# Efficacy of Artesunate + Sulphadoxine-Pyrimethamine (AS + SP) and Amodiaquine + Sulphadoxine-Pyrimethamine (AQ + SP) for Uncomplicatedfalciparum Malaria in Equatorial Guinea  (Central Africa)

Authors: Pilar Charle; Pedro Berzosa; Miguel Angel Descalzo; Aida de Lucio; Jose Raso; Jacqueline Obono; Magdalena Lwanga; Natividad Nlang; Araceli Nchama; Catalina Mangue; Anastasio Micha; Natividad Nsee; Rosario Mesie; Agustín Benito; Jesús Roche
Journal: Journal of Tropical Medicine
 (2009)

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## Abstract

Objectives. The objectives of the study were (i) to evaluate the efficacy of combination drugs, such as artesunate + sulphadoxine-pyrimethamine (AS + SP) and amodiaquine + sulphadoxine-pyripethamine (AQ + SP) in treatment of uncomplicated falciparum malaria (ii) to differentiate recrudescence from reinfection by analysing msp-1 and msp-2 genes of Plasmodium falciparum in treatment failure cases.
Methods. We carried out an in vivo study in the year 2005 in 206 children between 6 to 59 months age groups. Of the 206, 120 received AQ + SP, and 86 received AS + SP. A clinical and parasitological followup during 14 days was undertaken. Finger-prick blood sample from each patient was taken on Whatman filter paper (no. 3) on days 0, 7, 14 and also the day when the parasite and symptoms reappeared for PCR analysis.
Results. Late treatment failure was observed in 3.5% (4/114) with AQ + SP, and 2.5% (2/79) with AS + SP. The success rate was 96.5% with AQ + SP and 97.5% with AS + SP. No deaths and severe reactions were recorded. Out of the 6 treatment failure cases, one was reinfection as observed by PCR analysis of msp-1 and msp-2 genes on day 14.
Discussion. Both the combinations found to be efficacious and safe and could be used as a first-line treatment for uncomplicated falciparum malaria in Equatorial Guinea.


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## Body

## 1.Background

Malaria is the major cause of morbidity and mortality
in children under five years in Sub-Saharan Africa [1]. In areas
of stable malaria transmission, like Equatorial Guinea, 25–40% of the
patients in clinics and 20–50% of hospital admissions are due to malaria. The
falciparum malaria is the cause of at least 20% of death in fewer than five
years old children [1].In Sub-Saharan Africa, the mortality in children under
five years old during the period between 1990 and 1998 was doubled compared
with the period from 1982 to 1989. One of the most important reasons of this
increase in the mortality rates is the spread of antimalarial drug resistance in Africa [2].The prevalence of malaria infection in children less
than five years is 22% and 48% in Malabo
(Island Region) and in Bata (Continental Region). The more frequent species in Malabo
arePlasmodium
falciparum (92%), P. malariae (3%), P.
ovale (1%),  and mixed infection (4%); in Bata are Plasmodium
falciparum (97%), P. malariae (2%),  and mix
infection (1%) (unpublished data).A lot of African countries have the necessity of
review its malaria treatment. In Equatorial Guinea, as per the
National Malaria Program, chloroquine (CQ) is the first line of treatment,
whereas quinine is for treatment of severe malaria. Due to the initiative of
Medical Care International Development (MCDI), The Equatorial Guinea Ministry
of Health has started to use the combination of AS + SP as a first line treatment
of uncomplicatedfalciparum malaria in children and pregnant women since 2004 in Bioko Island.In different in vivo studies carried out in
Equatorial Guinea (Bioko Island) by the Institute of Health Carlos III (Madrid,
Spain), in collaboration with the Ministry of Health of this country from 1992
to 1999, it was concluded that resistance to cloroquine (CQ) was over 40% in
children under five, and resistance to sulphadoxine-pyrimethamine (SP) was
around 25% in the same age group. In the continental area of the country,
studies undertaken during 2001  gave the same results as in the island [3].During the years 2002 and 2003, the sensitivity ofPlasmodiumfalciparum to AS + SP combination was
evaluated in Equatorial
Guinea
(unpublished study),  and an adequate
clinical response of 95% was achieved.As a result of the problem of drug resistance
developed inP. falciparum, the Word Health Organization (WHO)
recommended to improve access of population to a quick and efficacy treatment
to decrease the burden of malaria [4].
The current recommendation is the use of a combination therapy following the
experience of treatments in other diseases, such as tuberculosis, leprosy,  and
HIV infection. A combination therapy would be helpful in simultaneous use of
two or more blood schizontocidal drugs with independent modes of action and
different biochemical targets in the parasite
[5].Present study was undertaken jointly with the Malaria
National Control Program for evaluation of AS + SP and AQ + SP combinations in
treatment of uncomplicatedfalciparum malaria. AQ + SP combination is presently
recommended by the WHO, if the efficacy is acceptable [6]. It is less expensive than a
combination with artesunate and can be used as a first line of treatment keeping
in view the economic resources of the country rather than introducing directly
a combination with artesunate. The second one  AS + SP is being used in the Island,  and its efficacy needs to be monitored for its
further use in the Malaria National Control Program.For any in vivo study, it is necessary to differentiate
recrudescence or reinfection, when the country is having high incidence of
malaria and with a high transmission rate. For this reason, polymorphic markers
asmsp-1 and msp-2 or microsatellite markers are useful for this
purpose [7].In this study, we evaluated the efficacy of the AS + SP
and AQ + SP combinations as  a first-line treatment for uncomplicated malaria in
Equatorial Guinea, and the outcome of infection and treatment was determined by
molecular analysis to present a real scenario on use of these two combinations.


