ANTIMICROBIAL AGENTS AND CHEMOTHERAPY, Feb. 2006, p. 407–413
Copyright © 2006, American Society for Microbiology. All Rights Reserved.
Vol. 50, No. 2
Low Levels of Pyrazinamide and Ethambutol in Children with
Tuberculosis and Impact of Age, Nutritional Status, and
Human Immunodeficiency Virus Infection
S. M. Graham,1,2,3* D. J. Bell,1,3,4S. Nyirongo,1R. Hartkoorn,3S. A. Ward,3and E. M. Molyneux2
Malawi-Liverpool-Wellcome Trust Clinical Research Programme,1Department of Paediatrics,2and Department of Medicine,4
College of Medicine, Blantyre, Malawi, and Liverpool School of Tropical Medicine, University of Liverpool,
Liverpool, United Kingdom3
Received 26 May 2005/Returned for modification 31 May 2005/Accepted 6 November 2005
Recent pharmacokinetic studies that included children found that serum drug levels were low compared to
those of adults for whom the same dosages were used. This study aimed to characterize the pharmacokinetics
of pyrazinamide and ethambutol in Malawian children and to examine the impact of age, nutritional status,
and human immunodeficiency virus (HIV) infection. We conducted a pharmacokinetic study of children
treated for tuberculosis with thrice-weekly pyrazinamide (n ? 27; mean age, 5.7 years) and of a separate group
of children treated with thrice-weekly ethambutol (n ? 18; mean age, 5.5 years) as portions of tablets according
to national guidelines. Malnutrition and HIV infection were common in both groups. Blood samples were taken
just prior to oral administration of the first dose, and subsequent samples were taken at intervals of 2, 3, 4,
7, 24, and 48 h after drug administration. Serum drug levels were low in all children for both drugs; in almost
all cases, the maximum concentration of the drug in serum (Cmax) failed to reach the MIC for Mycobacterium
tuberculosis. The Cmaxof pyrazinamide was significantly lower in younger children (<5 years) than in older
children. The Cmaxof pyrazinamide was also lower for HIV-infected children and children with severe
malnutrition, but these differences did not reach statistical significance. No differences were found for etham-
butol in relation to age, HIV infection, or malnutrition, but the Cmaxwas <2 mg/liter in all cases. Studies of
pharmacokinetic parameters and clinical outcomes obtained by using higher dosages of drugs for treatment of
childhood tuberculosis are needed, and recommended dosages may need to be increased.
There are very few pharmacokinetic (PK) studies of anti-
tuberculosis (anti-TB) drugs in children (8). Dosages for chil-
dren are based on weight and extrapolated from data from
studies with adults, yet pharmacokinetics for children, espe-
cially young children, is likely to be different than for adults.
Studies of ethambutol and pyrazinamide have found lower
plasma drug levels and shorter half-lives in children than in
adults using the same dosages, and the authors have suggested
that dosages per kilogram of body weight need to be higher for
children than for adults (36, 37). Similar conclusions were
drawn from a recent study of isoniazid pharmacokinetics in
South African children (29).
Until recently, the use of the same dosage recommendation
as for adults may not have been an important issue, since
studies that followed these schedules for children found that
outcomes were very good and serious adverse events were rare
(1, 2, 6, 33, 34). These data suggest that adequate levels of drug
were being achieved within a range that was safe. Recent
reports of outcomes for child TB, however, have found much
poorer treatment response than earlier studies (5, 15, 20, 23).
In these studies, human immunodeficiency virus (HIV) is the
most important risk factor for poor treatment response. One
reason for this may be malabsorption of oral anti-TB drugs by
the HIV-infected individual, especially with advanced disease
(10, 11, 26). There are no published reports on the effect of
HIV on the pharmacokinetics of anti-TB drugs in children.
We aimed to characterize the pharmacokinetics of pyrazin-
amide and ethambutol in Malawian children treated with the
standard recommended regimen for TB and to examine the
impact of age, nutritional status, and HIV infection.
MATERIALS AND METHODS
Patient population. Children admitted to Queen Elizabeth Central Hospital,
Blantyre, Malawi, between August 2000 and February 2001 with a diagnosis of
TB and scheduled to be started on anti-TB treatment were considered eligible
and were included in the study following informed consent by their guardians.
