acute plasmodium falciparum malaria is associated with varying degrees of neurological involvement , depending on the severity . few studies have , however , investigated the effect of the disease on hearing specifically in those with uncomplicated malaria . furthermore , several antimalarial drugs , including quinine [ 1 , 2 ] , chloroquine , and mefloquine , have been associated with ototoxicity , and certain artemisinin derivatives have also been associated with neuro- or ototoxicity in various animal species [ 59 ] . although human studies that have evaluated possible artemisinin - related effects on hearing have , with the exception of one study , concluded lack of any clinically relevant ototoxicity or neurotoxicity [ 1115 ] , the majority of these studies have been done in adults , in spite of the fact that children are the subgroup of patients who , because of their still developing nervous systems , are more susceptible to such potential treatment - related effects . the lack of studies evaluating the potential effects of newly introduced antimalarial drugs on hearing in children is due to the difficulties in conducting serial audiometric measurements in childhood populations , especially in resource - poor malaria - endemic countries where the disease is most prevalent . in most malaria - endemic countries , there is , indeed , near - absence of pre - illness hearing threshold data with which to compare data obtained during acute illness . this makes it difficult to conduct hearing assessment studies during malarial illness , as it becomes nearly impossible to distinguish between disease - specific and emergent or drug - related effects in such areas . in spite of these challenges , however , studies investigating trends in hearing thresholds during malaria are important , because malaria , even when uncomplicated , has been implicated as a cause of elevated audiometric thresholds in experimentally infected ( nonimmune ) adults with uncomplicated malaria , as well as in animals . in addition , repeated attacks of uncomplicated malaria have been implicated as a cause of poor cognitive performance in school children , but few studies have focused audiometric studies specifically on children . we have , in this study , conducted serial audiometric measurements with a follow - up time ranging between 9 and 15 months , in children with uncomplicated malaria treated with artesunate - amodiaquine , artemether - lumefantrine , or amodiaquine monotherapy . these measurements have been done not only to evaluate artemisinin - based and non - artemisinin - based antimalarial treatments , but also to compare artemisinin combination therapy ( act ) regimens that have been associated with differential propensities for neurotoxicity in animal studies . the audiometric measurements were also compared with those of age - matched children from the same area . the study design also permits comparison of audiograms obtained during acute illness with audiograms done at different postrecovery time points , allowing assessment of the potential effect of the acute malarial disease and recovery on the dynamics of hearing threshold changes over the study period . the audiometric assessment was a subcomponent of a clinical trial that was done to evaluate the efficacy and safety of artemisinin - based combination therapies for uncomplicated malaria in ghana . approval for the study was granted by the ethics and protocol review committee of the university of ghana medical school . the clinical trial was initiated at a time when chloroquine was still the official first - line treatment for uncomplicated malaria in ghana . the full description of the study site and trial results have been previously reported . briefly , enrolled children aged 0.514 years with uncomplicated malaria were treated with ( i ) artesunate ( plasmotrim , mepha ; switzerland ) , 4  mg / kg body weight as a single daily dose + amodiaquine ( camoquine pfizer ; dakar , senegal ) , 10  mg / kg body weight single daily dose , for 3 days , or ( ii ) artemether - lumefantrine ( coartem , novartis pharma ag , basel , switzerland ; 20  mg artemether and 120  mg lumefantrine ) , given at 0 and 8 hours on the first day and then twice daily for the two subsequent days according to body weight : 914  kg , 1 tablet / dose ; 1524  kg , 2 tablets / dose ; 2534  kg , 3 tablets / dose ; 35  kg and over , 4 tablets / dose . a limited number of children were treated with amodiaquine monotherapy ( same dosage as above ) ; however , this ( monotherapy ) treatment arm was discontinued in early 2005 when the ghana national first - line antimalarial treatment policy was changed from chloroquine to artesunate - amodiaquine . this monotherapy - treated - group , however , is crucial for