## 2. Material and Methods

### 2.1. Area and Study Population

Equatorial Guinea is situated in Central Africa, in the Guinea Gulf.
It is divided in two regions, the continental area, called Rio Muni between Cameroon and Gabon,
and the Island Region (Bioko, Annobón and Corisco Valley).
Bioko, the biggest island, is placed 40 km from Cameroon
coast. Population
estimation is around 484 000 habitants in an area of 28 051km2 between both regions
[8].
Around 75% of the population lives in the Continental Region.Malabo is the capital of the
country and is placed in the Island of Bioko. There is a
tropical climate with a rainy season from May to October and a dry period from
December to March. It is a mesoendemic transmission area with a Plasmodium
index of 21.7% in 2005 [9].
Bata is the most important city in the continental region and has also a
tropical climate with two dry seasons (December–March, June–September) and two
rainy seasons (March–June, September–December) alternate. It is a mesohyperendemic
area with a Plasmodium index between 41% and 75% in children under five years
old.  (Unpublished study from a National
Prevalence Survey 2006, Ministry of Health and Instituto de Salud Carlos III.)In
December 2005, an in vivo study was carried out in the two Regional
Hospitals of Malabo
and Bata. The study followed the WHO methodology [10]. The cases that were included in
the assessment came from the paediatric external consultation.It
was a two-armed prospective evaluation of the clinical and parasitological
responses to different antimalarial combinations, as AS + SP and AQ + SP. The
minimal sample size was different depending on the combinations. In order to
have a confidence level of 95% and a precision of 10% and as the proportion of
treatment failure with AS + SP was already known, a minimum of 50 patients had to
be included. With AQ + SP, there was no evidence of treatment failure, so the
minimum sample size had to be 96 for having a precision of 10% with a
confidence level of 95% [10].