Baseline clinical data included demographic data, findings on history and exam-
ination, weight, and hematocrit. Investigations included the Mantoux test, radi-
ography of the chest or spine when appropriate, sputum when available for smear
and culture, and HIV status. Mantoux testing was performed using 0.1 ml (10
IU) of tuberculin purified protein derivative RT23 (1:1,000). Results were read
between 48 and 72 h and recorded as the transverse diameter of palpable
induration. An induration of 10 mm or more was regarded as positive irrespec-
tive of Mycobacterium bovis BCG or HIV status. An induration greater than 5
mm was also regarded as positive if the child was HIV infected. HIV testing using
previously described methods (19) was undertaken on all children whose parents
or guardians gave consent. Nutritional status was graded according to the Well-
come Classification on the basis of percentage of expected weight for age
(WFA): ?80% WFA was graded as normal, 60 to 80% WFA as undernutrition,
and ?60% WFA as marasmus. No children were receiving antiretroviral therapy
at the time of enrollment, and no child had clinical evidence or a history of renal
or liver disease.
Drug dosage and dosing regimen for pyrazinamide. Pyrazinamide, in combi-
nation with isoniazid and rifampin, is recommended for the intensive phase of
TB therapy in Malawi for all forms of TB (21, 35). The recommended dose for
thrice-weekly therapy is 35 mg/kg of body weight, with a range of 30 to 40
mg/kg/dose. Liquid preparations are not available, and pyrizinamide (Phar-
* Corresponding author. Mailing address: Malawi-Liverpool-
Wellcome Trust Clinical Research Programme, P.O. Box 30096,
Blantyre 3, Malawi. Phone: 265-1-676444. Fax: 265-1-675774. E-mail:
mamed, Amsterdam, The Netherlands) was administered orally as 400-mg
tablets or portions of 400-mg tablets three times per week, on Monday,
Wednesday, and Friday, according to the recommended dosages for the
children’s weight ranges: 5 to 8.9 kg, 1/2 tablet; 9 to 14.9 kg, 1 tablet; 15 to
19.9 kg, 1 1/2 tablets; 20 to 24.9 kg, 2 tablets; and 25 to 39.9 kg, 3 tablets (21).
Pyrazinamide was administered orally as the first dose of the initial phase of
therapy at the same time each day, around 6 am, prior to breakfast. All
patients received isoniazid and rifampin at the same time as pyrazinamide,
but none received ethambutol.
Drug dosage and dosing regimen for ethambutol. Ethambutol is recommended
for the intensive phase of TB therapy in Malawi as one of a four-drug combi-
nation (R3H3Z3E3) for new cases of smear-positive pulmonary TB (PTB), cases
of smear-negative PTB with extensive parenchymal involvement, and severe
cases of extrapulmonary TB except for TB meningitis (21). The recommended
dose for thrice-weekly therapy is 30 mg/kg, with a range of 25 to 35 mg/kg/dose
(35). Ethambutol (Pharmamed, Amsterdam, The Netherlands) was administered
orally as 400-mg tablets or portions of 400 mg tablets three times per week, on
Monday, Wednesday, and Friday, according to the recommended dosages for
the children’s weight ranges: 5 to 8.9 kg, 1/2 tablet; 9 to 14.9 kg, 1 tablet; 15
to 24.9 kg, 1 1/2 tablets; and 25 to 34.9 kg, 2 tablets (21). Ethambutol was
administered orally as the first dose of the initial phase of therapy at the same
time as other prescribed anti-TB drugs, around 6 am, prior to breakfast. All
patients receiving ethambutol also received isoniazid, pyrazinamide, and ri-
fampin at the same time.
Sampling schedule. Following informed consent, the study patient was admit-
ted to the research ward, and an intravenous cannula was inserted for regular
blood sampling. The first sample was taken just prior to oral administration (0 h)
of the first dose of anti-TB therapy, and subsequent samples were taken at
intervals of 2, 3, 4, 7, 24, and 48 h after drug administration. Breakfast of maize
porridge and tea was usually consumed within 30 min of taking anti-TB medi-
cation, but the patient remained in bed. The 48-hour sample was taken before
the administration of the next prescribed dose. Blood samples were allowed to
clot, then centrifuged for 10 min, and serum was stored at ?70°C. Specimens
were transported at the completion of the study from Malawi to the University
of Liverpool for assay. On completion of the 48-h sampling procedure, the study
patient was transferred to the pediatric TB ward for ongoing management and
education to encourage adherence.
