HIV/AIDS Programme MEETING REPORT ON WHO RESNET ANNUAL MEETING 4 MARCH 2012 BILL AND MELINDA GATES FOUNDATION, SEATTLE, USA

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1 HIV/AIDS Programme MEETING REPORT ON WHO RESNET ANNUAL MEETING 4 MARCH 2012 BILL AND MELINDA GATES FOUNDATION, SEATTLE,

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3 HIV/AIDS Programme MEETING REPORT ON WHO RESNET ANNUAL MEETING 4 MARCH 2012 BILL AND MELINDA GATES FOUNDATION, SEATTLE,

4 WHO Library Cataloguing-in-Publication Data Meeting report on WHO RESNET annual meeting, 4 March 2012, Bill and Melinda Gates Foundation, Seattle,. 1.Drug resistance, Viral 2.Anti-HIV agents therapeutic use. 3.Antiretroviral therapy, Highly active therapeutic use. 4.HIV infections epidemiology. 5.Population surveillance. 6.World health. 7.International cooperation. 8.Financing, Organized - utilization. I.World Health Organization. ISBN (NLM classification: QV 268.5) World Health Organization 2012 All rights reserved. Publications of the World Health Organization are available on the WHO web site ( or can be purchased from WHO Press, World Health Organization, 20 Avenue Appia, 1211 Geneva 27, Switzerland (tel.: ; fax: ; bookorders@who.int). Requests for permission to reproduce or translate WHO publications whether for sale or for noncommercial distribution should be addressed to WHO Press through the WHO web site ( copyright_form/en/index.html). The designations employed and the presentation of the material in this publication do not imply the expression of any opinion whatsoever on the part of the World Health Organization concerning the legal status of any country, territory, city or area or of its authorities, or concerning the delimitation of its frontiers or boundaries. Dotted lines on maps represent approximate border lines for which there may not yet be full agreement. The mention of specific companies or of certain manufacturers products does not imply that they are endorsed or recommended by the World Health Organization in preference to others of a similar nature that are not mentioned. Errors and omissions excepted, the names of proprietary products are distinguished by initial capital letters. All reasonable precautions have been taken by the World Health Organization to verify the information contained in this publication. However, the published material is being distributed without warranty of any kind, either expressed or implied. The responsibility for the interpretation and use of the material lies with the reader. In no event shall the World Health Organization be liable for damages arising from its use. This publication contains the collective views of an international group of experts and does not necessarily represent the decision or the policies of the World Health Organization. Printed in Switzerland.

5 WHO ResNet Annual Meeting BACKGROUND The World Health Organization s Global Network HIVResNet, which benefits from technical support of HIVDR experts worldwide, was formed to address concerns regarding HIV drug resistance (HIVDR) emergence and to develop strategies to monitor and prevent HIVDR. In 2005, under ResNet endorsement, the World Health Organization (WHO) developed a public health strategy to minimize and assess HIVDR in countries scaling up antiretroviral therapy (ART) with a special focus on resourcelimited settings (RLS). Since 2005, standardized methods to inform population-based selection of first- and second-line ART regimens and support national programs to optimize patient care and minimize the emergence and transmission of HIVDR have been developed. Through the implementation of WHO recommended protocols, a considerable amount of data on transmitted and acquired HIV drug resistance has been produced. The objectives of this ResNet consultation was to provide an update on the ongoing activities in HIV ResNet, obtain ResNet endorsement for newly developed components in the global HIV drug resistance surveillance and monitoring strategy, and discuss the scenarios to strengthen implementation and the strategy in the future. 1

6 Bill and Melinda Gates Foundation 4 March 2012, Seattle, SUMMARY PRESENTATIONS SESSION A IMPLEMENTATION, RESULTS AND INTERPRETATION OF HIVDR WHO RECOMMENDED SURVEYS WHO HIVDR data from HIVDR Early Warning Indicators and Surveillance of acquired and transmitted HIVDR were presented. 1. MONITORING OF HIVDR EARLY WARNING INDICATORS (EWI) FROM ART CLINICS HIVDR EWIs assess factors at individual clinics which are known to be associated with emergence of HIVDR. Results provide an alert for corrective action to improve ART clinic performance and programme functioning. As of 2011, 124 rounds of EWI monitoring in 58 countries in >2000 clinics were reported. The implementation of EWI monitoring has progressively increased over time with 144 performed cumulatively since inception. EWIs 1, 2, 3 and 5 (see table 1) were those most frequently reported, particularly in Africa and Asia. To facilitate EWI monitoring, WHO developed an EXCEL based tool which has proven to be simple to use and which supports standardization. TABLE 1 EARLY WARNING INDICATORS EWI EWI Target 1. Prescribing practices 100% 2. Lost to follow-up at 12 months 20% 3. Retention on first-line ART at 12 months 70% 4. On-time drug pick up 90% 5. On-time appointment keeping 80% 6. Drug supply continuity 100% 8. Viral load <1000 copies/ml at 12 months 70% In August 2011, an expert panel meeting review meeting was held in Geneva to consider revisions to currently recommended EWIs and targets. After a critical review of available medical literature using the GRADE methodology, panel made several recommendations to WHO. Specifically recommendations were made to simplify EWI definitions, account for implementation challenges, harmonize with other routinely reported indicators and adjust based on new evidence. The suggested revised set of indicators 2

