Correspondence
Proportion of viable parasites (%)
100
10
1
0·1 Wild-type Arg539Thr Cys580Tyr Tyr493His
0·01
0·001 Fast
Slow Parasite clearance
and slow-clearing parasites with concordant survival rates had wildtype or mutant alleles (Tyr493His and Cys580Tyr), respectively. These data suggest that parasite survival rates in the ring-stage survival assay are more relevant than parasite clearance half-lives for identifying artemisinin-resistant P falciparum, and further validate K13-propeller polymorphisms as a molecular marker of artemisinin resistance in vitro and in vivo. We declare no competing interests.
Figure: Association between parasite survival according to the in-vitro ring-stage survival assay (0–3 h) and parasite clearance in patients treated with artemisinin-based combination therapy, by K13-propeller genotype
(5·30%, 19·32%, and 51·39%) but a resistant stage-dependent pattern (decrease in survival as parasites matured of 1·2%, 17·3%, and 50·2%, respectively). We postulated that these three parasites were indeed artemisinin-resistant, but that their clearance was fast because circulating parasites seen in blood films were predominantly older, drug-sensitive rings at the time the patients received artemisinin drugs. Of the 13 slow-clearing parasites, one had a paradoxically low survival (0·16%). We believe that this parasite was in fact artemisinin-sensitive, but that its clearance was slow because the patient had low levels of parasite-clearing immunity2 or a poor pharmacokinetics profile. We reported K13-propeller polymorphism as a new molecular marker for artemisinin-resistant P falciparum malaria,3 and postulated that K13propeller genotypes would definitively resolve the four discordant results. To test this possibility we genotyped all 26 parasites and found that the three parasites with discordantly high survival rates each carry a different mutant K13-propeller allele (Tyr493His, Cys580Tyr, and Arg539Thr), and that the parasite with a discordantly low survival rate carried a wildtype allele (figure). As expected, we also found that all the fast-clearing 450
Chanaki Amaratunga, Benoit Witkowski, Nimol Khim, Didier Menard, *Rick M Fairhurst [email protected] Laboratory of Malaria and Vector Research, NIAID/National Institutes of Health, Rockville, MD 20852, USA (CA, RMF); and Malaria Molecular Epidemiology Unit, Institut Pasteur du Cambodge, Phnom Penh, Cambodia (BW, NK, DM) 1
2
3
Witkowski B, Amaratunga C, Khim N, et al. Novel phenotypic assays for the detection of artemisinin-resistant Plasmodium falciparum malaria in Cambodia: in-vitro and ex-vivo drug-response studies. Lancet Infect Dis 2013; 13: 1043–49. Lopera-Mesa TM, Doumbia S, Chiang S, et al. Plasmodium falciparum clearance rates in response to artesunate in Malian children with malaria: effect of acquired immunity. J Infect Dis 2013; 207: 1655–63. Ariey F, Witkowski B, Amaratunga C, et al. A molecular marker of artemisinin-resistant Plasmodium falciparum malaria. Nature 2014; 505: 50–55.
monotherapy with artemisinin drugs for treatment of falciparum malaria— particularly the chloroquine-resistant strain—is 5–7 days.2 A study of patients treated with dihydroartemisinin (60 mg) daily (120 mg on day 1) for 3, 5, or 7 days showed no recrudescences in the 7 day group, compared with recrudescence in 66% of patients in the 5 day group and 52% in the 3 day group. Therefore, it was recommended that the 7 day course, with a total dose of 480 mg of dihydroartemisin, is the optimum regimen for treatment of P falciparum infection.3 Another feature of clinical malaria management in China is the quarantine of patients during treatment to prevent transmission and hamper the spread of resistance. Combination of artemisinin drugs with other antimalarial treatments gives more effective treatment at the early stage of malaria infections, although, it is becoming clear that the number of treatment days has a major impact on efficacy of artemisinin treatment either as monotherapy or in combination, especially if ineffective partner drugs are used.4 Although we do not have evidence to demonstrate this hypothesis so far, increasing the duration of artemisinin treatment and use of efficacious partner drugs could sustain the efficacy of artemisinin combination therapies. We declare no competing interests.
