ESID: A Visual Analytics Tool to Epidemiological Emergencies

Visual analysis tools can help illustrate the spread of infectious diseases and enable informed decisions on epidemiology and public health issues. To create visualisation tools that are intuitive, easy to use, and effective in communicating information, continued research and development focusing on user-centric and methodological design models is extremely important. As a contribution to this topic, this paper presents the design and development of the visual analytics application ESID (Epidemiological Scenarios for Infectious Diseases). The goal of ESID is to provide a platform for rapid assessment of the most effective interventions for infectious disease control. ESID provides spatial-temporal analysis, forecasting, comparison of simulations, interactive filters, and accessibility options. In its current form, it shows the simulations of a hybrid graph-equation-based model as introduced in for infection control. The model can be stratified for different age groups and takes into account the properties of the infectious disease as well as human mobility and contact behaviour.Comment: 6 pages, 5 images and 1 table, Eurovis workshop on visual analytics (EuroVA) 202

ESID: Exploring the Design and Development of a Visual Analytics Tool for Epidemiological Emergencies

Visual analytics tools can help illustrate the spread of infectious diseases and enable informed decisions on epidemiological and public health issues. To create visualisation tools that are intuitive, easy to use, and effective in communicating information, continued research and development focusing on user-centric and methodological design models is extremely important. As a contribution to this topic, this paper presents the design and development process of the visual analytics application ESID (Epidemiological Scenarios for Infectious Diseases). ESID is a visual analytics tool aimed at projecting the future developments of infectious disease spread using reported and simulated data based on sound mathematical-epidemiological models. The development process involved a collaborative and participatory design approach with project partners from diverse scientific fields. The findings from these studies, along with the guidelines derived from them, played a pivotal role in shaping the visualisation tool

Prevalence of Transmitted Drug Resistance and Impact of Transmitted Resistance on Treatment Success in the German HIV-1 Seroconverter Cohort

BACKGROUND: The aim of this study is to analyse the prevalence of transmitted drug resistance, TDR, and the impact of TDR on treatment success in the German HIV-1 Seroconverter Cohort. METHODS: Genotypic resistance analysis was performed in treatment-naïve study patients whose sample was available 1,312/1,564 (83.9% October 2008). A genotypic resistance result was obtained for 1,276/1,312 (97.3%). The resistance associated mutations were identified according to the surveillance drug resistance mutations list recommended for drug-naïve patients. Treatment success was determined as viral suppression below 500 copies/ml. RESULTS: Prevalence of TDR was stable at a high level between 1996 and 2007 in the German HIV-1 Seroconverter Cohort (N = 158/1,276; 12.4%; CI(wilson) 10.7-14.3; p(for trend) = 0.25). NRTI resistance was predominant (7.5%) but decreased significantly over time (CI(Wilson): 6.2-9.1, p(for trend) = 0.02). NNRTI resistance tended to increase over time (NNRTI: 3.5%; CI(Wilson): 2.6-4.6; p(for trend)= 0.07), whereas PI resistance remained stable (PI: 3.0%; CI(Wilson): 2.1-4.0; p(for trend) = 0.24). Resistance to all drug classes was frequently caused by singleton resistance mutations (NRTI 55.6%, PI 68.4%, NNRTI 99.1%). The majority of NRTI-resistant strains (79.8%) carried resistance-associated mutations selected by the thymidine analogues zidovudine and stavudine. Preferably 2NRTI/1PIr combinations were prescribed as first line regimen in patients with resistant HIV as well as in patients with susceptible strains (susceptible 45.3%; 173/382 vs. resistant 65.5%; 40/61). The majority of patients in both groups were treated successfully within the first year after ART-initiation (susceptible: 89.9%; 62/69; resistant: 7/9; 77.8%). CONCLUSION: Overall prevalence of TDR remained stable at a high level but trends of resistance against drug classes differed over time. The significant decrease of NRTI-resistance in patients newly infected with HIV might be related to the introduction of novel antiretroviral drugs and a wider use of genotypic resistance analysis prior to treatment initiation