7 WHO ResNet Annual Meeting which is designed to be implemented as a package is presented in Table 2. Notably, the total number of indicators was reduced from seven to four. The fifth indicator (viral load suppression at 12 months), is considered conditional and is designed to be implemented only at sites where routine viral load monitoring is performed for all patients 12 months after ART initiation. TABLE 2. REVISED EWI 2011 HIV Drug Resistance Early Warning Indicator Score Card Early Warning Indicator Status Target 1. On-time pill pick-up Red <80% adherence in 90% of patients Amber 80 95% adherence in 90% of patients Green >95% adherence in 90% of patients 2. Retention in care Red <75% retained after 12 months ART Amber, 75 85% retained after 12 months ART Green >85% retained after 12 months ART 3. Pharmacy stock-outs Red <100% of a 12 month period with no stock-outs Green 100% of a 12 month period with no stock-outs 4. Prescribing practices Red >0% dispensing of mono or dual therapy Green 0% dispensing of mono or dual therapy 5. Virological suppression# Red <70% viral load suppression after 12 months of ART Amber 70 85% viral load suppression after 12 months of ART Green >85% viral load suppression after 12 months of ART Notes: Red (poor performance, below desired level) Amber (fair performance, not yet at desired level but progressing towards desired level) Green (excellent performance, achieving desired level) Grey (data not available) # Targets for virological suppression in children <2 years old Red <60% viral load suppression after 12 months of ART Amber 60 70% viral load suppression after 12 months of ART Green >70% viral load suppression after 12 months of ART The revised set of indicators is anticipated to require substantially less data abstraction and whenever possible indicator definitions were harmonized with UNGASS or PEPFAR indicators and a target appropriate to HIVDR was established. To further stimulate widespread uptake of EWIs, sustainability and ownership, data abstraction and reporting responsibilities will be delegated to ART clinics. 3

8 Bill and Melinda Gates Foundation 4 March 2012, Seattle, 2. SURVEILLANCE OF ACQUIRED HIVDR Thirty-four monitoring surveys were conducted at 34 ART clinics in 10 countries (Burundi, India, Malawi, Mozambique, Nigeria, Zimbabwe, Cameroon, Swaziland, Indonesia, and Namibia) for a total of 4251 patients surveyed between 2006 and In a combined analysis of all available data, approximately 6% (N=3604 from the 15 surveys with available data) of patients initiating ART at the clinics (naïve and ARV exposed) had detected HIVDR at time of ART initiation. The proportion of patients with virological suppression (<1,000 copies/ml) at 12 months (N=2150) was 90% for patients retained in care and alive and 70% when LTFU and ART stops are included and classified virological failure. Approximately 35% of those with virological failure at 12 month had wild type suggesting that adherence counselling should be strengthened. At 12 months, in patients failing ART (N=128) the prevalence of thymidine analogue mutations (TAMs) remained limited (4.7% 3 TAMs). 67% had NNRTI resistance including 52% with 184I/V and 6.3% with K65R. Surveys proved useful as they provided site-specific assessments of viral load suppression, especially in sites where viral load monitoring was not routinely performed. However, survey results cannot be used to inform on the appropriate composition of second line regimens beyond months as the number of specimens from patients was too small to allow generalisation and in many RLS patient may remain on failing regimens for long durations and subsequently accrue additional HIVDR. During the discussion it was highlighted how measuring viral load is critical to minimize HIVDR by detecting early failure, thus preventing accrual of additional HIVDR, and that individual HIVDR testing is not critical to inform switch to second-line as resistance to protease inhibitors is not observed and drug options are limited. Drawbacks of current methodologies are addressed in a new cross-sectional survey of acquired HIVDR which is presented as part of the new global strategy in section C. 3. SURVEILLANCE OF TRANSMITTED HIVDR Ninety-four surveys (54 surveys with available data from 22 countries) were conducted between , mostly between Overall, 81% of surveys showed low TDR (<5%), suggesting that TDR was low in the areas and populations assessed at the time the surveys were conducted. 19% of surveys showed moderate TDR (5-15%) in and among the 11 surveys conducted in 2009, 5 (45%) showed moderate TDR (5-15%). Data from surveys conducted from 2005 and 2009 suggest that TDR remained low in most settings surveyed, but moderate TDR results alert programme planners to potential increases. Data from the surveys conducted thus far are insufficient to identify trends of TDR over time and no treatment guidelines changes are warranted before further investigation. However, moderate results warrant attention and concern and appropriate programme and public health action should be taken to confirm results and perform additional assessments as recommended in WHO TDR survey guidance. 4

