratios—with half-lives of less than 1 day. In 23 patients with CD4 counts ranging from 10 to 900 per μl, the average rate of CTL-mediated lysis corresponds to a target cell half-life of 0.7 day. We develop mathematical models to calculate the turnover rate of infected cells subjected to immune-mediated lysis and viral cytopathicity and to estimate the fraction of cells that are killed by CTL as opposed to virus. The models provide new interpretations of drug treatment dynamics and explain why the observed rate of virus decline is roughly constant for different patients. We conclude that in HIV type 1 infection, CTL-mediated lysis can reduce virus load by limiting virus production, with small effects on the half-life of infected cells.