### 2.2. Procedures, Treatment and Followup

All
the children were medically screened. The inclusion criteria were age between
6 to 59 months, no signs of severe malnutrition, a slide confirmed
monoespecific infection ofP.
falciparum, parasite density between 2000 and 200 000 asexual
parasites/microlitre, axillary temperature >37.5°C, easy access to the
hospital, absence of history of hypersensitivity reactions to any of the drugs
being evaluated and informed consent provided by parents or caregivers.  Any danger signs of severe malaria such as
inability to drink or breastfeed, vomiting, recent history of convulsions,
lethargy, and inability to sit or stand up were considered as exclusion
criteria.All
the children were randomly assigned to one therapeutic arm. They were evaluated
clinically and parasitologically during 14 days; treatment was given during
the first three days under direct observation.A
record form, which included: age, sex, address, name of the caregiver, contact
telephone (if available), was maintained. During the followup, parameters such
as temperature (during the six days), doses of drug (days 0, 1, and 2),
parasitaemia (days 0, 1, 2, 3, 7, and 14 or whatever day that children was bringing
to the hospital), haematocrit (days 0 and 14) and filter paper (days 0, 14,  or any
day in case of treatment failure for analysis of the molecular markers) were
recorded. Finger prick blood samples that were collected in Whatman paper and in a thick-and-thin smear stained
with Giemsa for microscopy exam were undertaken.Parasitaemia
was quantified by a standard approximation method, that is,  number of asexual
parasites seen per 200 white blood cells in a high-power examination of a thick
blood film. A positive smear was defined as the presence of at least one
asexual parasite seen after examining 100 thick smear fields. Quality control
of blood smears was done by rereading 10% of slides selected randomly.
Discordant results were subjected to a third microscopist. Haematocrit was
measured by microhaematocrit centrifugation on days 0 and 14.Drugs used were amodiaquine 150 mg (Holden Medical, set. 04L02. Expired 11/2007), artesunate 50 mg (Action medeor. Lot.
055585. Expired 11/2008), sulphadoxine 500 mg + pyrimethamine 25 mg (IDA. Lot. SPF-200 Expired 11/2008), quinine 200 mg (Holden
Medical. Lot MFF 371/05 A 01. Expired 12/2007).Amodiaquine was given as 10 mg/kg of body weight per
day for three days. Artesunate given as 4 mg/kg of body weight per day for three
days and sulphadoxine-pyrimethamine given as sulphadoxine 500 mg + pyrimethamine
25 mg/kg of body weight in a single dose  for the first day. In case of treatment
failure, quinine at 10 mg/kg of body weight every 8 hours for seven days was
administered.


### 2.3. Definition of Treatment Failure

The efficacy outcome was measured based on
parasitological clearance rates on day 14. The criteria to determine the
treatment failure were the following: early treatment failure (ETF) was
considered if development of severe malaria or danger signs during days 1, 2, or
3 in presence of parasitaemia, or parasitaemia on day 2 is higher than day 0
irrespective of axillary temperature, or parasitaemia on day 3 with axillary
temperature>37.5°C, or parasitaemia on day 3 more than 25% of day 0.Late clinical failure (LCF) was considered by
development of severe malaria or danger signs after day 3 with parasitaemia
without previous criteria of ETF, or parasitaemia and temperature>37.5°C on any day between 4
and 14 without previous criteria of ETF. Late parasitological failure (LPF) was
determined based on the presence of parasitaemia between days 7 and 14 with
temperature >37.5°C,
without previous criteria of ETF or LCF. Adequate clinical and parasitological response
was determined (ACPR) by the absence of parasitaemia on day 14, irrespective of
axillary temperature, without meeting the criteria of ETF, LCF, and LPF.Some of the cases were considered as Withdrawal, when
caregivers of children decided not to continue with the study despite all
efforts and lost to follow up; children could not be found at hospital, in the
community, or in the study area.


### 2.4. DNA Extraction and Molecular Study

We extracted DNA from individual blood sample collected
on Whatman filter
paper. We cut a circle of paper (40 mm Ø) with blood, and DNA was extracted by Phenol/Chloroform
method. This DNA was used for the different molecular assays: (a) semi-nested-Multiplex PCR for the diagnosis of Malaria to confirm thePlasmodium species (b) study of the msp-1 and msp-2 genes of P.
falciparum through nested-PCR [7]. In order to distinguish recrudescence from
reinfection cases, we studied the three different allelic families of msp-1
gene: Mad20, RO33 and K1,  and msp-2 gene. Recrudescence was defined as
the same population of parasites appeared on day 7 or day 14 as in day 0. Reinfection
was defined as a new population of parasites detected on day 14 compared to day
0. After the PCR, we analysed the result in the Bioanalyzer 2100 expert
(Agilent Technologies), this method has a very high resolution and we can
distinguish amplification fragments with a little difference in the basepairs.


### 2.5. Statistical Analysis

Baseline
characteristics, means,  and percentages were compared in study subjects of both areas
by Studentt-test and by chi-square test respectively. The non-parametric
alternative for t-test and  Mann-Whitney test was used to compare the baseline
parasite density. Therapeutic efficacy was calculated by dividing the number of
ACPR cases by the number of patients evaluated on 14th day. The 95% confidence
intervals for drug efficacy were also calculated. Log-rank test for equality of
survivor functions was calculated in order to compare time to parasitological
clearance.