Benoit Witkowski and colleagues1 reported a simple and rapid in vitro method to detect artemisinin resistance of P falciparum. They show that the slow clearance of parasites by artemisinin combination therapies is a result of a drop in susceptibility among young ring form parasites whereas susceptibility is maintained in mature stages. However, the clinical relevance of this finding to treatment failure is unclear since these infections could still be successfully treated with artemisinin combination therapies. Experiences in China also suggest that such resistance could be overcome if the right strategies are used. In China, the recommended duration of
*Zhao-Rong Lun, Pedro E Ferreira, Lin-Chun Fu [email protected] Key Laboratory of Gene Engineering of Ministry of Education, State Key Laboratory of Biocontrol, School of Life Sciences, Key Laboratory of Tropical Disease Control, Zhongshan School of Medicine, Sun Yat-Sen University, Guangzhou 510275, China (ZRL); School of Biological Sciences, Nanyang Technological University, Singapore (PEF); and Tropical Medicine Institute, Guangzhou University of Chinese Medicine, Guangzhou, China (LCF) 1
2
Witkowski B, Amaratunga C, Khim N, et al. Novel phenotypic assays for the detection of artemisinin-resistant Plasmodium falciparum malaria in Cambodia: in-vitro and ex-vivo drug-response studies. Lancet Infect Dis 2013; 13: 1043–49. The Ministry of Health of China. National guideline for use and regimen of antimalarials. Chin J Parasit Dis Control 2002; 15: 129–30.
www.thelancet.com/infection Vol 14 June 2014
Correspondence
3
4
Fu LC, Guo XB, Wang WL, et al. An exploration on the dosage and treatment course of dihydroartemisinin for falciparum malaria. Trad Chin Drug Res Clin Pharm 1996; 7: 17–18. Ferreira PE, Culleton R, Gil JP, Meshnick SR. Artemisinin resistance in Plasmodium falciparum: what is it really? Trends Parasitol 2013; 29: 318–20.
Stratified infection medicine: a call to arms Human susceptibility to infectious disease is strongly heritable.1 In the past decade, genetic studies have found more than 30 common genetic variants that have profound effects on susceptibility to infection, course of disease, and response to treatment.2 A major challenge for infectious disease medicine in the next decade will be making use of this knowledge to benefit patients. Stratification of patients, by customising management according to their genotypes, makes little impact on clinical treatment of infectious disease. We are aware of only one example—hepatitis C virus infection— in which this new paradigm has been exploited. In infection with hepatitis C virus genotype 1, a single nucleotide polymorphism in the IFNL3 gene is associated with a sustained virological response to treatment, and this can be clinically tested to predict response to therapy.3 By contrast, oncologists successfully and routinely apply knowledge of both patient’s and tumour’s genotypes to allocate patients to ever-smaller groups for whom optimum therapeutic strategies have been found. For example, HER2positive breast cancer responds to trastuzumab, and lung cancers carrying EGFR mutations respond to oral EGFR tyrosine kinase inhibitors. Several single nucleotide polymorphisms associated with infectious disease phenotypes could have implications for clinical practice.
www.thelancet.com/infection Vol 14 June 2014
The IFNL3 mutation associated with hepatitis C virus is also associated with the severity and frequency of reactivation of orofacial herpes simplex virus type 1 infection,4 which highlights two major concepts: first, that there are genetic variants that confer susceptibility to multiple pathogens; and second, that there are likely to be genotype-determined comorbidities that have not yet been observed and reported. Influenza A virus is a pathogen of global importance and a genetic polymorphism in IFITM3 is strongly associated with risk of severe influenza in human beings.5 If the clinical impact of this mutation can be quantified, and genotyping can be made both rapid and cost effective, then patients at high risk of life-threatening disease could be identified early and treated aggressively. The mortality associated with infective endocarditis is substantial and surgery can be lifesaving, but current guidelines recognise that indications for surgery are not supported by strong evidence. Staphylococcus aureus is the most common pathogen causing infective endocarditis and a single nucleotide polymorphism in the platelet glycoprotein GP1b receptor has been associated with increased risk of septic emboli in S aureus infective endocarditis.6 If confirmed by clinical studies, this finding could help stratify patients with S aureus infective endocarditis by providing an indication for early surgery in carriers of the risk allele, so reducing embolic complications in these patients. A better understanding of infectionassociated host genetic variation and the detection of novel biomarkers associated with the course of disease clearly have the potential to influence the clinical management of patients with infectious diseases. As well as continuing to apply genomic methods to important infections, there is a strong and timely need in infectious
disease medicine to translate the results of these new genomic studies into clinical practice. In their Comment in The Lancet Infectious Diseases, Jake Dunning and colleagues7 make the case for opensource cooperation to stimulate clinical research of infectious diseases. Such an approach will be especially important if we are to understand fully how genes affect the susceptibility of people to infectious diseases. To accomplish this, worldwide cooperation will be essential, and clinicians and scientists will have to embrace new ways of working together.7 Only through new strategies such as this can we meet the huge challenges, in both high-income and low-income countries, posed by soaring health-care costs, multidrug resistant pathogens, and emerging infections. We declare no competing interests.