9 WHO ResNet Annual Meeting SESSION B CONTEXTUALIZATION WHO HIVDR DATA IN THE EXISTING PUBLISHED LITERATURE 1. HIVDR IN RECENTLY AND CHRONICALLY INFECTED POPULATIONS : A META-ANALYSIS (RAVI GUPTA) Dr. Gupta undertook a meta-analysis commissioned by the WHO. Papers and conference abstracts published between January 2001 and July 2011 describing resistance in untreated individuals in RLS were searched. Additional unpublished data on HIVDR from national HIV treatment programs were included. Of a total of 26,635 patient genotypes, 14,098 came from studies in sub-saharan Africa, 5735 from Asia and 6802 from Central and South America. The prevalence of any drug resistance at nine years post rollout is estimated to be between 10 and 15% in Eastern Africa (9% for NNRTI). East Africa had the highest average modeled rate of increase [38.9 %/yr (p<0.0001]] since rollout, with NNRTI resistance increasing at 32.9%/yr (p<0.0001). The estimated rate of increase in West and Southern Africa was 10.4% (p=0.0196) and 9.6% (p=0.0341) per year respectively with current prevalence at 6 and 3.5% respectively. There was no change in resistance over time in Central and South America, with a current prevalence estimated at 7.4%. The findings, soon to be published, highlight the importance of HIVDR surveillance. Further analysis will be considered to account for the change in coverage over time, and assess difference between WHO surveys result and published literature. 2. ACQUIRED HIVDR: A META-ANALYSIS (DANIEL DAVIS) Dr. Davis presented a systematic review undertaken under commission of WHO. The aim of the review was to establish the prevalence of acquired HIVDR in RLS and determine how prevalence varies with duration on ART., Papers and conference abstracts published between January 2001 and July 2011 were reviewed. Despite a wide variation in reporting of study data, 22 studies for a total of 2041 patients were included in the quantitative synthesis. Overall prevalence was highest in East Africa 0.65, West Africa 0.57, and South Africa 0.62 (p 0.001). Evidence for increasing prevalence with duration on ART was found: prevalence (any mutation) increased by 12% per year of ART treatment (p=0.06); NRTI prevalence increased by 5% per year of ART treatment (p=0.47); M184V prevalence increased by 14% per year of ART treatment (p=0.09); and TAM prevalence increased by 14% per year of ART treatment (p=0.04). The effect does not appear to vary by region, but the analysis is likely under-powered to assess regional variations. As suggested by the group further analysis may include: sensitivity analysis using patients in virological failure as denominator; accumulation of multiple TAMs; influence of gender. 5

10 Bill and Melinda Gates Foundation 4 March 2012, Seattle, SESSION C WHO HIVDR PREVENTION AND ASSESSMENT STRATEGY NEW DIRECTIONS 1. SURVEILLANCE OF ACQUIRED HIVDR IN POPULATIONS FAILING ART- A CROSS SECTIONAL METHOD Despite providing comprehensive results, the implementation of the prospective survey was challenging and its widespread implementation was limited. These challenges have led to descriptions of HIVDR which are unlikely to represent the true picture of HIVDR in populations failing ART. For this reason a simpler retrospective cohort survey has been developed to: classify the proportion of adult patients with virological failure and months after initiation of first-line ART; the proportion of paediatric patients with virological failure 2 months after initiation of first-line ART; and provide a cumulate dataset of detected HIVDR amongst patients with virological failure. The new survey method is based on Lot quality assurance sampling (LQAS) and is designed to have a maximum specimen collection period of 3 months. It is anticipated that this retrospective method will be easier to implement in large numbers of sentinel ART clinics thus providing a more representative description of acquired HIVDR within countries scaling up ART. The retrospective method of acquired HIVDR will be piloted in 2012 in Kenya, Namibia, Tanzania, and South Africa (KwaZulu-Natal). Countries considering surveillance of acquired HIVDR should contact WHO to assess whether it is more relevant to use the existing prospective survey of acquired HIVDR or to pilot the new cross-sectional method. 2. SURVEILLANCE OF HIVDR IN POPULATIONS INITIATING ART In discussions held between WHO, the US Centers for Disease Control and Prevention (CDC), Office of the Global AIDS Coordinator (OGAC) and the Public Health Agency of Canada (PHAC), development and implementation of a m method to assess HIVDR in populations initiation ART (pre-treatment) has been considered. Surveillance of HIVDR in populations initiating ART would support public health decision making. Specifically, data would inform the choice of first-and support cost-effectiveness modelling of more individual viral load testing scale-up and individual patient HIVDR genotyping. A protocol to assess HIVDR in a representative sample of patients initiating ART within a national ART program is being developed. 3. SURVEILLANCE OF HIVDR IN HIV-INFECTED AND ART NAIVE INFANTS <18 MONTHS The key features of this protocol were presented to the group. The protocol assess the prevalence of HIVDR by using remnant dried blood spots (DBS) from early infant diagnosis (EID) services and explores factors associated with resistance in this population. A nationally representative sample is selected and samples are processed in a WHO-appointed laboratory (Atlanta, or Montpellier, France). Survey results are 6