## 2.1. Area and Study Population

Equatorial Guinea is situated in Central Africa, in the Guinea Gulf.
It is divided in two regions, the continental area, called Rio Muni between Cameroon and Gabon,
and the Island Region (Bioko, Annobón and Corisco Valley).
Bioko, the biggest island, is placed 40 km from Cameroon
coast. Population
estimation is around 484 000 habitants in an area of 28 051km2 between both regions
[8].
Around 75% of the population lives in the Continental Region.Malabo is the capital of the
country and is placed in the Island of Bioko. There is a
tropical climate with a rainy season from May to October and a dry period from
December to March. It is a mesoendemic transmission area with a Plasmodium
index of 21.7% in 2005 [9].
Bata is the most important city in the continental region and has also a
tropical climate with two dry seasons (December–March, June–September) and two
rainy seasons (March–June, September–December) alternate. It is a mesohyperendemic
area with a Plasmodium index between 41% and 75% in children under five years
old.  (Unpublished study from a National
Prevalence Survey 2006, Ministry of Health and Instituto de Salud Carlos III.)In
December 2005, an in vivo study was carried out in the two Regional
Hospitals of Malabo
and Bata. The study followed the WHO methodology [10]. The cases that were included in
the assessment came from the paediatric external consultation.It
was a two-armed prospective evaluation of the clinical and parasitological
responses to different antimalarial combinations, as AS + SP and AQ + SP. The
minimal sample size was different depending on the combinations. In order to
have a confidence level of 95% and a precision of 10% and as the proportion of
treatment failure with AS + SP was already known, a minimum of 50 patients had to
be included. With AQ + SP, there was no evidence of treatment failure, so the
minimum sample size had to be 96 for having a precision of 10% with a
confidence level of 95% [10].


## 2.2. Procedures, Treatment and Followup

All
the children were medically screened. The inclusion criteria were age between
6 to 59 months, no signs of severe malnutrition, a slide confirmed
monoespecific infection ofP.
falciparum, parasite density between 2000 and 200 000 asexual
parasites/microlitre, axillary temperature >37.5°C, easy access to the
hospital, absence of history of hypersensitivity reactions to any of the drugs
being evaluated and informed consent provided by parents or caregivers.  Any danger signs of severe malaria such as
inability to drink or breastfeed, vomiting, recent history of convulsions,
lethargy, and inability to sit or stand up were considered as exclusion
criteria.All
the children were randomly assigned to one therapeutic arm. They were evaluated
clinically and parasitologically during 14 days; treatment was given during
the first three days under direct observation.A
record form, which included: age, sex, address, name of the caregiver, contact
telephone (if available), was maintained. During the followup, parameters such
as temperature (during the six days), doses of drug (days 0, 1, and 2),
parasitaemia (days 0, 1, 2, 3, 7, and 14 or whatever day that children was bringing
to the hospital), haematocrit (days 0 and 14) and filter paper (days 0, 14,  or any
day in case of treatment failure for analysis of the molecular markers) were
recorded. Finger prick blood samples that were collected in Whatman paper and in a thick-and-thin smear stained
with Giemsa for microscopy exam were undertaken.Parasitaemia
was quantified by a standard approximation method, that is,  number of asexual
parasites seen per 200 white blood cells in a high-power examination of a thick
blood film. A positive smear was defined as the presence of at least one
asexual parasite seen after examining 100 thick smear fields. Quality control
of blood smears was done by rereading 10% of slides selected randomly.
Discordant results were subjected to a third microscopist. Haematocrit was
measured by microhaematocrit centrifugation on days 0 and 14.Drugs used were amodiaquine 150 mg (Holden Medical, set. 04L02. Expired 11/2007), artesunate 50 mg (Action medeor. Lot.
055585. Expired 11/2008), sulphadoxine 500 mg + pyrimethamine 25 mg (IDA. Lot. SPF-200 Expired 11/2008), quinine 200 mg (Holden
Medical. Lot MFF 371/05 A 01. Expired 12/2007).Amodiaquine was given as 10 mg/kg of body weight per
day for three days. Artesunate given as 4 mg/kg of body weight per day for three
days and sulphadoxine-pyrimethamine given as sulphadoxine 500 mg + pyrimethamine
25 mg/kg of body weight in a single dose  for the first day. In case of treatment
failure, quinine at 10 mg/kg of body weight every 8 hours for seven days was
administered.