*Clark D Russell, Samantha J Griffiths, J Kenneth Baillie, Juergen Haas [email protected] Division of Pathway Medicine, University of Edinburgh, The Chancellor’s Building, 49 Little France Crescent, Edinburgh EH16 4SB, UK (CDR, SJG, JH); and The Roslin Institute, University of Edinburgh, Midlothian, UK (JKB) 1
2
3
4
5
6
7
Sørensen TI, Nielsen GG, Andersen PK, et al. Genetic and environmental influences on premature death in adult adoptees. N Engl J Med 1988; 318: 727–32. Chapman SJ, Hill AV. Human genetic susceptibility to infectious disease. Nat Rev Genet 2012; 13: 175–88. Ge D, Fellay J, Thompson AJ, et al. Genetic variation in IL28B predicts hepatitis C treatment-induced viral clearance. Nature 2009; 461: 399–401. Griffiths SJ, Koegl M, Boutell C, et al. A systematic analysis of host factors reveals a Med23-interferon-λ regulatory axis against herpes simplex virus type 1 replication. PLoS Pathog 2013; 9: e1003514. Everitt AR, Clare S, Pertel T, et al. IFITM3 restricts the morbidity and mortality associated with influenza. Nature 2012; 484: 519–23. Daga S, Shepherd JG, Hung RK, et al. GPIb VNTR C/C genotype may predict embolic events in infective endocarditis. J Heart Valve Dis 2013; 22: 133–41. Dunning JW, Merson L, Rohde GG, et al. Open source clinical science for emerging infections. Lancet Infect Dis 2014; 14: 8–9.
451
Proportion of viable parasites (%)
100
10
1
0·1 Wild-type Arg539Thr Cys580Tyr Tyr493His
0·01
0·001 Fast
Slow Parasite clearance
and slow-clearing parasites with concordant survival rates had wildtype or mutant alleles (Tyr493His and Cys580Tyr), respectively. These data suggest that parasite survival rates in the ring-stage survival assay are more relevant than parasite clearance half-lives for identifying artemisinin-resistant P falciparum, and further validate K13-propeller polymorphisms as a molecular marker of artemisinin resistance in vitro and in vivo. We declare no competing interests.
Figure: Association between parasite survival according to the in-vitro ring-stage survival assay (0–3 h) and parasite clearance in patients treated with artemisinin-based combination therapy, by K13-propeller genotype
(5·30%, 19·32%, and 51·39%) but a resistant stage-dependent pattern (decrease in survival as parasites matured of 1·2%, 17·3%, and 50·2%, respectively). We postulated that these three parasites were indeed artemisinin-resistant, but that their clearance was fast because circulating parasites seen in blood films were predominantly older, drug-sensitive rings at the time the patients received artemisinin drugs. Of the 13 slow-clearing parasites, one had a paradoxically low survival (0·16%). We believe that this parasite was in fact artemisinin-sensitive, but that its clearance was slow because the patient had low levels of parasite-clearing immunity2 or a poor pharmacokinetics profile. We reported K13-propeller polymorphism as a new molecular marker for artemisinin-resistant P falciparum malaria,3 and postulated that K13propeller genotypes would definitively resolve the four discordant results. To test this possibility we genotyped all 26 parasites and found that the three parasites with discordantly high survival rates each carry a different mutant K13-propeller allele (Tyr493His, Cys580Tyr, and Arg539Thr), and that the parasite with a discordantly low survival rate carried a wildtype allele (figure). As expected, we also found that all the fast-clearing 450
Chanaki Amaratunga, Benoit Witkowski, Nimol Khim, Didier Menard, *Rick M Fairhurst [email protected] Laboratory of Malaria and Vector Research, NIAID/National Institutes of Health, Rockville, MD 20852, USA (CA, RMF); and Malaria Molecular Epidemiology Unit, Institut Pasteur du Cambodge, Phnom Penh, Cambodia (BW, NK, DM) 1
2
3
Witkowski B, Amaratunga C, Khim N, et al. Novel phenotypic assays for the detection of artemisinin-resistant Plasmodium falciparum malaria in Cambodia: in-vitro and ex-vivo drug-response studies. Lancet Infect Dis 2013; 13: 1043–49. Lopera-Mesa TM, Doumbia S, Chiang S, et al. Plasmodium falciparum clearance rates in response to artesunate in Malian children with malaria: effect of acquired immunity. J Infect Dis 2013; 207: 1655–63. Ariey F, Witkowski B, Amaratunga C, et al. A molecular marker of artemisinin-resistant Plasmodium falciparum malaria. Nature 2014; 505: 50–55.