11 WHO ResNet Annual Meeting anticipated to inform the choice of first-line therapy and future treatment strategies. Through the support of WHO and CDC, the survey will be piloted in Uganda, Malawi, Zimbabwe, Swaziland and Cote d Ivoire. 4. FEEDBACK FROM WHO EXPERT MEETING (GENEVA, FEB 14 15, 2012) 4.1 Revision of epidemiological criteria for definition of recently infection for surveys of transmitted drug resistance Challenges faced during survey implementation were discussed during the meeting. Challenges include achieving the required sample size to make prevalence classification. In particular, TDR survey implementation was challenging in countries with low prevalence generalized epidemics or epidemics concentrated in specific risk groups. The group discussed possible changes to survey methods including use of CD4 criteria to maximize sample enrichment with individuals likely to be recently infected. However, the sparse availability of CD4 testing and the spatial and temporal disconnection between CD4 testing and HIV testing require are simpler approach. A literature review to inform the revision of these criteria is currently being performed. Results of this review will be shared with the group. At present, no decisions have been taken to change TDR survey methods. 4.2 Public health actions based on HIVDR surveys result Moderate or high prevalence classifications of TDR or baseline HIVDR should trigger public health and programmatic actions. Those actions were discussed and the following consensus was achieved: Low level TDR (<5%) (Antenatal clinic (ANC), Voluntary counselling and testing/sexually transmitted infection clinic (VCT/STI), Most at risk population (MARPs)) Repeat survey in 2 years in same geographic area Consider expansion in 2 years to additional areas based on ART coverage and roll out No changes in current ARV guidelines prevention of mother to child transmission (PMTCT), ART, post exposure prophylaxis (PEP), pre-exposure prophylaxis ( PrEP) based on survey results Moderate Level 5-15% TDR in pregnant women (ANC) (NNRTI threshold): Following verification of moderate prevalence classification after quality assurance of laboratory and epidemiological data, a TDR survey should be repeated immediately to confirm moderate TDR in the same geographic area and in additional areas of the country. Depending on TDR results, the following actions should be taken: Review HIVDR EWI data, available VL suppression rates and acquired HIVDR surveillance data at ART sites in the area of TDR survey; Assess performance of HIV prevention programmes in minimizing transmission of drug resistant virus; 7

12 Bill and Melinda Gates Foundation 4 March 2012, Seattle, Assess coverage of HIV testing services (high coverage of HIV testing increases awareness of HIV status and reduce the risk of unintended HIVDR transmission); Perform HIVDR surveillance in populations prior to ART initiation (pre-therapy) to inform policy on: first- line regimens, use of pre-treatment HIVDR genotyping or use of intensified individual viral load monitoring Consider full scale national surveillance in ALL HIV-infected PREGNANT WOMEN to estimate a point prevalence of TDR and trigger programmatic and public health actions such as switch to PI-based PMTCT, or genotyping of HIV-infected pregnant women, based on national costeffectiveness thresholds. High Level >15% TDR in ANC sites (NNRTI threshold): verification of moderate prevalence classification after quality assurance of laboratory and epidemiological data, the following programmatic and publis health actions are recommended: Review HIVDR EWI data, available VL suppression rates and acquired HIVDR surveillance data at ART sites in the area of TDR survey; Assess performance of HIV prevention programmes in minimizing transmission of drug resistant virus; Assess coverage of HIV testing services (high coverage of HIV testing increases awareness of HIV status and reduce the risk of unintended HIVDR transmission); Perform HIVDR surveillance in populations prior to ART initiation (pre-therapy) to inform policy on: first- line regimens, use of pre-treatment HIVDR genotyping or use of intensified individual viral load monitoring Perform full scale national surveillance in ALL HIV-infected PREGNANT WOMEN to estimate point prevalence of TDR and trigger appropriate programmatic and public health actions such as switch to PI-based PMTCT, or genotyping of HIV-infected pregnant women, based on national cost-effectiveness thresholds. 4.3 Need for cost effectiveness analysis to identify appropriate thresholds to trigger public health actions: developing models to predict HIVDR impact (Andrew Philips) With the aim of informing the definition of appropriate HIVDR prevalence thresholds to trigger public health action a modeling exercise would be required. Potential factors to be included in the model were illustrated addressing the potential complexity of this approach. This work may also provide an essential preliminary piece to conduct a value of information analysis and evaluate whether HIVDR surveillance offer a financial benefit to program. However, the general consensus expressed by the group was that simpler and more effective approaches to advocate for HIVDR surveillance should be considered. 8

13 WHO ResNet Annual Meeting SESSION D WINDS OF CHANGE IN THE WHO HIVDR LABORATORY NETWORK (NEIL PARKIN) An update was given to the group regarding current activities, achievements and challenges encounter by the WHO HIVDR laboratory Network. The Laboratory at Division of Research on Virology and Immunology (DRVI), NCAIDS, Chinese Center for Disease Control and Prevention, Beijing China was recently accredited as a regional laboratory. In 2011, three new national drug resistance laboratories (NDRL) were added: Rio de Janeiro (Brazil); Abidjan (Cote d Ivoire); and Addis Ababa (Ethiopia). Seven additional laboratories are at various stages of the application process: Vietnam (2), Brazil (2), Uganda, Mexico, and the Russian Federation. In addition to performing laboratory testing for HIVDR surveillance, the laboratory network developed a dried blood spot (DBS) validation panel to assess DBS extraction methods of member laboratories. DBS storage and shipment conditions were assessed though operational research performed by WHO, the National Institutes of Health, and the WHO accredited laboratory in Uganda. Additionally, innovative lower cost genotyping assays have been explored and were discussed. Some challenges were reported. Of particular concern is difficulty in maintaining accreditation for laboratories using only plasma as a specimen type and the need to accelerate DBS-based genotyping despite its higher cost to facilitate representative results. Changes in the accreditation mechanism were proposed in order to tackle existing challenges and severely restricted funding availability. At present the NIH supported proficiency panel program is undergoing re-competition and will be re-funded. However, at present no new laboratories may receive proficiency panels. THE FUTURE OF HIVDR SURVEILLANCE These and further elements of the WHO HIVDR Laboratory Network were extensively discussed during the WHO ResNet Laboratory Meeting held the 5th of March 2012 in Seattle and a comprehensive report of the meeting is hereby presented in Annex 1. A number of positive signals related to HIVDR surveillance implementation have been noticed and a fair number of surveys have been already been conducted. However significant effort is required to expand and maintain surveillance coverage particularly in high priority countries. Similarly, additional commitment should be put in place to ensure TDR surveys are repeated following WHO guidance and to engender enthusiasm for use of these data for informed decision making. Scale-up of EWI monitoring to all ART sites should be encouraged and should be integrated as part of routine national components of monitoring and evaluation. Countries which have already participated in the global HIVDR surveillance and prevention strategy should be reenergized and more countries should become involved once data management capacity has been secured. 9