## 2.3. Definition of Treatment Failure

The efficacy outcome was measured based on
parasitological clearance rates on day 14. The criteria to determine the
treatment failure were the following: early treatment failure (ETF) was
considered if development of severe malaria or danger signs during days 1, 2, or
3 in presence of parasitaemia, or parasitaemia on day 2 is higher than day 0
irrespective of axillary temperature, or parasitaemia on day 3 with axillary
temperature>37.5°C, or parasitaemia on day 3 more than 25% of day 0.Late clinical failure (LCF) was considered by
development of severe malaria or danger signs after day 3 with parasitaemia
without previous criteria of ETF, or parasitaemia and temperature>37.5°C on any day between 4
and 14 without previous criteria of ETF. Late parasitological failure (LPF) was
determined based on the presence of parasitaemia between days 7 and 14 with
temperature >37.5°C,
without previous criteria of ETF or LCF. Adequate clinical and parasitological response
was determined (ACPR) by the absence of parasitaemia on day 14, irrespective of
axillary temperature, without meeting the criteria of ETF, LCF, and LPF.Some of the cases were considered as Withdrawal, when
caregivers of children decided not to continue with the study despite all
efforts and lost to follow up; children could not be found at hospital, in the
community, or in the study area.


## 2.4. DNA Extraction and Molecular Study

We extracted DNA from individual blood sample collected
on Whatman filter
paper. We cut a circle of paper (40 mm Ø) with blood, and DNA was extracted by Phenol/Chloroform
method. This DNA was used for the different molecular assays: (a) semi-nested-Multiplex PCR for the diagnosis of Malaria to confirm thePlasmodium species (b) study of the msp-1 and msp-2 genes of P.
falciparum through nested-PCR [7]. In order to distinguish recrudescence from
reinfection cases, we studied the three different allelic families of msp-1
gene: Mad20, RO33 and K1,  and msp-2 gene. Recrudescence was defined as
the same population of parasites appeared on day 7 or day 14 as in day 0. Reinfection
was defined as a new population of parasites detected on day 14 compared to day
0. After the PCR, we analysed the result in the Bioanalyzer 2100 expert
(Agilent Technologies), this method has a very high resolution and we can
distinguish amplification fragments with a little difference in the basepairs.


## 2.5. Statistical Analysis

Baseline
characteristics, means,  and percentages were compared in study subjects of both areas
by Studentt-test and by chi-square test respectively. The non-parametric
alternative for t-test and  Mann-Whitney test was used to compare the baseline
parasite density. Therapeutic efficacy was calculated by dividing the number of
ACPR cases by the number of patients evaluated on 14th day. The 95% confidence
intervals for drug efficacy were also calculated. Log-rank test for equality of
survivor functions was calculated in order to compare time to parasitological
clearance.


