P15-13. HVTN 503(Phambili) trial discontinuation of enrolment/vaccination: the impact on trial participant attitudes to vaccine trials and scientific research

Background: The HVTN 503/Phambili trial, a phase IIB study of the Merck Ad-5 HIV vaccine, suspended enrollment and vaccinations following results of the HVTN502/STEP study. Participants were notified of their treatment allocation and continue to be followed. We investigated the impact of the suspension on trial participants' attitudes towards HIV vaccine trials and scientific research. Methods: A self-administered questionnaire investigating knowledge and attitudes utilising a five-point likert scale, was administered 12–18 months post suspension at the 4 Phambili trial sites implementing the latest protocol. Results: Of 544 participants enrolled at 4 sites, 490 completed the questionnaire. 51% understood that vaccination, and 31% that enrolment had been permanently discontinued. Although 85% knew Phambili DSMB recommendations were based on results of the STEP study, only 51% knew that the results showed more infections in the vaccine arm. Although 90% knew they should continue protecting themselves against HIV, only 33% acknowledged trialists' concerns that the vaccine may make them more susceptible to HIV. Knowledge was similar across sites. 12% had tested for HIV antibodies outside of the trial before unblinding to establish their treatment allocation. 46% felt disappointed, angry or afraid, primarily because the vaccine would not prevent HIV in their community. 79% said they would engage in less risk behaviour. 94% thought the study team had made the correct decision by unblinding them and 96% thought site-participant communication was appropriate. 91% felt they made the right decision to enrol in Phambili and 91% planned to continue attending scheduled visits. 86% support scientific research and 80% would want to participate in future HIV vaccine. Conclusion: Despite site communications, knowledge levels were low. Importantly, most knew they should continue to protect themselves against HIV. Although almost half of participants reported negative feelings, most attitudes towards trial sites, scientific research and HIV vaccine trials were positive

P14-07. Offering new prevention modalities in HIV vaccine trials: experience with male circumcision in the Phambili trial

Background: New prevention options will be added to the 'standard of prevention' offered in HIV vaccine efficacy trials as new methods prove effective. The HVTN503/Phambili trial was initiated in January 2007, shortly after results from 3 randomized controlled trials of male circumcision (MC) demonstrated that MC reduces the risk of HIV acquisition. Thus, HVTN503 investigators made plans to offer MC at no cost to enrolled male participants. Methods: All participants were informed of the benefits of MC as a component of HIV risk reduction counseling, including how and where MC could be accessed. One site offered on-site MC and the others provided referral to local services for men who requested MC. We present data on uptake of MC post-enrollment. Results: Prior to discontinuation of enrolment, 441 men joined the trial, of whom 312 (70.7%) were uncircumcised. Of these, 82 (26.3% of uncircumcised men) requested MC after enrolment. Uptake varied by site, being lower (70%) at the eThekwini site, the site with lowest baseline MC prevalence. Among 3 sites with intermediate baseline MC prevalence, uptake varied from 3.3 to 37.6%, being highest at the site providing MC on site. Uptake was similar in vaccine and placebo arms of the trial [42 (26.1%) vs. 40 (26.5%)]. There was no significant difference by arm in the timing of circumcision relative to randomization assignment being provided to participants following release of the STEP trial results [post-unblinding, vaccine 18 (42.9%) vs. placebo 13 (32.5%), p = 0.37]. Conclusion: MC, a new prevention modality, was offered as part of HIV prevention services in HVTN503. Uptake varied by provision of care model and inversely with baseline MC prevalence, but did not differ between treatment arms, and remained similar even after provision of treatment

Drug resistance to sulphadoxine-pyrimethamine in Plasmodium falciparum malaria in Mlimba, Tanzania