Sample analysis for pyrazinamide. Plasma pyrazinamide concentrations were
determined by a fully validated high-performance liquid chromatography
(HPLC) method with UV detection. Plasma samples (100 ?l) were transferred to
clean 1.5-ml microcentrifuge tubes followed by the addition of 200 ?l of an
internal standard (acetazolamide at 10 ?g/ml in acetonitrile). After vortex mixing
for 30 s, proteins were precipitated by centrifugation (10 min, 12,000 ? g). The
clear supernatant was transferred to a clean LSL tube and evaporated to dryness
under a stream of nitrogen in a water bath at 37°C. Samples were reconstituted
in mobile phase (300 ?l) and were mixed by vortexing, and the contents were
transferred to an autosampler vial. A 60-?l volume of sample was injected into
the HPLC system. Chromatographic separation was achieved on a HyPurity C18
column (5 ?m particle size; 150 by 4.6 mm diameter) (Thermo Electron Corpo-
ration, Runcorn, Cheshire, United Kingdom), protected by a LiChroCart pre-
column guard using an isocratic mobile phase of water containing 0.06%
trifluoroacetic acid and acetonitrile (95/9, vol/vol), at a flow rate of 1.2
ml/min. Analyte detection was performed on a Spectra 100 variable UV
detector operating at 268 nM (Thermo Electron). The assay was linear in the
range of 0 to 80 ?g/ml, with a lower limit of detection of 100 ng/ml. Inter- and
intraassay variabilities were less than 15%.
Sample analysis for ethambutol. Plasma ethambutol concentrations were de-
termined by a fully validated liquid chromatography-tandem mass spectrometry
(LC-MS-MS) method. Plasma samples (200 ?l) were transferred to clean 1.5-ml
microcentrifuge tubes, followed by the addition of 200 ?l of an internal standard
(propanolol at 1 ?g/ml in acetonitrile). Proteins were precipitated by the addi-
tion of 400 ?l of acetonitrile followed by centrifugation (10 min, 12,000 ? g). A
200-?l volume of supernatant was transferred to an autosample vial. A 2-?l
volume of sample was injected into the HPLC-MS-MS system. Chromatographic
separation was achieved on a HYPERSIL silica column (5 ?m; 50 ? 4.6 mm)
(Thermo Electron Corporation, Runcorn, Cheshire, United Kingdom), pro-
tected by a precolumn guard (Si 60; 5 ?m; Merck, Germany) using an isocratic
mobile phase of 4 mM ammonium acetate and acetonitrile (20:80, vol/vol), at a
flow rate of 0.4 ml/min. Analyte detection was performed on a TSQ7000 triple
quad mass spectrometer operating in the MS-MS mode (Thermo Electron). For
ethambutol, the daughter ion at 115.6 m/z produced from the parent ion at 205
m/z was used for quantitation. For the internal standard, the daughter ion at 116
m/z from the parent ion at 260 m/z was used for quantitation. The assay was
linear in the range of 0 to 12.8 ?g/ml, with a lower limit of detection of 100 ng/ml.
Inter- and intraassay variabilities were less than 15%.
PK analysis. The maximum concentration of the drug in serum (Cmax), the
time to reach Cmax(Tmax), and the area under the concentration-time curve
(AUC) were determined from the concentration-time profile of each patient by
noncompartmental methods using the PK software package KINETICA (version
4.1.1; InnaPhase Corporation). AUC was estimated using the trapezoidal rule.
For pyrazinamide, concentration-time data were available for most of the
children only up to 24 h and not beyond. Consequently, it is impossible to
confidently estimate the terminal elimination phase, and estimations of the AUC
to infinity produced an extrapolated AUC that was unacceptably large (more than
20% of the total AUC from 0 h to infinity). For this reason, the AUC to 24 h
(AUC24) was determined, and we have not attempted to calculate the apparent
clearance or elimination half-life (t1/2) for pyrazinamide. For 13 of the 18 children
with ethambutol concentration-time profiles, we were able to define the terminal
elimination phase, and detailed PK analysis has been restricted to this group.
Published MICs of pyrazinamide for drug-susceptible strains of Mycobacterium
tuberculosis are 6 to 50 mg/liter (28), and those of ethambutol are 1.0 to 2.5
mg/liter (36). For intermittent dosing, reference cutoff points for Cmaxare de-
fined as low at 25 mg/liter and very low at 20 mg/liter for pyrazinamide and as low
at 4 mg/liter for ethambutol (27, 36, 37).
Statistical analysis. Data were analyzed using SPSS (version 11.0.0; SPSS
Inc.). Comparisons of PK data were made using Mann-Whitney tests for HIV
status, nutrition status, age, and reactivity to a tuberculin skin test (TST).
Differences between groups were considered statistically significant at a
P value of ?0.05.
Ethical approval. The study was approved by the Research Ethics Committee,
College of Medicine, University of Malawi.
Pyrazinamide results. Twenty-seven children received thrice-
weekly pyrazinamide as the first dose of the initial phase for TB
treatment. Table 1 shows the clinical characteristics of these
children. Of those with a TST result available, 4 of 18 HIV-
infected children had a reactive TST compared to 2 of 7 who
were not HIV infected (22% versus 29%). The mean WFA was
TABLE 1. Characteristics of study patients
(n ? 27)
(n ? 18)
Mean age (range) (yr)5.7 (0.9–14) 5.5 (1–12)
Mean wt (range) (kg)14.3 (6–30)14.8 (7–33)
Mean % WFA (range)67 (43–104)72 (56–102)
Mean dosage (range)
33 (25–48)33 (24–44)
Other TB drugsRifampin,
TST result (n)
HIV status (n)
408GRAHAM ET AL.ANTIMICROB. AGENTS CHEMOTHER.