## 3. Results

Between
August and December 2005, 206 children were randomised from the external
consultation of Malabo and Bata Regional Hospitals
in order to
evaluate the efficacy of two antimalarial combinations: AQ + SP and AS + SP.
Baseline characteristics of children are shown in Table1. As expected, no
differences were found between groups, except in the weight.Table 1
Baseline characteristics.
Amodiaquine  +  sulphadoxine-pyrimethamineArtesunate + sulphadoxine-pyrimethamineSexMale
(%)6253Age in months (mean ± sd)28.2 (15)26.7 (15)Weight  in Kg(mean±sd)*12.7 (4)11.7 (3)Temperature inºC (mean ± sd)38.2 (1)38.3 (1)Parasites/microlitre
(median/IR)23,882 (9,360–30,480)24,978 (20,040–31,640)Haematocrit30 (4)31 (4)P*<.05; $P<.01;
sd (standard deviation); IR (Interquartile rank).The enrolled children were given one of the two
combinations in a random way: 120 were treated with AQ + SP and 86 received AS + SP.
In the first group 95% (114/120) finished the study and 91.9% (79/86) in the
second group. The trial profile appears in the Figure1.Figure 1
Trial profile.There
were 13 children who lost to follow up in AQ + SP group. The percentage of lost
to follow up was 5% (6/120) and in the AS + SP arm was 8.1% (7/86). In general
children who lost to follow up were fewer (6.3%; 13/206) than 10% of the children as per WHO
recommendations.With
AQ + SP, 4 cases of LPF were registered and the proportion of treatment failure
was 3.5% (4/114). The ACPR
was 96.5% (confidence interval 95%: [91.3%–99%]). With AS + SP, 2 cases of LPF 2.5% (2/79) were
registered, the cure rate was 97.5% (C.I. 95%: [91.2%–99.7%]). All of the
failure cases received rescue treatment with oral quinine and cured (see Table2). After the molecular correction, the ACPR changed from 96.5% to 97.3% with
AQ + SP combination.Table 2
Treatment
outcomes observed in the patients of Bata and Malabo Hospitals.
The results appear in the different groups with two combination drugs.
TreatmentTotalAQPRETFLPFLCFNº%Nº%Nº%Nº%amodiaquine + sulphadoxine-pyrimethamine11411096.500.043.500.0artesunate + sulphadoxine-pyrimethamine797797.500.022.500.0Both combinations were well tolerated, no deaths were
notified and no severe reactions were recorded, except 3 patients (2.5%) had
complaints of itching in AQ + SP treatment group, they were given antihistamine. Four
patients (3.3%) in AQ + SP and 1 patient (1.2%) in AS + SP treatment groups vomited,
which could not be confirmed whether this was the consequence of malaria or the
treatment effect.Theparasite
clearance (Figure 2) was faster
in the group who received the combination with artesunate (log-rank test
P<.001). In AS + SP treatment group, 77.2% (61/79) and in AQ + SP treatment
group, 94.7% (108/114) presented parasitemia on day 1. On day 2, 3.8% (3/79) of
AS + SP and 55.3% (63/114) of AQ + SP treatment groups were parasitemic. By day 3,
all of them were negative.Figure 2
Proportion of parasites clearance with the two combinations.Baselinegametocyte carriage was less than 5% in both arms. After receiving
treatment, gametocyte density was found to be higher in AS + SP patients than
AQ + SP. With the first combination, by day 2, 12% of children had gametocytes on
the thick film, compared with the 7.5% with AQ + SP. By day 3, the 7.8% with
AS + SP and the 6% with AQ + SP had gametocytes. By day 14, gametocytemia was under
2% with both the combinations.Haematocrit at enrolment showed that  37%
of the children had moderate anaemia with haematocrit 24–29, 49% had mild
anaemia with haematocrit 30–34,  and 14% were nonanaemic. After treatment the
moderate anaemia decreased from 37% to 23%,  and nonanaemic cases increased from
14% to 22%.Molecular correction was done to distinguish recrudescent from reinfection in 6
treatment failure cases: 4 cases from Bata, 3 in the AQ + SP combination,
and 1 in the AS + SP, and 2 in Malabo
with two
combinations (AQ + SP and AS + SP). Table 3 shows the number of alleles of msp-1 allelic family, and msp-2 detected in the six treatment
failure cases. We could confirm the resistant
cases from Bata by molecular analysis; because the parasite genotypes detected
by PCR on day 14 were similar with those on day 0. However, in the AQ + SP treatment
failure cases from Malabo,
one of them was reinfection because a different set of genotypes appeared on
day 14 as shown in Table 3.Table 3
Number of alleles ofmsp-1 allelic
family, and the alleles detected in msp-2.
In case of isolate 293,  new populations appeared on day 14, one for RO33 (150 bp)
and two for msp-2 (600 bp and 700 bp).
msp-1LocalityTreatmentK1MAD20RO33msp-236D0BataAQ/SP135/155/22500570/75536D141550011D0BataAQ/SP190/205/290150500/530/59011D14500102D0BataAQ/SP210/250/265195/210/240150535/590102D1415075D0BataAS/SP160/220/250150575/67075D1415029D0MalaboAQ/SP135/225490/525/66529D14150600 / 7002D0MalaboAS/SP210/250/265195/210/240150535/5902D14****