Structural and Functional Analysis of the LTR Promoter Activity of Group M, N and O

HIV-1 wird in drei genetisch distinkte Gruppen, M (major) N (non-M, non-O) und O (outlier) klassifiziert. Innerhalb der M-Gruppe wurden bislang 9 Subtypen und eine zunehmende Anzahl rekombinanter Viren (CRF, circulating recombinant form) identifiziert. Die Subtypen und CRF sind weltweit in Bezug auf die geographische Region und die Transmissionsgruppe sehr unterschiedlich verbreitet. Neben soziodemographischen Faktoren können die biologischen Eigenschaften des Wirtes und des Virus diese unterschiedliche Prävalenz beeinflussen. Die subtyp-spezifische Sequenzvariation des LTR-Promotors von HIV-1 (long terminal repeat) könnte zu einer divergenten Transkriptionskontrolle der Genexpression führen, die zu subtyp-spezifischen Unterschieden in der Replikationseffizienz und letztendlich in der Ausbreitungsdynamik der Subtypen führen könnte. Auch eine subtyp-spezifische Replikationseffizienz in den unterschiedlichen Zielzellen von HIV-1 wird diskutiert. Zu Beginn dieser Arbeit waren subtyp-spezifische strukturelle Unterschiede der LTR anhand eines limitierten Sets publizierter LTR-Sequenzen abzuleiten. Ausgehend von einem Isolat von Subtyp B, C und von CRF01_AE waren auch funktionale Unterschiede der LTR mit Unterschieden in der LTR-Architektur korreliert worden. Diese Ergebnisse stützten die Hypothese einer divergenten Transkriptionsregulation in Abhängigkeit vom Subtyp der LTR. In dieser Arbeit wurde erstmals die strukturelle Promotororganisation und die funktionale Aktivität des LTR-Promotors nicht nur bei verschiedenen Subtypen und CRF der Gruppe M, sondern auch im Vergleich zur Gruppe N und Gruppe O analysiert. Dazu wurde die provirale LTR von jeweils zwei Vertretern der Subtypen A bis G, von der rekominaten Form CRF01_AE und von jeweils einem Vertreter der Gruppe N und der Gruppe O in einen Luziferase-Reporter-Vektor kloniert. Pro Isolat wurden 4-10 Klone sequenziert und analysiert. In phylogenetischen Analysen wurde die ermittelte LTR-Sequenz im Vergleich zu den aktuellen Referenzsequenzen der LTR aus der HIV-Datenbank klassifiziert und eine detaillierte Strukturanalyse der vorliegenden Transkriptions-faktorbindungsstellen (cis-regulatorische Elemente) durchgeführt. Die LTR-Promotoraktivität repräsentativer Klone wurde in standardisierten transienten Transfektionsassays in der Zelllinie COS-Z-28 untersucht. Aus dem Vergleich der LTR-Promotororganisation folgt, dass es keine Strukturmerkmale gibt, in denen sich die Isolate eines Subtyps spezifisch unterscheiden. Meist wurde bei einem einzelnen Vertreter eines anderen Subtyps oder einer Gruppe ein ähnliches Bindungsmotiv identifiziert. Zum Beispiel wurde das für CRF01_AE bisher als charakteristisch beschriebene veränderte Motiv der TATAA-28-Box, ein wesentliches Element zur Bildung des basalen Transkriptionsinitiationskomplexes, auch bei dem Isolat der N-Gruppe identifiziert. Auch der NFkB-Enhancer gilt als subtyp-spezifisches Merkmal. Bislang galten drei NFkB-Bindungsmotive als charakteristisch für die LTR von Subtyp C-Viren. Es wurde jedoch ein Subtyp C Isolat identifiziert, das nur zwei NFkB-Bindungstellen aufweist, während bei einem Gruppe O-Isolat ein drittes NFkB-Motiv nach Kultivierung der Viren beobachtet wurde. Insgesamt ist der LTR-Promotor des N-Isolates der Gruppe M sehr ähnlich, der Aufbau der O-LTR unterscheidet sich dagegen deutlich von der LTR der M- und der N-Gruppe. Die Promotoren aller Gruppen wiesen eine geringe basale Transkriptionsaktivät auf und waren durch das Tat-Protein von Subtyp B transaktivierbar. Die LTR- Aktivität war bei den einzelnen Virusisolaten unterschiedlich, die Aktivitätsunterschiede waren jedoch nicht auf die Zugehörigkeit zu einer Gruppe oder zu einem Subtyp zurückzuführen. Vielmehr scheinen individuelle Strukturmerkmale der LTR für die unterschiedliche Promotoraktivität verantwortlich zu sein. LTR-Sequenzen mit drei NFkB-Enhancer-Motiven zeigen eine erhöhte basale Promotoraktivität gegenüber LTR-Sequenzen mit den für HIV-1 üblichen zwei NFkB-Enhancer-Motiven, unabhängig von der Subtyp- oder Gruppenzugehörigkeit der LTR. Isolate mit nur einer Bindungsstelle für NFkB zeigen dagegen keinen signifikanten Aktivitätsverlust. Die LTR-Aktivität ist somit nur eingeschränkt korrelierbar mit der Anzahl der NFkB-Enhancer-Motive. Die Transkriptionsaktivität der LTR von HIV-1-Isolaten der Gruppen M, N und O ist