64% for HIV-infected children compared to 74% for non-
HIV-infected children, and 63% for those with a nonreactive
TST compared to 66% for those with a reactive TST.
Table 2 shows the results of analysis of the pyrazinamide
pharmacokinetic profiles for the 27 study patients and the
impact on pyrazinamide levels of HIV status, age, and nutri-
tional status. The table also shows the Cmaxand AUC24nor-
malized for the dose taken. The overall range for Cmaxwas
wide (5.78 to 84.1 mg/liter), with 10 patients (37%) recording
Cmaxvalues below the low reference cutoff point of 25 mg/liter
and 9 (33%) below the very low point of 20 mg/liter (27).
Young age was associated with a significantly lower Cmax
(Table 2), although when normalized for the dose given, the
difference did not reach statistical significance (P ? 0.06). Cmax
values were also lower in HIV-infected children, but this dif-
ference was not significant. HIV prevalence was higher for
children under the age of 5 years in this study than for older
children: 13 of 15 (87%) compared to 5 of 12 (42%), respec-
tively (P ? 0.05). The mean (? standard deviation) dose re-
ceived by the younger children was also lower (30.5 ? 5.4
mg/kg) than that for the older age group (35.0 ? 6.4 mg/kg),
but the difference was not significant. The mean (? standard
deviation) dose received by HIV-infected children was similar
to that received by non-HIV-infected children (32 ? 6.3 mg/kg
versus 34 ? 3.2 mg/kg, respectively). The Cmaxwas significantly
higher in children with a reactive TST. The sample size was not
large enough to examine the importance of HIV status as a
as common in older children as in younger children and was
associated with lower values, but these were not significant.
Figure 1 demonstrates the relationship between dosage in
mg/kg of body weight and Cmaxfor both age groups. All five
patients who received a dose of 25 mg/kg had admission weights
of between 7.9 and 8.2 kg, for which the recommended dose is 1/2
tablet, or 200 mg. One child received a dose of 48 mg/kg, which is
beyond the recommended range but in line with recommended
dosing by tablets for weight groups: the child’s weight was 25 kg,
for which the recommended dose in Malawi is 3 tablets, or 1,200
mg. Both children who recorded Cmaxvalues of more than 60
mg/liter were above the age of 5 years, not HIV infected, and not
severely malnourished. Figure 2 shows comparisons of mean con-
centrations (? standard errors) with time in relation to age.
Ethambutol results. Eighteen children received thrice-weekly
ethambutol as part of the initial phase of treatment. Table 1
shows the clinical characteristics of these children. No child had a
diagnosis of gastrointestinal or abdominal TB. All children re-
ceived the recommended regimen for cases of severe PTB or
extrapulmonary TB, R3H3Z3E3(21). Of those who had a TST
result available, 1 of 5 HIV-infected children had a reactive TST
compared to 8 of 10 non-HIV-infected children. The mean WFA
for HIV-infected children was 68%, compared to 75% for non-
HIV-infected children. The mean WFA was the same (72%) for
those with reactive or nonreactive TST.
Table 3 shows results of analysis of ethambutol pharmacoki-
netic profiles for 18 study patients and the impact of HIV, age,
and nutritional status. The overall range for Cmaxwas wide (0.32
to 3.68 mg/liter), with all patients recording a Cmaxbelow the low
reference cutoff point for intermittent dosing of 4 mg/liter (36)
was 2 to 7 h, and Tmaxwas 4 or 7 h for seven (39%) patients. No
significant differences were recorded in Cmaxand Tmaxin relation
to age, nutritional status, or TST result. Tmaxwas significantly
(43%) of the 7 children below the age of 5 years were HIV
Fisher’s exact test). The mean dose received by the younger chil-
dren was the same (33 mg/kg) as that received by older children,
and the mean dose received by HIV-infected children was sim-
ilar to that received by non-HIV-infected children (32 mg/kg
versus 34 mg/kg, respectively). Table 4 shows the volume of
distribution and t1/2data for 13 children. There were no sig-
TABLE 2. Results of pharmacokinetic analysis for pyrazinamidea
All patients 2736.6 (19.7) 3.4 (1.5)376 (328) 1.1 (0.6)11.3 (10.0)
9 41.9 (22.9)
13.1 (12.0) 18
5 48.6 (4.4)b
aAll results are presented as means (standard deviations).
bStatistically significant difference (P ? 0.05).
cP ? 0.09.