monotherapy with artemisinin drugs for treatment of falciparum malaria— particularly the chloroquine-resistant strain—is 5–7 days.2 A study of patients treated with dihydroartemisinin (60 mg) daily (120 mg on day 1) for 3, 5, or 7 days showed no recrudescences in the 7 day group, compared with recrudescence in 66% of patients in the 5 day group and 52% in the 3 day group. Therefore, it was recommended that the 7 day course, with a total dose of 480 mg of dihydroartemisin, is the optimum regimen for treatment of P falciparum infection.3 Another feature of clinical malaria management in China is the quarantine of patients during treatment to prevent transmission and hamper the spread of resistance. Combination of artemisinin drugs with other antimalarial treatments gives more effective treatment at the early stage of malaria infections, although, it is becoming clear that the number of treatment days has a major impact on efficacy of artemisinin treatment either as monotherapy or in combination, especially if ineffective partner drugs are used.4 Although we do not have evidence to demonstrate this hypothesis so far, increasing the duration of artemisinin treatment and use of efficacious partner drugs could sustain the efficacy of artemisinin combination therapies. We declare no competing interests.
Benoit Witkowski and colleagues1 reported a simple and rapid in vitro method to detect artemisinin resistance of P falciparum. They show that the slow clearance of parasites by artemisinin combination therapies is a result of a drop in susceptibility among young ring form parasites whereas susceptibility is maintained in mature stages. However, the clinical relevance of this finding to treatment failure is unclear since these infections could still be successfully treated with artemisinin combination therapies. Experiences in China also suggest that such resistance could be overcome if the right strategies are used. In China, the recommended duration of
*Zhao-Rong Lun, Pedro E Ferreira, Lin-Chun Fu [email protected] Key Laboratory of Gene Engineering of Ministry of Education, State Key Laboratory of Biocontrol, School of Life Sciences, Key Laboratory of Tropical Disease Control, Zhongshan School of Medicine, Sun Yat-Sen University, Guangzhou 510275, China (ZRL); School of Biological Sciences, Nanyang Technological University, Singapore (PEF); and Tropical Medicine Institute, Guangzhou University of Chinese Medicine, Guangzhou, China (LCF) 1
2
Witkowski B, Amaratunga C, Khim N, et al. Novel phenotypic assays for the detection of artemisinin-resistant Plasmodium falciparum malaria in Cambodia: in-vitro and ex-vivo drug-response studies. Lancet Infect Dis 2013; 13: 1043–49. The Ministry of Health of China. National guideline for use and regimen of antimalarials. Chin J Parasit Dis Control 2002; 15: 129–30.
www.thelancet.com/infection Vol 14 June 2014
Correspondence
3
4
Fu LC, Guo XB, Wang WL, et al. An exploration on the dosage and treatment course of dihydroartemisinin for falciparum malaria. Trad Chin Drug Res Clin Pharm 1996; 7: 17–18. Ferreira PE, Culleton R, Gil JP, Meshnick SR. Artemisinin resistance in Plasmodium falciparum: what is it really? Trends Parasitol 2013; 29: 318–20.