14 Bill and Melinda Gates Foundation 4 March 2012, Seattle, Issues around sustainability have been clarified, particularly in the current context of limited funding availability. Improvement in performance can be instrumental to advocate for sustained and better support by partners and member states. Meeting participants made suggestions for increased training between laboratories and collaborators in the network. A window of opportunity has to be created to move commitment from the technical level to the political level. This can be facilitated by strong communication. Specific suggestions were made by the participants; e.g. produce short briefs for public health decision makers regarding the value and importance of HIVDR surveillance and prevention efforts. Global HIVDR surveillance and monitoring will be highlighted in an upcoming WHO global report. A proposal to increase recognition of member institutions and reinforce mutual support within ResNet was briefly discussed. It was agreed that WHO will create a strategy /implementation working group including, in addition to ResNet members, public health decision makers to guide the future functioning of ResNet. Finally, there was clear consensus that WHO should continue to lead and provide coordination for the HIV ResNet group. 10

15 WHO ResNet Annual Meeting CONCLUSION AND NEXT STEPS The HIV ResNet group endorsed the interpretation of results presented as well as the new components of the global strategy. WHO was invited to continue providing global coordination and guidance. The individual commitment and collective responsibility of the ResNet members in advocating and support the global strategy activities were reinforced. Bilateral collaboration between member institution should be further explored in an effort to support sustainability The fast release of the new revised global strategy was seen as an important element to be used for advocacy to obtain support from the Global Fund and other funding agencies. Additionally, dissemination of information may also increase uptake of the strategy by countries and enhance motivation which have recently been adversely affected by the termination of funding. Prioritization of funding was also seen as a significant element which may help countries to implement one strategically selected assessment element of the HIVDR strategy if it were unable to commit to implementation of the entire recommended surveillance package. 11

16 Bill and Melinda Gates Foundation 4 March 2012, Seattle, ANNEX 1. WHO LABORATORY NETWORK MEETING REPORT Seattle, WA, 5 March 2012 BACKGROUND The global surveillance for HIV drug resistance (DR) has been spearheaded by the World Health Organization (WHO) and its partners, with contributions from an international network of epidemiologists, researchers, and laboratories. Financial support to WHO since 2007 has been primarily though a grant from the Bill and Melinda Gates Foundation (BMGF), and to CDC from the President s Emergency Program for AIDS Research (PEPFAR). In addition, the provision of materials and logistical support for the genotyping external quality assurance program, run by the Virology Quality Assurance (VQA) program at Rush University in Chicago, IL, is supported by a grant from the NIAID (NIH). In 2013, the WHO grant from BMGF will end, threatening the continuity of coordination in the Network. Furthermore, the capacity for VQA and NIH to support the WHO laboratory network EQA program has reached its capacity. In order for the laboratory operations component of the HIV DR surveillance program to continue, additional sources of support must be identified, and a critical review of the manner in which activities are carried out and of their efficiency is needed. The WHO HIVDR Laboratory Network Advisory Group assembled on March 5, 2012 at the offices of the Bill and Melinda Gates Foundation in Seattle, WA. Agenda topics, discussion and outcomes are summarized below. A list of participants and meeting agenda are also attached. 1. LABORATORY NETWORK STATUS AND SUSTAINABILITY CHALLENGES Laboratory accreditation status and activity update: During 2011, 3 new laboratories were added to the network. There are now 27 laboratories accredited and 7 more in various stages of the accreditation process. EQA program update: the VQA continues to support the WHO EQA program by providing proficiency panels, logistical support, distribution, and data processing functions. Recent results: panel 022g has been tested by 28 laboratories so far, with a total of 39 submissions (several labs use more than 1 assay type); 38 have met the criteria and mean nucleotide identity scores was 99.7%. VQA expansion limitations: due to the nature of the contract from the Division of AIDS (DAIDS) at NIH, no new labs can be added to the list of those receiving EQA panels. Future of the EQA program: A new contract award will be made by NIH/DAIDS in June 2012, and the new awardee will continue to provide the currently enrolled labs with proficiency panels. However, unless additional funding can be identified, additional labs cannot be added. Since passing the annual panel is a requirement for accreditation, this means that no new labs can be added to the network. At the same time, the demand for dried blood spot (DBS)-based 12