VOL. 50, 2006 PK STUDY OF TB DRUGS WITH MALAWIAN CHILDREN409
nificant differences recorded in these data in relation to age,
nutritional, HIV or TST status.
This study provides original pharmacokinetic data for pyra-
zinamide and ethambutol in children. Recommended dosages
for TB in children are from pharmacokinetic studies of adults,
yet there are likely to be important age-related differences in
drug absorption, metabolism, and clearance. We have found
that serum drug levels achieved using intermittent pyrazin-
amide or ethambutol therapy at recommended doses are very
low in Malawian children. Only 2 of 27 children in our study
recorded a maximum serum pyrazinamide concentration above
the median of 66 mg/liter recorded using intermittent dosing
for North American adults and children (37). Both these chil-
FIG. 1. Cmaxof pyrazinamide in relation to dose received. Data for children below the age of 5 years (n ? 15) and for children 5 years old and
older (n ? 12) are compared.
FIG. 2. Comparison of PK profiles for pyrazinamide in relation to age.
410GRAHAM ET AL.ANTIMICROB. AGENTS CHEMOTHER.
dren were 5 years old or older, not HIV infected, and not
severely malnourished. For ethambutol, none of the children
in our study recorded a maximum concentration in serum
above what is considered the low cutoff point of 4 mg/liter for
intermittent dosing from earlier studies of adults (36).
Studies have found lower concentrations and delayed ab-
sorption of anti-TB drugs in children compared to adults re-
ceiving the same dose (29, 36, 37). An important recent study
of 64 South African children under the age of 13 years (median
age, 3.8 years) found that younger children eliminate isoniazid
faster than older children and that children require a higher
mg/kg dose to achieve concentrations comparable to those for
adults (29). An earlier study compared mean Cmaxvalues
among 28 European children of different age ranges who re-
ceived 35 mg/kg ethambutol. That study found lower levels for
the younger children—1.5 mg/liter for 2- to 5-year-olds com-
pared to 2.3 mg/liter for 6- to 9-year-olds and 3.0 mg/liter for
10- to 14-year-olds (14). We did not find a similar trend with
We did find that younger children reached significantly
lower serum pyrazinamide concentrations than older children.
There may be confounders. Younger children in the pyrazin-
amide arm of the study had a significantly higher HIV preva-
lence and received lower mean drug dosages than older chil-
dren. The sample size was not large enough to allow multivariate
analysis. There is some evidence that adults with HIV/AIDS do
not absorb some anti-TB drugs, especially rifampin, as well as
non-HIV-infected patients (10, 11, 26). Absorption might be
especially reduced in HIV-infected individuals with severe immu-
nosuppression and HIV-related enteropathy, but we did not per-
form CD4 cell counts. Pyrazinamide and ethambutol levels
were not significantly lower in HIV-infected or severely mal-
nourished children in our study. A recent study of 48 HIV-
infected adults with TB in the United States, 75% with a CD4
cell count of ?200/mm3, found that adequate concentrations
were achieved with intermittent dosing of pyrazinamide (27).
We compared pharmacokinetic parameters in relation to
malnutrition. Levels of pyrazinamide but not ethambutol were
lower for more malnourished children, but these differences
did not reach the level of significance. Severe malnutrition was
as common in older children as in younger children, and HIV
prevalence was not significantly higher in children with maras-
mus. Previous studies of isoniazid and rifampin in children
have been carried out to examine the impact of malnutrition
and did not find a major effect (30–32).
It is known that reduced absorption occurs if the drugs are
taken with a meal, especially a high-fat meal (24, 25). We are
not sure what impact the taking of a low-fat meal around 30
min after the drugs had on absorption in our study group. The
practice in this study was consistent with the usual practice in
Malawian hospitals when anti-TB treatment is administered.
Important reasons for undertaking this study were the worsen-
ing outcomes for child TB in Malawi and elsewhere in the
region and the relatively recent recommendation for etham-
TABLE 3. Results of pharmacokinetic analysis for ethambutola
All patients18 1.8 (1.2) 3.5 (1.8)0.04 (0.03)
12 1.8 (1.1)
7 1.8 (1.2)
3 1.0 (0.9)
aAll results are presented as means (standard deviations).
bStatistically significant difference.
TABLE 4. Volume of distribution and elimination half-life data for ethambutola
Clearance (liters/h)V (liters)b
All patients1322.2 (12.9)8.6 (4.7)49.0 (62.2) 413.3 (304.3)0.53 (0.33)
aAll results are presented as means (standard deviations). No comparisons show significant differences.
bV, volume of distribution.