somit nicht subtyp- oder gruppen-spezifisch, sondern isolat-spezifisch reguliert. Die Ergebnisse können jedoch eine divergente Transkriptionsregulation in primären Zielzellen von HIV-1 nicht ausschließen.HIV-1 is classified into three distinct genetic groups: M (major), N (non-M; non-O) and O (outlier). Within the group M actually nine subtypes and an increasing number of circulating recombinant forms have been identified. The global spread of subtypes and CRF (circulating recombinant form) differs with respect to geographic regions and the routes of transmission. Besides sociodemographic factors, biological properties of the host and the virus may influence the different prevalences of the subtypes. Subtype-specific sequence variation of the HIV-1 LTR promoter (long terminal repeat) could result in divergent transcriptional control of the gene expression, which in turn may lead to divergent replicative properties and may contribute to the dynamic spread of subtypes. Subtype-specific efficiency of replication depending on the HIV target cells is also discussed. When the project was initiated subtype-specific structural differences of the LTR have been derived by means of a limited set of published LTR sequences. Starting from single isolates of subtype B, C and CRFO1_AE, functional differences of the LTR were correlated with differences of the LTR architecture. These results raise the possibility that the genetic diversity of the LTR may result in HIV-1 strains with different transcriptional regulation . In this study the LTR enhancer/promoter regions of different subtypes of HIV-1 group M were characterized with regard to nucleotide sequence and promotor activity and for the first time they were analysed in comparison to group N and group O LTR promoters. The proviral LTR of the subtypes A to G, of the recombinant form CRFO1_ AE and of one group N and one group O strain was cloned into a luciferase reporter vector. Four to ten clones of each isolate were sequenced and analysed. The LTR-sequences were classified in phylogenetic analysis together with actual reference LTR sequences of the HIV database. A detailed structural analysis of cis- regulating enhancer/promoter elements was performed and the activity of representative LTR promoters was analysed in standardized transient transfection assays in COS-Z-28 cells. There are only few subtype-specific structural features which are common to all isolates of a given subtype. In most cases within a subtype or group individual LTR sequences were identified being similar to other genotypes in single binding motivs. The TATAA-28-box motiv, for example, an essential element of the basic transcription initiation complex, has a characteristic mutation in the LTR of CRF01_AE isolates, but this modification was also found in the N group. Also the structure of the NFkB enhancer is described to be subtype-specific. So far, the LTR of subtype C strains considered to carry three binding sites for NFkB. However, a subtype C isolate was identified, counting only two functional NFkB sites, whereas in the LTR of an group O isolate a third NFkB motiv appeared upon cultivation of the virus. Summarized, the organisation of the LTR enhancer/promoter of the N isolate is very similar to group M isolates, in contrast to the structure of the O-LTR which is distinct from the LTR of the M and the N group. The promoters of all virus strains had low basal transcription activity and were activated by the viral transactivator protein Tat of subtype B. The LTR activities in individual isolates were different, but there was no correlation of these differences with the phylogenetic clade to which they belonged to. Individual structural features of the LTR are not likely to be responsible for the different promoter activities. LTR sequences with three NFkB motivs showed a higher basal promoter activity in comparison to LTR sequences with two NFkB motivs, no matter which subtype or group they belong to, whereas isolates with only one binding site for NFkB had no significant loss of LTR activity. Therefore the correlation of the LTR activity and the number of NFkB motivs is restrictive. In conclusion the LTR-directed transcriptional regulation of HIV-1 isolates from group M, N and O is not specific of the subtype or group, but specific of the individual isolate. Finally, divergent transcriptional regulation in primary target cells of HIV-1 cannot be excluded