17 WHO ResNet Annual Meeting genotyping services in increasing, and some laboratories in the network are underutilized due to low demand in their regions and/or high cost of testing. WHO will review the activity of all laboratories since 2007 and those that have not contributed genotyping data for surveys or are unlikely to do so in the near future will be reclassified as Affiliated Laboratories and will not receive proficiency panels until further notice. These labs will not be able to test specimens for WHO surveys until a need is demonstrated and the lab tests and passes the annual panel. New DBS accreditation requirements: in order to increase DBS-based testing capacity, all Regional and Specialized laboratories will be required to have demonstrated proficiency with DBS. If they do not, they will not receive proficiency panels (plasma or DBS) until further notice. These labs will not be able to test specimens for WHO surveys until they meet the DBS requirements and successfully test the annual panels. The DBS accreditation requirements are as follows: 1. Existing accreditation by WHO for the performance of HIVDR genotyping from plasma. 2. At least 6 months of experience in DBS-based genotyping. 3. At least 100 DBS specimens successfully amplified and sequenced, with a success rate of at least 80% when viral loads are over 5000 copies/ml and DBS have been well handled (i.e. according to WHO recommendations for preparation, storage and shipping). 4. Successful testing of a WHO-recognized proficiency panel consisting of DBS specimens. 5. Successful validation of a DBS-based in-house assay for genotyping using the standardized criteria described separately. 6. Submission to WHO of a report summarizing the above elements for review. To summarize the proposed changes: 1. RDRL and SDRL are requested to submit DBS assay validation data by December WHO will perform a needs assessment and review lab activity since Labs with good performance with DBS and a record of previous contribution will continue to receive plasma and DBS panels in Other labs will not receive panels and will not be able to test specimens for WHO surveys. 5. Roles and responsibilities of excluded labs to be determined (training, operational research, etc.). 2. THE CEPHEID XPERT MTB/RIF ASSAY IN RESOURCE-LIMITED SETTINGS: POTENTIAL FOR APPLICATION TO HIVDR SURVEILLANCE Dr. David Persing (Chief Scientific Officer at Cepheid, Sunnyvale, CA) presented an overview of Cepheid s technology platform for molecular diagnostics. The Cepheid Xpert MTB/RIF nucleic amplification assay is based on real-time PCR and provides information on 5 different sites in the genome using molecular beacon detection technology. The MTB/RIF assay performs simultaneous detection of TB and rifampicin resistance directly from a sputum specimen, and was developed in partnership with the Foundation for Innovative New Diagnostics (FIND) and the University of Medicine and Dentistry of New Jersey, with support from the US National Institutes of Health. The MTB assay target is the 81 bp region (Rifampicin Resistance-Determining Region or RRDR) of the rpob gene. Molecular beacon probes have some advantages over linear probes, such as an increased number of possible fluorophores that can be used as 13

18 Bill and Melinda Gates Foundation 4 March 2012, Seattle, reporters and better ability to discriminate single base-pair mismatches. Five probes are used, each targeting a different area of the amplified target. Each probe is labeled with a different fluorophore, permitting simultaneous detection of the presence of wild type. The GeneXpert platform is a self-contained workstation that supports multiple clinical diagnostic assays. It has several configurations that allow for simultaneous loading of up to 80 cartridges, each designed for the performance of a specific test. The technology was endorsed by WHO in December FIND has negotiated preferential pricing for both the GeneXpert instrument and Xpert MTB/RIF cartridges for specific resource-limited countries. The current negotiated price of a GeneXpert System 4-module unit with desktop computer for the public sector in eligible countries is 17,000 USD; eligible countries can purchase test cartridges for approximately 17 USD each. As of the end of 2011, nearly 300 instruments have been ordered for use in 47 countries, 21 of which are also countries in which HIV DR surveillance is currently being carried out (Botswana, Brazil, Cambodia, Ethiopia, India, Indonesia, Kenya, Lesotho, Malawi, Mozambique, Nigeria, Russia, Senegal, South Africa, Swaziland, Tanzania, Thailand, Uganda, Zambia, Zimbabwe, and Viet Nam; more information can be found at en/index.html). Therefore it is of great interest to determine whether it is possible to develop an HIV DR genotyping assay that can be adapted to the GeneXpert platform. It is important to note that the instrumentation (systems and modules) required for the MTB/ RIF assay may not require that an HIV DR assay also use molecular beacons for detection and discrimination. For example, an AS-PCR or OLA assay might be used instead and adapted to run on the same system and/or module. Capitalizing on the international roll-out of the Gene X-pert instruments for TB testing in resource limited settings for HIV DR applications would require Cepheid to develop the HIV kits, since it is a closed system. Although Cepheid is interested in the possibility of developing such an assay, assigning company resources to the effort required is predicated on sufficient return on the investment by parallel marketing of the assay in resource-rich as well as resource-poor settings. Since it is not clear that a market exists for a Cepheid assay for HIV drug resistance mutations, at this time the company is not planning to initiate this development effort. If outside sources of funding could be identified, this decision could be revisited. 3. GENOTYPING COST REDUCTION Between 2007 and 2011, genotyping costs for specimens collected for WHO surveys in up to 30 countries in resource-limited settings were covered using funds from the Gates Foundation grant to WHO. The annual cost for 1 survey of transmitted resistance and 1 monitoring survey using either of the commercial kits (ViroSeq or TruGene) is approximately 75% of the budget allocated, which is also intended to cover the cost of viral load, shipping, training, and data management. Beginning in 2012, these funds are no longer widely available. The capacity of the small number of public health labs that can provide no cost genotyping has been reached. Therefore the cost of genotyping assays must be dramatically reduced if HIVDR testing is to continue. One approach to reducing genotyping assay cost is to switch to point mutation assays (PMA), since full PR-RT sequencing may not be required for public health surveillance purposes, and 14