VOL. 50, 2006PK STUDY OF TB DRUGS WITH MALAWIAN CHILDREN 411
butol usage for all childhood age groups. The death rate for
2,739 Malawians treated for TB in 1998 was 17%, and the
outcome was unknown for an additional 21% (13). Evidence
from the region including Malawi suggests that coinfection
with HIV is a major reason for the high death rate (15, 17, 18,
23). Malabsorption of anti-TB drugs may be one reason for
poor outcomes for HIV-infected children, but there are likely
to be others, such as wrong diagnosis (17), coinfection with
other pathogens (4, 15), inadequate dosages received (12), and
poorer compliance (9). Another factor contributing to poor
outcome may be that ethambutol is not as effective as rifampin
in the continuation phase of therapy (16). Ethambutol re-
placed thiacetazone in regions of HIV endemicity because
thiacetazone caused severe and often fatal adverse reactions in
HIV-infected adults and children (3, 22). There were concerns
about the use of ethambutol for young children because of
their inability to report the early symptoms of optic neuritis,
the most important side effect, that can lead to blindness. This
is a dose-related side effect, so this risk is considered negligible
if recommended doses are used (7). Ethambutol is now rec-
ommended and commonly used for children of all ages in
standard regimens (21, 35). This study suggests an important
potential problem with ethambutol in that currently recom-
mended doses result in inadequate therapeutic drug levels
rather than any risk of toxicity.
In the majority of developing countries, where most child-
hood TB occurs, anti-TB therapy is available only in tablet
form. This means that the same portions of tablets are given to
all children within a particular age range (21). This is a poten-
tial problem, especially for children with low weights. For ex-
ample, children weighing 5.0 kg and 8.9 kg receive the same
dose. Figure 1 shows that all five patients between 7.9 and 8.2
kg (who received a recommended dose of 200 mg, or 25 mg/kg
) recorded maximum serum drug concentrations of ?30
mg/liter. These recommendations may need to be revised.
We examined the relationship between a reactive TST and
drug levels. This is because a reactive TST is likely to be a
readily available surrogate marker for immunocompetence in
regions where CD4 counts are not available. Although num-
bers were small, the maximum pyrazinamide concentration was
significantly higher in children with reactive TSTs, but there
was no difference for ethambutol. A nonreactive TST can be
due to immunosuppression due to advanced HIV disease or
severe malnutrition, but it may also indicate a wrong diagnosis.
Therefore, the potential use of the TST result in determining
drug dosage would be limited to those with a positive result.
In conclusion, this pharmacokinetic study has found poor
absorption of pyrazinamide and ethambutol in Malawian chil-
dren. It has also found that low serum drug levels are common
using intermittent therapy at recommended doses and that
young age is an important risk factor for low levels. Studies
are needed that compare pharmacokinetic parameters using
higher doses and that measure the impact of higher doses on
outcome, as well as the incidence of adverse reactions.
S.M.G. and D.J.B. are supported by the Wellcome Trust, United
Kingdom, and drug analysis was performed under the Wellcome Trust-
funded LOTLink award. We acknowledge the Research Development
Fund of the University of Liverpool and S. B. Squire of the TB Knowl-
edge Programme, Liverpool School of Tropical Medicine, for provid-
ing support for S.N. to receive training in drug analysis at the Univer-
sity of Liverpool.
We thank I. G. Edwards for assistance with proposal development.
There were no conflicts of interest.
1. Al Dossary, F. S., L. T. Ong, A. G. Correa, and J. R. Starke. 2002. Treatment
of childhood tuberculosis with a six month directly observed regimen of only
two weeks of daily therapy. Pediatr. Infect. Dis. J. 21:91–97.
2. Biddulph, J. 1990. Short course chemotherapy for childhood tuberculosis.
Pediatr. Infect. Dis. J. 9:794–801.
3. Chintu, C., C. Luo, G. Bhat, M. Raviglione, H. DuPont, and A. Zumla. 1993.
Cutaneous hypersensitivity reactions due to thiacetazone in the treatment of
tuberculosis in Zambian children infected with HIV-I. Arch. Dis. Child.
4. Chintu, C., V. Mudenda, S. Lucas, A. Nunn, K. Lishimpi, D. Maswahu, F.
Kasolo, P. Mwaba, G. Bhat, H. Terunuma, and A. Zumla. 2002. Lung
diseases at necropsy in African children dying from respiratory illnesses: a
descriptive necropsy study. Lancet 360:985.
5. Espinal, M. A., A. L. Reingold, G. Perez, E. Camilo, S. Soto, E. Cruz, N.
Matos, and G. Gonzalez. 1996. Human immunodeficiency virus infection in
children with tuberculosis in Santo Domingo, Dominican Republic: preva-
lence, clinical findings, and response to antituberculosis treatment. J. Acquir.
Immune Defic. Syndr. Hum. Retrovirol. 13:155–159.