BACKGROUND: Sulphadoxine-pyrimethamine (SP) has been and is currently used for treatment of uncomplicated Plasmodium falciparum malaria in many African countries. Nevertheless, the response of parasites to SP treatment has shown significant variation between individuals. METHODS: The genes for dihydrofolate reductase (dhfr) and dihydropteroate synthase (dhps) were used as markers, to investigate parasite resistance to SP in 141 children aged less than 5 years. Parasite DNA was extracted by Chelex method from blood samples collected and preserved on filter papers. Subsequently, polymerase chain reaction (PCR) and restriction fragment length polymorphism (PCR-RFLP) were applied to detect the SP resistance-associated point mutations on dhfr and dhps. Commonly reported point mutations at codons 51, 59, 108 and 164 in the dhfr and codons 437, 540 and 581 in the dhps domains were examined. RESULTS: Children infected with parasites harbouring a range of single to quintuple dhfr/dhps mutations were erratically cured with SP. However, the quintuple dhfr/dhps mutant genotypes were mostly associated with treatment failures. High proportion of SP resistance-associated point mutations was detected in this study but the adequate clinical response (89.4%) observed clinically at day 14 of follow up reflects the role of semi-immunity protection and parasite clearance in the population. CONCLUSION: In monitoring drug resistance to SP, concurrent studies on possible confounding factors pertaining to development of resistance in falciparum malaria should be considered. The SP resistance potential detected in this study, cautions on its useful therapeutic life as an interim first-line drug against malaria in Tanzania and other malaria-endemic countries

Molecular surveillance of Plasmodium vivax dhfr and dhps mutations in isolates from Afghanistan

<p>Abstract</p> <p>Background</p> <p>Analysis of dihydrofolate reductase (<it>dhfr</it>) and dihydropteroate synthase (<it>dhps</it>) mutations in <it>Plasmodium vivax </it>wild isolates has been considered to be a valuable molecular approach for mapping resistance to sulphadoxine-pyrimethamine (SP). The present study investigates the frequency of SNPs-haplotypes in the <it>dhfr </it>and <it>dhps </it>genes in <it>P. vivax </it>clinical isolates circulating in two malaria endemic areas in Afghanistan.</p> <p>Methods</p> <p><it>P. vivax </it>clinical isolates (n = 171) were collected in two different malaria endemic regions in north-west (Herat) and east (Nangarhar) Afghanistan in 2008. All collected isolates were analysed for SNP-haplotypes at positions 13, 33, 57, 58, 61, 117 and 173 of the <it>pvdhfr </it>and 383 and 553 of the <it>pvdhps </it>genes using PCR-RFLP methods.</p> <p>Results</p> <p>All 171 examined isolates were found to carry wild-type amino acids at positions 13, 33, 57, 61 and 173, while 58R and 117N mutations were detected among 4.1% and 12.3% of Afghan isolates, respectively. Based on the size polymorphism of <it>pvdhfr </it>genes at repeat region, type B was the most prevalent variant among Herat (86%) and Nangarhar (88.4%) isolates. Mixed genotype infections (type A/B and A/B/C) were detected in only 2.3% (2/86) of Herat and 1.2% (1/86) of Nangarhar isolates, respectively. The combination of <it>pvdhfr </it>and <it>pvdhps </it>haplotypes among all 171 samples demonstrated six distinct haplotypes. The two most prevalent haplotypes among all examined samples were wild-type (86%) and single mutant haplotype I₁₃P₃₃F₅₇S₅₈T₆₁<b>N </b>₁₁₇I_173/A₃₈₃A₅₅₃(6.4%).</p> <p>Double (I₁₃P₃₃S₅₇<b>R</b>₅₈T₆₁<b>N</b>₁₁₇I₁₇₃/A₃₈₃A₅₅₃) and triple mutant haplotypes (I₁₃P₃₃S₅₇<b>R </b>₅₈T₆₁<b>N</b>₁₁₇I₁₇₃/<b>G</b>₃₈₃A₅₅₃) were found in 1.7% and 1.2% of Afghan isolates, respectively. This triple mutant haplotype was only detected in isolates from Herat, but in none of the Nangarhar isolates.</p> <p>Conclusion</p> <p>The present study shows a limited polymorphism in <it>pvdhfr </it>from Afghan isolates and provides important basic information to establish an epidemiological map of drug-resistant vivax malaria, and updating guidelines for anti-malarial policy in Afghanistan. The continuous usage of SP as first-line anti-malarial drug in Afghanistan might increase the risk of mutations in the <it>dhfr </it>and <it>dhps </it>genes in both <it>P. vivax </it>and <it>Plasmodium falciparum </it>isolates, which may lead to a complete SP resistance in the near future in this region. Therefore, continuous surveillance of <it>P. vivax </it>and <it>P. falciparum </it>molecular markers are needed to monitor the development of resistance to SP in the region.</p