19 WHO ResNet Annual Meeting an alternative assay that only interrogates a subset of DR-associated mutations might suffice. Such an assay might be less costly and simpler to perform. PMA technologies to consider include the Cepheid GeneXpert platform, allele-specific PCR (AS-PCR), Oligonucleotide Ligation assay (OLA), Allele Specific Primer Extension (ASPE), and others. Factors to consider include:»» Selection of which mutations to test; this is likely to change as treatment recommendations evolve.»» Impact on sensitivity to detect DR HIV, since some DR HIV will be misclassified as drug sensitive if they lack all the mutations being tested for.»» Sequence diversity between subtypes will likely necessitate regional differences in primer sequence and assay conditions»» Quality assurance (for example, the inability to perform phylogenetic analysis)»» Training, support and implementation»» Cost A PMA feasibility report is being prepared that will outline the salient technology platforms, model the effects of mutation selection on DR HIV detection sensitivity, and assess the likelihood that a PMA can replace full sequencing in a cost-effective manner. The general consensus of meeting participants was that outside of very localized areas where subtype diversity is minimal, it is unlikely that a single PMA could be developed that would be both able to test multiple subtypes and be cost-effective (i.e. less costly than current sequencing assays). Allele Specific Primer Extension (ASPE). The ASPE method involves two phases, a primerextension reaction, primed with oligonucleotides complementary to specific variants at the codons of interest, followed by capture on solid microsphere surfaces for detection. The microspheres have an oligo bound to them that is complementary to a capture sequence that is part of the allele specific oligonucleotide. In combination with an instrument capable of highorder multiplexing such as the Illumina MAGPIX, a large number of variants at each position of interest can be tested. The WHO Specialized DR Laboratory at the US CDC in Atlanta, GA had experimented with a prototype ASPE assay for HIV DR mutations in PR and RT. Forty five genotyping primers have been designed for twenty allele-specific mutations associated with HIVDR to NRTI, NNRTI and PI (M41L, K65R, K70R, L74V, Y115F, Q151M, M184V, K219E, L100I, K101E, K103N, V106A/M, Y181C, Y188L, G190A, V32I, I47V, L76V, I84V and L90M). A DNA fragment containing the target mutation sites is amplified, and amplicons are extended using the 45 primers in a single reaction tube. Sensitivity for detection of mutant variants in mixtures is about 7%. As an alternative to PMAs, it was proposed that the network adopt a lower cost version of current sequencing assays. For example amplification and sequencing of RT only, perhaps with as few as 2 primers and allowing for some regions of single-stranded coverage, and reduced manual editing of basecalls (see below) could be performed for $50 USD per test or less. Some labs are running such low cost assays in their labs and can be a resource for protocol sharing and training to other labs. 15

20 Bill and Melinda Gates Foundation 4 March 2012, Seattle, Dr. Richard Harrigan (University of British Columbia, Vancouver, Canada) has offered to provide DNA sequencing at no cost to WHO network labs in need. Furthermore Dr. Harrigan has developed an automated software system (called RECall) that accepts raw chromatogram data from ABI sequencers, and generates Individual nucleotide sequence text files, batch nucleotide sequences in fasta format, PR-RT resistance assessment (Stanford algorithm), and batch summaries with quality indicators, mutations list, and relative peak heights at mixed positions. See and Woods et al., J. Clin. Microbiol. June 2012 vol. 50 no for more information. In the second half of 2012, WHO staff and consultants will explore ways to make the best use of these resources to maintain the current level of laboratory support for the global surveillance of HIVDR. 16

21 WHO ResNet Annual Meeting ANNEX 2. LIST OF PARTICIPANTS AFRO Tendani Gaolathe Botswana Harvard AIDS Initiative Partnership Princess Marina Hospital Gaborone Botswana Nicaise Ndembi Institute of Human Virology Pent House, Maina Court Abuja, Nigeria Lynn Morris AIDS Virus Research Unit, Natl. Inst. for Communicable Diseases Johannesburg, South Africa Gillian Hunt National Institute for Communicable Diseases (NICD) Sandringham Johannesburg, South Africa AMRO Amilcar Tanuri UFRJ Rio de Janeiro, Brazil Mark Wainberg McGill University Montreal, Quebec Canada Richard Harrigan BC Centre for Excellence in HIV/AIDS University of British Columbia Vancouver, BC Canada Paul Sandstrom National HIV and Retrovirology Laboratories Public Helath Agency of Canada Canada Surbhi Modi Pediatric HIV Team Maternal and Child Health Branch CDC Atlanta, GA Chunfu Yang International Laboratory Branch, GAP, NCHSTP, CDC Atlanta, GA Elliot Raizes CDC Atlanta, GA Margaret Davis Maternal and Child Health Branch Global AIDS Program, CDC Atlanta, GA Michael R. Jordan Tufts University Boston, Steven Y. Hong Tufts Medical Center and Tufts School Of Medicine Boston, Neil Parkin Data First Consulting, Inc. Belmont, CA 17