Characterization of natural polymorphic sites of the HIV-1 integrase before the introduction of HIV-1 integrase inhibitors in Germany

Introduction: The aim of our study was to analyze the occurrence and evolution of HIV-1 integrase polymorphisms during the HIV-1 epidemic in Germany prior to the introduction of the first integrase inhibitor raltegravir in 2007. Materials and Methods: Plasma samples from drug-naïve HIV-1 infected individuals newly diagnosed between 1986 and 2006 were used to determine PCR-based population sequences of the HIV-1 integrase (amino acids 1–278). The HIV-1 subtype was determined using the REGA HIV-1 subtyping tool. We calculated the frequency of amino acids at each position of the HIV-1 integrase in 337 subtype B strains for the time periods 1986–1989, 1991–1994, 1995–1998, 1999–2002, and 2003–2006. Positions were defined as polymorphic if amino acid variation was >1% in any period. Logistic regression was used to identify trends in amino acid variation over time. Resistance-associated mutations were identified according to the IAS 2013 list and the HIVdb, ANRS and GRADE algorithms. Results: Overall, 56.8% (158/278) amino acid positions were polymorphic and 15.8% (25/158) of these positions exhibited a significant trend in amino acid variation over time. Proportionately, most polymorphic positions (63.3%, 31/49) were detected in the N-terminal zinc finger domain of the HIV-1 integrase. Motifs and residues essential for HIV-1 integrase activity were little polymorphic, but within the minimal non-specific DNA binding region I220-D270 up to 18.1% amino acid variation was noticed, including four positions with significant amino acid variation over time (S230, D232, D256, A265). No major resistance mutations were identified, and minor resistance mutations were rarely observed without trend over time. E157Q considered by HIVdb, ANRS, and GRADE algorithms was the most frequent resistance-associated polymorphism with an overall prevalence of 2.4%. Conclusions: Detailed knowledge of the evolutionary variation of HIV-1 integrase polymorphisms is important to understand the development of resistance in the presence of the drug. Our results will contribute to define the relevance of integrase polymorphisms in HIV-strains resistant to integrase inhibitors and to improve resistance interpretation algorithms

Minor Drug-Resistant HIV Type-1 Variants in Breast Milk and Plasma of HIV Type-1-Infected Ugandan Women after Nevirapine Single-Dose Prophylaxis