inclusion as non - artemisinin treated control subjects in the context of the objectives of this study and has been maintained in the data analysis despite its limited size . in addition , a control group of randomly selected age- and sex - matched children were enrolled from a school in the study area as controls . children with conditions such as serous otitis media , active or inactive chronic suppurative otitis media , impacted cerumen , eardrum perforation , tympanic membrane scarring , or other clinically evident outer and middle ear abnormalities , were excluded . children with a past medical history or clinically obvious symptoms and signs of sickle cell anaemia , renal or liver disease , malnutrition , craniofacial abnormalities , or dysmorphism , as well as children , when they were newborns , were reported to have been admitted into neonatal intensive care for any cause or those with a history of head trauma were excluded . additionally , children with a past medical history suggesting possible birth asphyxia , neonatal jaundice , and meningitis , and those who were known to have taken aminoglycoside antibiotics , loop diuretics , or herbal medications within the past three months were excluded . air conduction threshold was obtained for each ear separately in a quiet ( nonairtight ) room , using a portable audiometer ( kamplex ks8 ; pc werth , london , uk ) , with noise - attenuated tdh-39 headsets and earphones . the level of ambient noise measured in the examination room was < 50  db a ( within the level recommended by the american national standard for ambient noise levels for audiometric test room standards ) . the unmasked psychoacoustical hearing thresholds were established at 0.125 , 0.25 , 0.5 , 0.75 , 1 , 1.5 , 2 , 3 , 4 , 6 , and 8  khz , for tonal stimuli , using a standard two - step - down , one - step - up method , as per the modified hughson - westlake procedure . briefly , pure tones at the specified frequency were applied and then decreased , in 10  db steps until no response was elicited from the subject . the applied tone was subsequently increased in 5  db steps until a response elicited . this procedure was repeated , and the hearing threshold at a particular frequency was determined as the average of the lowest sound intensity the subject responded to . the control subjects were tested once , under similar conditions , using the same equipment , and following the same procedure as the subjects . venous blood was collected into edta and heparinised tubes on days 0 , 3 , 7 , 14 , and 28 and on any day of recurrent symptomatic parasitaemia for routine haematological and biochemical investigations . parasite counts were determined in giemsa - stained blood films relative to 200 white blood cells ( wbcs ) and the measured wbc count . total wbcs and differential counts were measured by an automated haematology analyzer ( cell dyn , abbott laboratories , usa ) . continuous normally distributed data were described by the mean and standard deviation or standard error and non normally distributed data by the median and range . for the latter data , all statistics were performed on ranks . percentages were given for categorical data , which were compared using the chi - square test with yates ' correction , or fischer 's exact test , as appropriate . one - way analysis of variance , with holm - sidak post hoc pairwise testing , or two - way repeated measures analysis of variance was used to compare variables between the treatment groups and controls or to test for intra - individual and within - group differences , respectively . for some comparisons , a delta - value was calculated by subtracting the values measured at two time points in the same individual . multiple linear regression analysis was used to evaluate the effect of selected subject characteristics on the audiometric thresholds . statistics were done , using sigmaplot 11 ( systat , chicago , il , usa ) . audiometric analysis was performed in 80 malaria patients ( artesunate - amodiaquine , n = 37 ; artemether - lumefantrine , n = 35 ; amodiaquine , n = 8) on days 0 , 3 , 7 , and 28 ; however , for each day of repeated testing , various children did not turn up , explaining the variable numbers of comparison in each analysis below . in all repeated comparisons , only patients tested at both time points were included in the data analysis . at the end of the follow - up period , audiometry was done in 58 of the originally recruited 80 treated children ( artesunate - amodiaquine , n = 30 ; artemether - lumefantrine , n = 23 ; amodiaquine , n = 5 ) and in 57 healthy controls .   