Stratified infection medicine: a call to arms Human susceptibility to infectious disease is strongly heritable.1 In the past decade, genetic studies have found more than 30 common genetic variants that have profound effects on susceptibility to infection, course of disease, and response to treatment.2 A major challenge for infectious disease medicine in the next decade will be making use of this knowledge to benefit patients. Stratification of patients, by customising management according to their genotypes, makes little impact on clinical treatment of infectious disease. We are aware of only one example—hepatitis C virus infection— in which this new paradigm has been exploited. In infection with hepatitis C virus genotype 1, a single nucleotide polymorphism in the IFNL3 gene is associated with a sustained virological response to treatment, and this can be clinically tested to predict response to therapy.3 By contrast, oncologists successfully and routinely apply knowledge of both patient’s and tumour’s genotypes to allocate patients to ever-smaller groups for whom optimum therapeutic strategies have been found. For example, HER2positive breast cancer responds to trastuzumab, and lung cancers carrying EGFR mutations respond to oral EGFR tyrosine kinase inhibitors. Several single nucleotide polymorphisms associated with infectious disease phenotypes could have implications for clinical practice.
www.thelancet.com/infection Vol 14 June 2014
The IFNL3 mutation associated with hepatitis C virus is also associated with the severity and frequency of reactivation of orofacial herpes simplex virus type 1 infection,4 which highlights two major concepts: first, that there are genetic variants that confer susceptibility to multiple pathogens; and second, that there are likely to be genotype-determined comorbidities that have not yet been observed and reported. Influenza A virus is a pathogen of global importance and a genetic polymorphism in IFITM3 is strongly associated with risk of severe influenza in human beings.5 If the clinical impact of this mutation can be quantified, and genotyping can be made both rapid and cost effective, then patients at high risk of life-threatening disease could be identified early and treated aggressively. The mortality associated with infective endocarditis is substantial and surgery can be lifesaving, but current guidelines recognise that indications for surgery are not supported by strong evidence. Staphylococcus aureus is the most common pathogen causing infective endocarditis and a single nucleotide polymorphism in the platelet glycoprotein GP1b receptor has been associated with increased risk of septic emboli in S aureus infective endocarditis.6 If confirmed by clinical studies, this finding could help stratify patients with S aureus infective endocarditis by providing an indication for early surgery in carriers of the risk allele, so reducing embolic complications in these patients. A better understanding of infectionassociated host genetic variation and the detection of novel biomarkers associated with the course of disease clearly have the potential to influence the clinical management of patients with infectious diseases. As well as continuing to apply genomic methods to important infections, there is a strong and timely need in infectious
disease medicine to translate the results of these new genomic studies into clinical practice. In their Comment in The Lancet Infectious Diseases, Jake Dunning and colleagues7 make the case for opensource cooperation to stimulate clinical research of infectious diseases. Such an approach will be especially important if we are to understand fully how genes affect the susceptibility of people to infectious diseases. To accomplish this, worldwide cooperation will be essential, and clinicians and scientists will have to embrace new ways of working together.7 Only through new strategies such as this can we meet the huge challenges, in both high-income and low-income countries, posed by soaring health-care costs, multidrug resistant pathogens, and emerging infections. We declare no competing interests.
*Clark D Russell, Samantha J Griffiths, J Kenneth Baillie, Juergen Haas [email protected] Division of Pathway Medicine, University of Edinburgh, The Chancellor’s Building, 49 Little France Crescent, Edinburgh EH16 4SB, UK (CDR, SJG, JH); and The Roslin Institute, University of Edinburgh, Midlothian, UK (JKB) 1
2
3
4
5
6
7
Sørensen TI, Nielsen GG, Andersen PK, et al. Genetic and environmental influences on premature death in adult adoptees. N Engl J Med 1988; 318: 727–32. Chapman SJ, Hill AV. Human genetic susceptibility to infectious disease. Nat Rev Genet 2012; 13: 175–88. Ge D, Fellay J, Thompson AJ, et al. Genetic variation in IL28B predicts hepatitis C treatment-induced viral clearance. Nature 2009; 461: 399–401. Griffiths SJ, Koegl M, Boutell C, et al. A systematic analysis of host factors reveals a Med23-interferon-λ regulatory axis against herpes simplex virus type 1 replication. PLoS Pathog 2013; 9: e1003514. Everitt AR, Clare S, Pertel T, et al. IFITM3 restricts the morbidity and mortality associated with influenza. Nature 2012; 484: 519–23. Daga S, Shepherd JG, Hung RK, et al. GPIb VNTR C/C genotype may predict embolic events in infective endocarditis. J Heart Valve Dis 2013; 22: 133–41. Dunning JW, Merson L, Rohde GG, et al. Open source clinical science for emerging infections. Lancet Infect Dis 2014; 14: 8–9.
451
