Drug resistant malaria in Papua New Guinea and molecular monitoring of parasite resistance

Malaria is a serious global health problem and in the absence of an effective vaccine, access
to safe and effective treatment still remains the mainstay in the control of the disease.
However, the efficacy of this control strategy is hampered by the emergence and spread of
drug resistant malaria which may lead to excess of mortality. One of the greatest challenges
for health authorities of malaria endemic countries is thus to decide on when and how
antimalarial drug policy should be changed, so that most of the patients will fully recover
from the disease and will be cleared from parasites.
The current ‘gold standard’ for the assessment of antimalarial resistance is the estimation of in
vivo drug efficacy, whereas in vitro drug sensitivity tests and the analysis of molecular
resistance markers in the parasite serve as complementary tools.
In the present study, we assessed the relevance of a new appraisal approach for malaria
resistance: community-based cross-sectional surveys versus clinical malaria studies, and the
usefulness of a new molecular technology for the identification of molecular markers in
different parasite genes. The frequencies of single nucleotide polymorphisms (SNPs) in given
resistance marker genes, as well as genotype patterns were analyzed in clinical samples and
their role in predicting in vivo treatment response was investigated. Furthermore, community
drug resistance profiles were correlated with the incidence risk of clinical treatment failure in
order to evaluate the relevance and usefulness of such a novel approach in the management of
drug use.
In Papua New Guinea (PNG), the 4-aminoquinoline drugs amodiaquine (AQ) and chloroquine
(CQ) have been first-line treatment against uncomplicated malaria until the late 1990s. At the
same time, resistance of Plasmodium falciparum and P. vivax to these drugs had reached
unacceptably high levels and health authorities were prompted to revise antimalarial treatment
policy in 1997. First efficacy trials with the combination of AQ or CQ plus SP conducted
between 1998 and 1999 showed good efficacy against falciparum and vivax malaria and the
PNG Department of Health chose these combination regimens to replace the monotherapy
with AQ or CQ as the standard first-line treatment against uncomplicated malaria in 2000.
The in vivo studies we conducted between 2003 and 2005 were the first ones to assess the
therapeutic efficacy of the newly introduced combination regimen against P. falciparum and
P. vivax malaria using the revised WHO standard protocol. In our studies conducted in three
different areas over the period of three consecutive years, we observed PCR-corrected
treatment failure rates up to 28% for P. falciparum and 12% for P. vivax malaria.
Regarding former drug history in PNG (i.e., long lasting 4-aminoquinoline use and sporadic
use of SP as mass chemprophylaxis or partner drug with quinine for second-line treatment),
we found a genetic background in the parasite population that is associated with high CQ as
well as moderate pyrimethamine resistance. We also observed the emergence of mutations
concordant with a sulphadoxine resistant phenotype, indicating that the efficacy of the sulpha
component is already compromised. Further results that identified key pfdhps mutations to be
most relevant in predicting treatment failure with the current first-line regimen corroborated
our findings that AQ and CQ as inefficacious partner drugs of SP in the new standard
treatment were not able to curb both, the progression of pyrimethamine resistance as well as
the emergence of sulphadoxine resistance in PNG.
We have shown that our community-based molecular monitoring approach was feasible in
PNG and that molecular monitoring of parasite resistance can indeed be a valuable
supplementary tool in malaria resistance surveillance. However, our data also clearly
highlighted several drawbacks of the presently applied methods for the assessment of
resistance, the most important being the lack of standardised methods that are applicable in
different epidemiological settings. In addition, our data indicate that currently suggested
public health models for the molecular monitoring of parasite resistance are not suitable for
universal application in settings which are different with regard to several factors such as
malaria endemicity, transmission intensity and drug use patterns.
To summarize, decreasing in vivo efficacy of the current first-line regimen in PNG and the
molecular drug resistance profile of the parasite population consistent with a CQ and SP
resistant phenotype strongly indicate that a policy change to artemisinin-based combination
therapy (ACT) has to be considered in the near future. We have shown that a careful baseline
evaluation of the molecular resistance background is needed for the identification of the most
relevant molecular markers for longitudinal monitoring in a given area. The novel DNA
microarray-based method which allows the parallel analysis of multiple drug resistanceassociated
SNPs has been proven to be a valuable tool to assess the usefulness of each known
molecular marker in a particular region with specific drug use. Moreover, the new technology
enabled the assessment of molecular markers on an epidemiological scale and hence opened
the avenue for the investigation of a more comprehensive community-based monitoring
programme.
To conclude, the novel technical tool for the assessment of molecular markers of parasite
resistance presented in the current study is cheap, easy to use, and applicable in laboratories
with limited infrastructure. Moreover, the technology is highly versatile and allows rapid
adaptation to specific monitoring needs, the most important at the moment being the close
monitoring of resistance to the highly effective artemisinin derivates and potential partner
drugs in ACTs. Though molecular markers have been proven to be useful as an early warning
system, their usefulness in predicting treatment response and the progression of resistance is
still limited. Hence, currently suggested public health models based on molecular data will
have to include additional parameters for important determinants of parasite resistance and to
be evaluated in varying epidemiological settings before molecular methods may eventually
replace in vivo efficacy studies for the surveillance of resistance.