Molecular markers of anti-malarial drug resistance in Lahj Governorate, Yemen: baseline data and implications

<p>Abstract</p> <p>Background</p> <p>This is an investigation of anti-malarial molecular markers coupled with a therapeutic efficacy test of chloroquine (CQ) against falciparum malaria in an area of unstable malaria in Lahj Governorate, Yemen. The study was aimed at assessment of therapeutic response to CQ and elucidation of baseline information on molecular markers for <it>Plasmodium falciparum </it>resistance against CQ and sulphadoxine/pyrimethamine (SP).</p> <p>Methods</p> <p>Between 2002 and 2003 the field test was conducted according to the standard WHO protocol to evaluate the therapeutic efficacy of CQ in 124 patients with falciparum malaria in an endemic area in Lahj Governorate in Yemen. Blood samples collected during this study were analysed for <it>P. falciparum </it>chloroquine resistance transporter gene (<it>pfcrt</it>)-76 polymorphisms, mutation <it>pfcrt-</it>S163R and the antifolate resistance-associated mutations dihydrofolate reductase (<it>dhfr</it>)-C59R and dihydropteroate synthase (<it>dhps</it>)-K540E. Direct DNA sequencing of the <it>pfcrt </it>gene from three representative field samples was carried out after DNA amplification of the 13 exons of the <it>pfcrt </it>gene.</p> <p>Results</p> <p>Treatment failure was detected in 61% of the 122 cases that completed the 14-day follow-up. The prevalence of mutant <it>pfcrt </it>T76 was 98% in 112 amplified pre-treatment samples. The presence of <it>pfcrt </it>T76 was poorly predictive of <it>in vivo </it>CQ resistance (PPV = 61.8%, 95% CI = 52.7-70.9). The prevalence of <it>dhfr </it>Arg-59 mutation in 99 amplified samples was 5%, while the <it>dhps </it>Glu-540 was not detected in any of 119 amplified samples. Sequencing the <it>pfcrt </it>gene confirmed that Yemeni CQ resistant <it>P. falciparum </it>carry the old world (Asian and African) CQ resistant haplotype CVIETSESI at positions 72,73,74,75,76,220,271, 326 and 371.</p> <p>Conclusion</p> <p>This is the first study to report baseline information on the characteristics and implications of anti-malarial drug resistance markers in Yemen. It is also the first report of the haplotype associated with CQR <it>P. falciparum </it>parasites from Yemen. Mutant <it>pfcrt</it>T76 is highly prevalent but it is a poor predictor of treatment failure in the study population. The prevalence of mutation <it>dhfr</it>Arg59 is suggestive of emerging resistance to SP, which is currently a component of the recommended combination treatment of falciparum malaria in Yemen. More studies on these markers are recommended for surveillance of resistance in the study area.</p

Molecular epidemiology of drug-resistant malaria in western Kenya highlands

<p>Abstract</p> <p>Background</p> <p>Since the late 1980s a series of malaria epidemics has occurred in western Kenya highlands. Among the possible factors that may contribute to the highland malaria epidemics, parasite resistance to antimalarials has not been well investigated.</p> <p>Methods</p> <p>Using parasites from highland and lowland areas of western Kenya, we examined key mutations associated with <it>Plasmodium falciparum </it>resistance to sulfadoxine – pyrimethamine and chloroquine, including dihydrofolate reductase (<it>pfdhfr</it>) and dihydropteroate synthetase (<it>pfdhps</it>), chloroquine resistance transporter gene (<it>pfcrt</it>), and multi-drug resistance gene 1 (<it>pfmdr1</it>).</p> <p>Results</p> <p>We found that >70% of samples harbored 76T <it>pfcrt </it>mutations and over 80% of samples harbored quintuple mutations (51I/59R/108N <it>pfdhfr </it>and 437G/540E <it>pfdhps</it>) in both highland and lowland samples. Further, we did not detect significant difference in the frequencies of these mutations between symptomatic and asymptomatic malaria volunteers, and between highland and lowland samples.</p> <p>Conclusion</p> <p>These findings suggest that drug resistance of malaria parasites in the highlands could be contributed by the mutations and their high frequencies as found in the lowland. The results are discussed in terms of the role of drug resistance as a driving force for malaria outbreaks in the highlands.</p