22 Bill and Melinda Gates Foundation 4 March 2012, Seattle, Christine Rousseau Bill and Melinda Gates Foundation Seattle, WA Chris Duncan Bill and Melinda Gates Foundation Seattle, WA Vincent Marconi Grady Health System s Infectious Disease Program at the Ponce de Leon Center and Emory School of Public Health Atlanta, GA Deborah Persaud John Hopkins School of Medicine Baltimore, MD Jon Kaplan Centers for Disease Control and Prevention (CDC) 1600 Cliffon Road NE Atlanta, GA Diane E. Bennett ESIB, Division of Global HIV/AIDS Center for Global Health Centers for Disease Control and Prevention MS E-30, 1600 Clifton Road NE Atlanta, GA Lynne Mofenson 6100 Executive Blvd Room 4B11E, MSC 7510 Bethesda Md Soo-Yon Rhee Director, Stanford HIV Drug Resistance Database Standford University California EURO Anne-Mieke Vandamme The AIDS Reference Laboratory Rega Institute and University Hospitals Minderbroedersstraat Leuven Belgium Charles Boucher Department of Virology Erasmus Medical Center, Erasmus University Rotterdam, The Netherlands Jens Lundgren Department of International Health, Immunology and Microbiology Panum Instituttet, Blegdamsvej, København N Denmark Jonathan M. Schapiro Director, AIDS Service National Hemophilia Center Tel Hashomer Israel Raph Hamers PharmAccess Foundation Academic Medical Center of the University of Amsterdam Amsterdam The Netherlands Kim Sigaloff PhD student, PharmAccess Foundation and Department of Global Health, Academic Medical Center, University of Amsterdam The Netherlands Rob Schuurman Department of Virology University Medical Center Utrecht Utrecht The Netherlands David van de Vijver Department of Virology University Medical Center Utrecht The Netherlands 18

23 WHO ResNet Annual Meeting Dr AM.J. Wensing Department of Virology University Medical Center Utrecht Utrecht, The Netherlands Roger Paredes Institut de Recerca de la SIDA - irsicaixa Badalona, Spain Deenan Pillay Health Protection Agency London, UK Daniel Davis University of Cambridge School of Clinical Medicine Cambridge, UK Ravindra Gupta 43 Derby Road London -E182PY UK Professor Clive Loveday International Clinical Virology Centre Trust (ICVC Charitable Trust) Great Missenden, UK Andrew Phillips HIV Epidemiology and Biostatistics Group Research Department of Infectious and Population Health UCL-Royal Free Campus London, UK SEARO Srikanth Tripathy National AIDS Research Institute Indian Council of Medical Research Bhosari, Pune India WPRO Anna Hearps Clinical Research Laboratory Burnet Inst. for Medical Research and Public Health Melbourne, Australia James McMahon Infectious Diseases Unit Alfred Hospital Level 2 Burnet Tower 85 Commercial Road Melbourne Australia Annette Sohn TREAT Asia/amfAR The Foundation for AIDS Research Sukhumvit Road, Suite 2104 Klongtoey, Bangkok Thailand WHO SECRETARIAT Joseph Perriëns HIV Technologies and Commodities Unit Department of HIV/AIDS World Health Organization Geneva, Switzerland Silvia Bertagnolio HIV Technologies and Commodities Department of HIV/AIDS WHO, Geneva, Switzerland Martina Penazzato Consultant HIV Technologies and Commodities Department of HIV/AIDS WHO, Geneva, Switzerland Philippa Easterbrook Consultant HIV Treatment and Care Department of HIV/AIDS WHO,Geneva, Switzerland 19

24 Bill and Melinda Gates Foundation 4 March 2012, Seattle, ANNEX 3. AGENDA WHO HIV ResNet Annual Meeting 4 March 2012 Seattle, Agenda Chair: Mark Wainberg Session topic / Presenter 09:00 09:10 Welcome and Introductions (Jos Perriens, Coordinator of the Technologies and Commodities (TCO) Team) 09:10 10:30 Implementation, Results and Interpretation of HIVDR WHO recommended surveys 1. Monitor HIVDR Early Warning Indicators from ART clinics 2. Surveillance of acquired HIVDR 3. Surveillance of transmitted HIVDR 10:30 11:00 Coffee break 11:00 12:30 Contextualization WHO HIVDR Data in the existing published literature: 4. HIVDR in recently and chronically infected populations : A Meta-analysis (Ravi Gupta, UCL, London, UK) 5. Acquired HIVDR: A Meta-analysis (Daniel Davis, Cambridge, UK) 12:30 13:30 Lunch 13:30 14:45 WHO HIVDR Prevention and Assessment Strategy -- New Directions 6. Updated HIVDR EWIs guidance 7. Surveillance of acquired HIVDR in populations failing ART- A cross sectional method 8. Surveillance of HIVDR in populations initiating ART 9. Surveillance of HIVDR in HIV-infected and ART naive infants < 18 months 10. Feedback from Expert Meeting (Geneva, Feb 14-15, 2012) Revision of epidemiological criteria for definition of recently infection Public health actions based on HIVDR surveys result - Need for cost effectiveness analysis to identify appropriate thresholds to trigger actions - Need for value of information analysis - Moving forward: developing models to predict HIVDR impact (Andrew Philips) 14:45 15:15 Discussion 15:15 15:30 Coffee break 15:30 15:45 Winds of change in Laboratory network 15:45 16:15 Next steps and future prospective (Jos Perriens, WHO-Geneva) 20

25 NOTES

26 NOTES

27 For more information, contact: World Health Organization Department of HIV/AIDS 20, avenue Appia 1211 Geneva 27 Switzerland ISBN hiv-aids@who.int