Background Nevirapine single-dose (NVP-SD) reduces mother-to-child transmission of HIV type-1 (HIV-1), but frequently induces resistance mutations in the HIV-1 genome. Little is known about drug-resistant HIV-1 variants in the breast milk of women who have taken NVP-SD. Methods Blood and breast milk samples of 39 HIV-1-infected Ugandan women were taken 6–12 weeks after NVP-SD intake. Samples were analysed by population sequencing and allele-specific real-time PCR (AS-PCR) with detection limits for NVP-resistant HIV-1 variants (K103N and Y181C) of &lt;1% of the total viral population. Results AS-PCR results for both plasma and breast milk were obtained for 19 women who constituted the final study group (HIV-1 subtype frequencies were A1 n=11, D n=5, G n=2 and C n=1). A total of 7 (37%) and 10 (53%) women carried NVP-resistant virus in breast milk and plasma, respectively. Overall, 71% (5/7) women with NVP-resistant HIV-1 in breast milk displayed &gt;1 drug-resistant variant. Resistance in breast milk was higher at week 6 (6/13 samples [46%]) compared with week 12 (1/6 samples [17%]). In total, 10 drug-resistant populations harbouring the K103N and/or Y181C mutation were detected in the 19 breast milk samples; 7 (70%) were caused by resistant minorities (&lt;5% of the total HIV-1 population). In the four women with drug-resistant virus in both plasma and breast milk, the mutation patterns differed between the two compartments. Conclusions Minor populations of drug-resistant HIV-1 were frequently found in breast milk of Ugandan women after exposure to NVP-SD. Further studies need to explore the role of minor drug-resistant variants in the postnatal transmission of (resistant) HIV-1. </jats:sec

Improved Testing of Recent HIV-1 Infections with the BioRad Avidity Assay Compared to the Limiting Antigen Avidity Assay and BED Capture Enzyme Immunoassay: Evaluation Using Reference Sample Panels from the German Seroconverter Cohort

Background: The variety and limitations of current laboratory methods for estimating HIV-incidence has driven attempts to improve and standardize the performance of serological ‘Tests for Recent HIV-Infections’ (TRI). Primary and follow-up HIV-1 positive plasma samples from individuals with well-defined dates of infection collected as part of the German Seroconverter Cohort provided specimens highly suitable for use in comparing the performance of three TRIs: the AWARE™ BED™ EIA HIV-1 Incidence test (BED-CEIA), Genetic systems HIV-1/HIV-2 Plus O EIA antibody avidity-based assay (BioRad Avidity) and Sedia™ HIV-1 LAg Avidity EIA (LAg Avidity). Methods: The evaluation panel included 180 specimens: 44 from antiretroviral (ARV)-naïve individuals with recently acquired HIV-infection (≤130 days; 25 B and 19 non-B subtypes) and 136 from long-term (>12 months) infected individuals [101 ARV-naïve subtype B, 16 non-B subtypes, 14 ARV-treated individuals, 5 slow progressors (SLP)]. Results: For long-term infected, ARV-naïve individuals the false recent rates (FRR) of both the BioRad and LAg Avidity assays were 2% (2/101 for subtype B) and 6% (1/16 for subtype ‘non-B’), while the FRR of the BED-CEIA was 7% (7/101 for subtype B) and 25% (4/16 for subtype ‘non-B’) (all p>0.05). Misclassification of ARV-treated individuals and SLP was rare by LAg (1/14, 0/5) and BioRad Avidity assays (2/14, 1/5) but more frequent by BED-CEIA (5/14, 3/5). Among recently-infected individuals (subtype B), 60% (15/25) were correctly classified by BED-CEIA, 88% (22/25) by BioRad Avidity and significantly fewer by LAg (48%, 12/25) compared to BioRad Avidity (p = 0.005) with a higher true-recency rate among non-B infections for all assays. Conclusions: This study using well-characterized specimens demonstrated lower FRRs for both avidity methods than with the BED-CEIA. For recently infected individuals the BioRad Avidity assay was shown to give the most accurate results

Molecular evolution of HIV-1 integrase during the 20 years prior to the first approval of integrase inhibitors