6. Gocmen, A., U. Ozcelic, N. Kiper, M. Toppare, S. Kaya, R. Cengizlier, and
F. Cetinkaya. 1993. Short course intermittent chemotherapy in childhood
tuberculosis. Infection 21:324–327.
7. Graham, S. M., H. M. Daley, A. Banerjee, F. M. Salaniponi, and A. D.
Harries. 1998. Ethambutol in tuberculosis: time to reconsider? Arch. Dis.
8. Graham, S. M., R. P. Gie, H. S. Schaaf, J. B. Coulter, M. A. Espinal, and N.
Beyers. 2004. Childhood tuberculosis: clinical research needs. Int. J. Tuberc.
Lung Dis. 8:648–657.
9. Graham, S. M., and A. D. Harries. 1999. Childhood TB/HIV co-infection:
correction, confusion and compliance. Int. J. Tuberc. Lung Dis. 3:1144.
Padmapriyadarsini, S. Swaminathan, S. Bhagavathy, P. Venkatesan, L.
Sekar, A. Mahilmaran, N. Ravichandran, and P. Paramesh. 2004. De-
creased bioavailability of rifampin and other antituberculosis drugs in pa-
tients with advanced human immunodeficiency virus disease. Antimicrob.
Agents Chemother. 48:4473–4475.
Padmapriyadarsini, S. Swaminathan, P. Venkatesan, L. Sekar, S. Kumar,
O. R. Krishnarajasekhar, and P. Paramesh. 2004. Malabsorption of ri-
fampin and isoniazid in HIV-infected patients with and without tuberculosis.
Clin. Infect. Dis. 38:280–283.
12. Harries, A. D., F. Gausi, and F. M. Salaniponi. 2004. Prescriptions and
dosages of anti-tuberculosis drugs in the National Tuberculosis Control
Programme of Malawi. Int. J. Tuberc. Lung Dis. 8:724–729.
13. Harries, A. D., N. J. Hargreaves, S. M. Graham, C. Mwansambo, P. Kazembe,
R. L. Broadhead, D. Maher, and F. M. Salaniponi. 2002. Childhood tuber-
culosis in Malawi: nationwide case-finding and treatment outcomes. Int. J.
Tuberc. Lung Dis. 6:424–431.
14. Hussels, H., U. Kroening, and K. Magdorf. 1973. Ethambutol and rifampicin
serum levels in children: second report on the combined administration of
ethambutol and rifampicin. Pneumonologie 149:31–38.
15. Jeena, P. M., P. Pillay, T. Pillay, and H. M. Coovadia. 2002. Impact of HIV-1
co-infection on presentation and hospital-related mortality in children with
culture proven pulmonary tuberculosis in Durban, South Africa. Int. J.
Tuberc. Lung Dis. 6:672–678.
16. Jindani, A., A. J. Nunn, and D. A. Enarson. 2004. Two 8-month regimens of
chemotherapy for treatment of newly diagnosed pulmonary tuberculosis:
international multicentre randomised trial. Lancet 364:1244–1251.
17. Kiwanuka, J., S. M. Graham, J. B. Coulter, J. S. Gondwe, N. Chilewani, H.
Carty, and C. A. Hart. 2001. Diagnosis of pulmonary tuberculosis in children
in an HIV-endemic area, Malawi. Ann. Trop. Paediatr. 21:5–14.
18. Madhi, S. A., R. E. Huebner, L. Doedens, T. Aduc, D. Wesley, and P. A.
Cooper. 2000. HIV-1 co-infection in children hospitalised with tuberculosis
in South Africa. Int. J. Tuberc. Lung Dis. 4:448–454.
19. Molyneux, E. M., A. L. Walsh, H. Forsyth, M. Tembo, J. Mwenechanya, K.
Kayira, L. Bwanaisa, A. Njobvu, S. Rogerson, and G. Malenga. 2002. Dexa-
methasone treatment in childhood bacterial meningitis in Malawi: a ran-
domised controlled trial. Lancet 360:211–218.
20. Mukadi, Y. D., S. Z. Wiktor, I. M. Coulibaly, D. Coulibaly, A. Mbengue,
A. M. Folquet, A. Ackah, M. Sassan-Morokro, D. Bonnard, C. Maurice, C.
Nolan, J. K. Kreiss, and A. E. Greenberg. 1997. Impact of HIV infection on
the development, clinical presentation, and outcome of tuberculosis among
children in Abidjan, Cote d’Ivoire. AIDS 11:1151–1158.
21. National Tuberculosis Control Programme, Malawi. 2002. Manual of the
412GRAHAM ET AL.ANTIMICROB. AGENTS CHEMOTHER.
National Tuberculosis Control Programme of Malawi. Ministry of Health Download full-text
and Population, Lilongwe, Malawi.