on day 0 ( before treatment ) , hearing thresholds of all the acutely ill subjects were significantly elevated compared with the hearing thresholds of the control subjects ( figure 1(a ) , two - way repeated measurement anova on ranks p < 0.001 , post hoc test significant for all frequencies ) . hearing threshold levels were similar for the three treatment groups , each of which was significantly different from the control group ( except 6000  hz , table 1 ) . there were no differences in hearing threshold levels from day 0 to day 3 or from day 3 to day 7 ( p between 0.1 and 0.9 , signed rank test for all frequencies / ears and both days , data not shown ) . there were significant improvements ( decreases ) in hearing threshold levels on day 28 compared with day 0 , at specific frequencies ( 250  hz and 500  hz for both ears and also at 1000  hz for the left ear , signed rank test performed for each frequency and ear , p < 0.05 , n = 47 , figure 2(a ) , for a representation across all wavelengths ) . of a particular note , despite the improvement , the mean hearing threshold levels remained significantly elevated on day 28 compared with followup at 915 months ( figure 2(b ) , p < 0.01 , n = 38 ) . the hearing threshold level measurements of treated children measured after at least 9 months were comparable , or even marginally ( but clinically nonsignificantly ) , lower than the hearing thresholds of controls ( figure 1(b ) , two - way repeated measurement anova on ranks p = 0.01 , median difference < 5  db ) , except for measurements for the right ear at 125  hz and 250  hz ( table 2 ) . at the final followup ( 915 months ) , there were no differences in the mean hearing threshold levels between the amodiaquine monotherapy and the artesunate - amodiaquine or artemether - lumefantrine arms ( figure 3(a ) , two - way repeated measurement anova on ranks , p = 0.7 ) . on day 7 , the hearing threshold levels in the amodiaquine monotherapy arm were higher , whereas in the two act arms they remained at the level of day 0 . thus , the hearing threshold levels on day 7 in the monotherapy arm were significantly higher than the act arms , especially at frequencies < 1000  hz ( figure 3(b ) , two - way repeated measurement anova on ranks , p < 0.001 , holm - sidak post hoc test for amodiaquine versus artemether - lumefantrine , p < 0.001 , of amodiaquine versus artesunate - amodiaquine , p = 0.003 ; amodiaquine , n = 5 , artemether - lumefantrine , n = 21 , artesunate - amodiaquine , n = 15 ) . a linear regression analysis performed to compare the effect of selected characteristics on hearing threshold levels showed that a lower mean age and disease severity ( i.e. , higher parasite density ) were the most consistent predictors of elevated hearing threshold measured on the exit ( follow - up ) audiogram ( data not shown ) . in this study , serial audiometric measurements were done before treatment and at prespecified times after treatment of uncomplicated malaria , with act regimens that have been associated with different propensities for toxicity in animal studies , as well as in subjects treated with a nonartemisinin ( amodiaquine ) antimalarial drug . the results showed elevated hearing thresholds in all three groups of children at presentation and through the acute illness stage but complete reversibility after a sufficiently long follow - up period . although the data shows progressive improvement ( decrease ) of hearing threshold levels during the initial days of treatment , especially in the two act groups , there were some residual elevation 28 days after treatment . the pattern of hearing threshold elevations , which was more pronounced at the lower frequencies , is consistent with that of the malaria - attributed hearing impairment that has been recently demonstrated in animals . it has also been shown , by fundoscopic examinations , that uncomplicated malaria may be associated with some neurological effects , a finding that has potential implications for development , especially in areas of intense malaria transmission . in this respect , the trend towards widening the target group for intermittent preventive treatment for malaria with the aim of reducing the number of malaria attacks in certain settings [ 21 , 22 ] could be partly supported by the findings from this study . the data did not show any differences between hearing threshold levels of children treated with artesunate - amodiaquine or artemether - lumefantrine . however , treatment with act - based antimalarials seemed to halt the persistence or progression of the threshold elevations during acute illness , whereas amodiaquine monotherapy treatment did not show such an effect . although the number of subjects in the amodiaquine monotherapy group was limited , precluding definitive conclusions on differences between act and non - act treatment groups , this apparent difference between the act and non - act antimalarial regimens could be due to the rapid parasite clearance and faster fever resolution in the act groups . the quicker symptom resolution in the act groups would be expected to have relatively lower effect on subject concentration and ability to respond to the tonal stimuli . however , the artemisinin derivatives may also play a direct role by modifying cerebral immunopathology , as described in a murine model of experimental cerebral malaria . the findings of this study therefore suggest that there were minimal , if any , detectable effect of the two act regimens , at the administered doses , on hearing as measured by audiometry . furthermore , since hearing threshold change is considered a better indicator of potential drug - induced ototoxicity than hearing threshold levels , the complete reversibility of the noted changes further suggest disease- rather than drug - induced changes . however , a more definitive statement of a lack of effect of treatment with these act regimens on hearing threshold changes could have been made if plasma concentration data of dihydroartemisinin , the main metabolite of artemisinins , were available . also , since preillness baseline audiometric threshold levels were not available in this study  as in most malaria audiometric studies  further studies that utilize preillness audiometric thresholds as a baseline should be considered  in so far as this is feasible in endemic areas . the treatment responses from the efficacy trial , however , suggest that therapeutic levels were attained . the complete reversibility of the elevated threshold elevations in all the three groups at the nine - month follow - up audiogram suggests acute malaria to be the likely cause of these changes , which is consistent with results from studies in adults [ 12 , 13 , 15 ] . furthermore , the ( slightly ) better performance of subjects compared with controls at the exit audiogram is possibly due to a learning effect , most likely from repeated testing , as it has been reported that test scores of pure tone audiometry in children , improve with age or after repeated examinations [ 2527 ] . it has been hypothesized that malaria could contribute to hearing impairment by impairing labyrinth artery microcirculation   and it has also been shown that antimalarial drugs could , by disrupting cochlear vasculature , increase susceptibility to ototoxicity . these findings taken together imply that not only does hearing impairment occur as part of the natural history of uncomplicated malaria , but also if audiometry is used to evaluate drug induced ototoxicity of newly introduced antimalarial drugs , the effect of the disease could confound any hearing threshold elevations if testing of different individuals is done at different posttreatment times . audiometric thresholds measured in children with uncomplicated malaria treated with different antimalarial regimens showed reversible hearing threshold elevations in all treated groups , implying that these changes were disease- rather than drug - related . this has potential implications for learning , development , and behaviour of children repeatedly exposed to malaria in endemic areas and also has implications for evaluating potential ototoxicity of newly introduced antimalarial drugs .
background . plasmodium falciparum malaria , as well as certain antimalarial drugs , 
 is associated with hearing impairment in adults . 
 there is little information , however , on the extent , if any , of this effect in children , and the evidence linking artemisinin combination therapies ( acts ) with hearing is inconclusive 
 . methods . 
 audiometry was conducted in children with uncomplicated malaria treated with artesunate - amodiaquine ( n = 37 ) , artemether - lumefantrine ( n = 35 ) , or amodiaquine ( n = 8) in accra , ghana . 
 audiometry was repeated 3 , 7 , and 28 days later and after 9 months . 
 audiometric thresholds were compared with those of a control group of children ( n = 57 ) from the same area . 
 findings . during the acute stage , hearing threshold levels of treated children were significantly elevated compared with controls ( p < 0.001 ) . 
 the threshold elevations persisted up to 28 days , but no differences in hearing thresholds were evident between treated children and controls after 9 months . 
 the hearing thresholds of children treated with the two act regimens were comparable but lower than those of amodiaquine - treated children during acute illness . 
 interpretation . 
 malaria is the likely cause of the elevated hearing threshold levels during the acute illness , a finding that has implications for learning and development in areas of intense transmission , as well as for evaluating potential ototoxicity of new antimalarial drugs .