Abstract Background Detailed knowledge of the evolutionary potential of polymorphic sites in a viral protein is important for understanding the development of drug resistance in the presence of an inhibitor. We therefore set out to analyse the molecular evolution of the HIV-1 subtype B integrase at the inter-patient level in Germany during a 20-year period prior to the first introduction of integrase strand inhibitors (INSTIs). Methods We determined 337 HIV-1 integrase subtype B sequences (amino acids 1–278) from stored plasma samples of antiretroviral treatment-naïve individuals newly diagnosed with HIV-1 between 1986 and 2006. Shannon entropy was calculated to determine the variability at each amino acid position. Time trends in the frequency of amino acid variants were identified by linear regression. Direct coupling analysis was applied to detect covarying sites. Results Twenty-two time trends in the frequency of amino acid variants demonstrated either single amino acid exchanges or variation in the degree of polymorphy. Covariation was observed for 17 amino acid variants with a temporal trend. Some minor INSTI resistance mutations (T124A, V151I, K156 N, T206S, S230 N) and some INSTI-selected mutations (M50I, L101I, T122I, T124 N, T125A, M154I, G193E, V201I) were identified at overall frequencies >5%. Among these, the frequencies of L101I, T122I, and V201I increased over time, whereas the frequency of M154I decreased. Moreover, L101I, T122I, T124A, T125A, M154I, and V201I covaried with non-resistance-associated variants. Conclusions Time-trending, covarying polymorphisms indicate that long-term evolutionary changes of the HIV-1 integrase involve defined clusters of possibly structurally or functionally associated sites independent of selective pressure through INSTIs at the inter-patient level. Linkage between polymorphic resistance- and non-resistance-associated sites can impact the selection of INSTI resistance mutations in complex ways. Identification of these sites can help in improving genotypic resistance assays, resistance prediction algorithms, and the development of new integrase inhibitors

First Line Treatment Response in Patients with Transmitted HIV Drug Resistance and Well Defined Time Point of HIV Infection: Updated Results from the German HIV-1 Seroconverter Study

Background: Transmission of drug-resistant HIV-1 (TDR) can impair the virologic response to antiretroviral combination therapy. Aim of the study was to assess the impact of TDR on treatment success of resistance test-guided first-line therapy in the German HIV-1 Seroconverter Cohort for patients infected with HIV between 1996 and 2010. An update of the prevalence of TDR and trend over time was performed. Methods: Data of 1,667 HIV-infected individuals who seroconverted between 1996 and 2010 were analysed. The WHO drug resistance mutations list was used to identify resistance-associated HIV mutations in drug-naïve patients for epidemiological analysis. For treatment success analysis the Stanford algorithm was used to classify a subset of 323 drug-naïve genotyped patients who received a first-line cART into three resistance groups: patients without TDR, patients with TDR and fully active cART and patients with TDR and non-fully active cART. The frequency of virologic failure 5 to 12 months after treatment initiation was determined. Results: Prevalence of TDR was stable at a high mean level of 11.9% (198/1,667) in the HIV-1 Seroconverter Cohort without significant trend over time. Nucleotide reverse transcriptase inhibitor resistance was predominant (6.0%) and decreased significantly over time (OR = 0.92, CI = 0.87–0.98, p = 0.01). Non-nucleoside reverse transcriptase inhibitor (2.4%; OR = 1.00, CI = 0.92–1.09, p = 0.96) and protease inhibitor resistance (2.0%; OR = 0.94, CI = 0.861.03, p = 0.17) remained stable. Virologic failure was observed in 6.5% of patients with TDR receiving fully active cART, 5,6% of patients with TDR receiving non-fully active cART and 3.2% of patients without TDR. The difference between the three groups was not significant (p = 0.41). Conclusion: Overall prevalence of TDR remained stable at a rather high level. No significant differences in the frequency of virologic failure were identified during first-line cART between patients with TDR and fully-active cART, patients with TDR and non-fully active cART and patients without TDR