22. Nunn, P., D. Kibuga, S. Gathua, R. Brindle, A. Imalingat, K. Wasunna, S.
Lucas, C. Gilks, M. Omwega, J. Were, et al. 1991. Cutaneous hypersensitivity
reactions due to thiacetazone in HIV-1 seropositive patients treated for
tuberculosis. Lancet 337:627–630.
23. Palme, I. B., B. Gudetta, J. Bruchfeld, L. Muhe, and J. Giesecke. 2002.
Impact of human immunodeficiency virus 1 infection on clinical presenta-
tion, treatment outcome and survival in a cohort of Ethiopian children with
tuberculosis. Pediatr. Infect. Dis. J. 21:1053–1061.
24. Peloquin, C. A., A. E. Bulpitt, G. S. Jaresko, R. W. Jelliffe, J. M. Childs, and
D. E. Nix. 1999. Pharmacokinetics of ethambutol under fasting conditions,
with food, and with antacids. Antimicrob. Agents Chemother. 43:568–572.
25. Peloquin, C. A., R. Namdar, M. D. Singleton, and D. E. Nix. 1999. Pharma-
cokinetics of rifampin under fasting conditions, with food, and with antacids.
26. Peloquin, C. A., A. T. Nitta, W. J. Burman, K. F. Brudney, J. R. Miranda-
Massari, M. E. McGuinness, S. E. Berning, and G. T. Gerena. 1996. Low
antituberculosis drug concentrations in patients with AIDS. Ann. Pharma-
27. Perlman, D. C., Y. Segal, S. Rosenkranz, P. M. Rainey, C. A. Peloquin, R. P.
cokinetics of pyrazinamide in HIV-infected persons with tuberculosis. Clin.
Infect. Dis. 38:556–564.
28. Salfinger, M., and L. B. Heifets. 1988. Determination of pyrazinamide MICs
for Mycobacterium tuberculosis at different pHs by the radiometric method.
Antimicrob. Agents Chemother. 32:1002–1004.
29. Schaaf, H. S., D. P. Parkin, H. I. Seifart, C. J. Werely, P. B. Hesseling, P. D. van
Helden, J. S. Maritz, and P. R. Donald. 2005. Isoniazid pharmacokinetics in
children treated for respiratory tuberculosis. Arch. Dis. Child. 90:614–618.
30. Seifart, H. I., P. R. Donald, J. N. De Villiers, D. P. Parkin, and P. P.
Jaarsveld. 1995. Isoniazid elimination kinetics in children with protein-
energy malnutrition treated for tuberculous meningitis with a four-compo-
nent antimicrobial regimen. Ann. Trop. Paediatr. 15:249–254.
31. Seth, V., A. Beotra, A. Bagga, and S. Seth. 1992. Drug therapy in malnutri-
tion. Indian Pediatr. 29:1341–1346.
32. Seth, V., A. Beotra, S. D. Seth, O. P. Semwal, S. Kabra, Y. Jain, and S.
Mukhopadhya. 1993. Serum concentrations of rifampicin and isoniazid in
tuberculosis. Indian Pediatr. 30:1091–1098.
33. Te Water Naude, J. M., P. R. Donald, G. D. Hussey, M. A. Kibel, A. Louw,
D. R. Perkins, and H. S. Schaaf. 2000. Twice weekly vs. daily chemotherapy
for childhood tuberculosis. Pediatr. Infect. Dis. J. 19:405–410.
34. Tsakalidis, D., P. Pratsidou, A. Hitoglou-Makedou, G. Tzouvelekis, and I.
Sofroniadis. 1992. Intensive short course chemotherapy for treatment of
Greek children with tuberculosis. Pediatr. Infect. Dis. J. 11:1036–1042.
35. World Health Organization. 2003. Treatment of tuberculosis: guidelines for
national programmes. World Health Organization, Geneva, Switzerland.
36. Zhu, M., W. J. Burman, J. R. Starke, J. J. Stambaugh, P. Steiner, A. E.
Bulpitt, D. Ashkin, B. Auclair, S. E. Berning, R. W. Jelliffe, G. S. Jaresko,
and C. A. Peloquin. 2004. Pharmacokinetics of ethambutol in children and
adults with tuberculosis. Int. J. Tuberc. Lung Dis. 8:1360–1367.
37. Zhu, M., J. R. Starke, W. J. Burman, P. Steiner, J. J. Stambaugh, D. Ashkin,
A. E. Bulpitt, S. E. Berning, and C. A. Peloquin. 2002. Population pharma-
cokinetic modeling of pyrazinamide in children and adults with tuberculosis.
VOL. 50, 2006 PK STUDY OF TB DRUGS WITH MALAWIAN CHILDREN413