HIV-1 Strain and Primary Drug Resistance in Canada
Surveillance Report to June 30, 2002

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An Overview of the Canadian HIV Strain and Drug Resistance Surveillance Program

The Canadian HIV Strain and Drug Resistance Surveillance Program (CHSDRSP) is a collaborative effort between the provinces and territories and the Centre for Infectious Disease Prevention and Control (CIDPC), Public Health Agency of Canada, Health Canada. It forms a key component in a national system for the enhanced and integrated surveillance of HIV/AIDS, STD, emerging retroviruses and other sexually transmitted blood-borne pathogens. The program was initiated to characterize and monitor the genetic diversity of the HIV epidemic in Canada, addressing the concerns of affected communities, public health authorities, primary care physicians and researchers.

Goals of the CHSDRSP

A 1998 consensus workshop in Vancouver established the following goals for this program:

To enhance the safety of the blood supply

In order to ensure the safety of the blood supply, all HIV tests need to reliably detect the strains circulating in the country. The precedent for this goal was the discovery of HIV-2 and highly divergent group O strains of HIV-1, which required modification of some serologic screening tests by the addition of new antigens to ensure detection. The sentinel arm of the CHSDRSP, through the reference services of the national HIV laboratories, addresses this goal by testing samples with unusual serologic, polymerase chain reaction (PCR) or other virologic test results. This relation between the national HIV laboratories and the provincial public health laboratories also serves other programs, including quality assurance and the monitoring of diagnostic kits.

To inform vaccine development

One of the primary, public health related reasons for conducting a systematic surveillance of HIV genetic variability in Canada is to inform vaccine research and development. The majority of vaccines undergoing clinical trials have been developed using strains of HIV-1 subtype B (predominant in North America and Europe). However, the ability of these vaccines to elicit cross-subtype responses is unclear. It is therefore imperative to collect data on the prevalence and incidence of HIV subtypes and other determinants of subtype diversity in Canada in order to inform vaccine strategies, since the efficacy and effectiveness of vaccines may be subtype specific.

To assess genetic markers of HIV drug resistance

Although antiretroviral therapies have led to a reduction in HIV related morbidity and mortality in Canada, there is concern that their widespread use, increased number of treatment failures, and high HIV infection rates may result in increased transmission of drug-resistant virus. Indeed, studies have shown that transmission of drug-resistant virus is increasing. The CHSDRSP aims to address the prevalence of primary drug resistance and the variation of this prevalence over time, geographic area and population risk group. The resulting information can be used to develop treatment guidelines at the population level for initial therapeutic regimens and to develop more effective HIV prevention strategies, including the prevention of mother-to-child transmission.

To determine HIV transmission, pathogenesis and progression to HIV-related diseases

Although genetic analyses have been used to assess the spread of HIV globally, there is little consensus on whether differences in HIV subtype and mutations conferring drug resistance affect the rate of transmission, pathogenesis and HIV-related disease progression. Knowing the distribution of HIV variants in Canada, along with corresponding epidemiologic information, will help to address these questions. The public health implications of such findings, including prevention and treatment strategies, are of special interest.

Methodology

Epidemiologic data and laboratory specimen collection and transfer

The provincial partners in the CHSDRSP send serum samples taken for diagnostic testing from treatment naVve individuals with newly diagnosed HIV infection to CIDPC, Health Canada. Subtype analysis and primary drug resistance genotyping is conducted at the National Laboratory for HIV Genetics. Incidence testing is conducted at the National HIV Reference Laboratory. Samples with unusual laboratory results are sent through the sentinel arm of the CHSDRSP and are analyzed for subtype and drug resistance (if this testing is requested) at the National Laboratory for HIV Genetics.

For each submitted laboratory sample, non-nominal epidemiologic information is also sent to CIDPC. The data include information routinely collected on the national or provincial HIV case reporting forms and, where available, additional information that helps interpret the laboratory results, including treatment history, CD4 count and viral load at diagnosis, and previous HIV testing history. Epidemiologic analyses are conducted at the Division of HIV/AIDS Epidemiology and Surveillance.

As of June 30, 2002, British Columbia (BC), Alberta, Manitoba, Saskatchewan, Newfoundland and Labrador, and Nova Scotia have participated in CHSDRSP. The results presented in this report represent samples that CIDPC had received as of June 30, 2002, on which HIV subtype and drug resistance testing had been completed successfully. Serum samples have also been received from Ontario, but since these samples were received through the sentinel arm of the CHSDRSP the results are reported in Section 5, under National HIV Reference Services.

Samples and epidemiologic data continue to flow to Health Canada from participating provinces, and results from these analyses will be presented in future reports. Discussions are currently under way to expand collection of samples and epidemiologic data to the remaining provinces and territories.

Genetic algorithm for HIV subtyping and drug resistance testing

After extraction of the RNA and an initial one-step reverse transcriptase PCR, nested PCR amplification of the pol gene (which encodes for the HIV protease and reverse transcriptase enzymes) is performed using a combination of published and in-house group M consensus primers. The PCR product is directly sequenced with internal PCR primers on a Li-Cor 4200L automated sequencer. In this way, a complete double stranded sequence for the entire protease gene and the first 253 amino acids of reverse transcriptase is used to assess subtype and primary mutations associated with drug resistance. Of note is that between July 1998 and December 2000, the C2-V5 region (233 amino acids) of the envelope protein was used to assess HIV subtype. When PCR products sequence poorly, at least two additional amplification and sequencing attempts are made with increasing enhancement algorithms to sequence the PCR product. If this is not successful, a final effort is made to resolve the problem by cloning the PCR product and screening 10 to 12 clones per person.

Consensus of major mutations associated with primary drug resistance

Interpretation of results from genetic algorithms requires knowledge of the association between specific mutations and virologic response to antiretroviral drugs. The associations are often complex and not necessarily additive. Consensus drug resistance mutation lists have been published through database banks (e.g. Stanford University, http://hivdb.stanford.edu/hiv and the Los Alamos HIV Sequence Database, http://resdb.lanl.gov/Resist_DB/) and by expert committees on HIV drug resistance (e.g. International AIDS Society - USA Drug Resistance Mutations Group). However, even the experts do not always agree on these so-called “rules-based” algorithms.

For this report, major mutations identified in the protease and reverse transcriptase genes of HIV were defined by a consensus of listings reported by the International AIDS Society-USA Drug Resistance Mutations Group^* and by Stanford University. The major mutations that have been added to our consensus list since the last surveillance report of samples received to June 30, 2001, include I50L, D67N, Y115F, L210W, K219E/Q, L100I, V108I, Y188C/H/L, P225H, M230L and P236L. Please refer to Appendix 2 for the complete, current list of mutations associated with clinical resistance that was used for this report.

Epidemiologic analyses

Laboratory and epidemiologic data are linked using unique identifiers. Significant associations between primary drug resistance or HIV-1 non-B subtypes and epidemiologic characteristics of individuals in the sample population were assessed using the c² test and where appropriate Fisher's exact test. Logistic regression analyses to further define independent factors associated with primary drug resistance and with non-B infections were conducted using SPSS 8.0^TM (SPSS Inc. Chicago, IL). Independent variables that were examined included age at diagnosis of HIV infection, gender, exposure category, ethnicity, and year of diagnosis of HIV infection.

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