## 4. Discussion

There is a need to give a thought about any monotherapy as a
first-line treatment in case of patients report with uncomplicatedfalciparum malaria. Keeping this in view, present study was undertaken on the
drug sensitivity of two different therapeutics combinations. This study would
be helpful in National Malaria Control Program of Equatorial Guinea to
introduce a new combination therapy for malaria treatment.Two combinations were assessed, the combinations AS + SP, where
the AS is an efficacious drug and reduces the parasitaemia faster than others,
and the second combination, AQ + SP, is also an effective combination but with the
risk to develop resistant in due course of time, but more accessible to the
population when compared with artemisinin derivatives.The study demonstrated that both combinations are safe and
efficacious for the treatment of nonseverefalciparum malaria in Equatorial Guinea.  These results are similar to other studies made
in different African's areas. A study carried out in Tanzania
in an area of high malaria
transmission compared the efficacy and safety of AQ and SP in monotherapy and
AQ + SP as a combination. The study demonstrated that AQ + SP combination was safe,
and the combination of the two drugs had higher efficacy (96.2%) than
monotherapy [11].
In a similar study carried out in Gambian children, concluded that AS + SP
combination was safe and the efficacy was 98.4% [12].
A study carried out in Guinea
showed an efficacy of 99% with this combination [13].
These results are similar to those of the present study with cure rates of
97.5% (77/79) in
AS + SP combination and 97.3% (111/114) in AQ + SP. The
situation in Equatorial
Guinea
is similar with other African
countries. The cure-rates with the two combination drugs, AQ + SP and AS + SP,  are
over the 95%, which is the limit that WHO suggested for a combination to be
used as a first line of treatment [6].In general terms, drugs were well tolerated, with a low
percentage of adverse effects. In the group that received AQ + SP, a small number
of children required retreatment after vomiting after treatment. Severe
reactions with amodiaquine as neutropenia have been described also in other
publications. In one study, it was shown that the risk was associated with amodiaquine used as a weekly
prophylaxis, not like a treatment [14].
In a study carried out in Madagascar, using combination of AQ + SP like our work,
no severe reaction was described using AQ in a 3-day treatment schedule [15].Clearance of parasites was faster in case of combination
with artemisinin derivatives, due to the mode of action of artemisine, which
reduces parasitaemia by a factor of 10 in each cycle. In Uganda, was
described the difference in parasitemia clearance using a combination with and
without artemisinin derivatives. By day two, in the first case just 3% were
positive compared to the second group, where 48% were positive [16].
These results coincide with our study, where by day two, 3.8% were parasitaemic
compared with the 55.3% of children who received AQ + SP on the same day. Due to faster
parasite clearance, clinical improvement and disappearance of symptoms were
faster in children treated with AS + SP. Because of this rapid effect, use of
artemisinin derivatives in a combination would be appropriate in terms of
decreasing the possible development of parasite resistance to the drug by rapid
clearance thus they are not in contact with the drug for a long time [17].Contrary to other studies, we have not seen any
difference in gametocytes carriage. From this study, we could not say that using
of artemisinin-based combination therapies may reflect in decrease of malaria
transmission in the area. One study in Tanzania, where different drugs in
monotherapy and combinations were compared, demonstrated that  AQ + SP
gametocytes on day 14 were  25.7% in children, which was double than using
the combination with AS, where just 11.9% of children had gametocytes [18].Both the combinations had SP.  It is necessary to assess other therapies
without this drug due to the following reasons. First, SP is the only safe drug
actually to be used as Intermittent Preventive Treatment for pregnant women.
Following WHO recommendations, we need to protect the future development of
resistance to this drug. And the second reason is that SP has a resistance
around 18% in monotherapy (unpublished data, 2002). Drugs that have been used for a long time as monotherapy and have
already demonstrated resistance have the risk to develop resistance in
combination in due course of time.From the results of this study, we may suggest that in
all in vivo studies a molecular assay for discriminating recrudescence
from reinfection should be done. The molecular correction is the only method
that allows us to give a real data about the level of resistance detected in an
endemic area. This way, it is possible to analyse different markers likemsp-1 and  msp-2 genes of P. falciparum, or other polymorphic markers
as microsatellites [19].We need to continue the epidemiological surveillance
and monitoring resistance in order to give enough information to the National Malaria
Program of Equatorial Guinea for revising malaria treatment policy in the
country. This should be done as per the recommendation of WHO before the drug resistance
reaches at levels not allowed to be used.The conclusions of this study are that  both the combinations are safe and efficacious
for their use as a first-line treatment in case of nonseverefalciparum malaria. To choose one of
them will depend on economic resources of Equatorial Guinea in the moment to
change Malaria Policy of the country; the molecular correction gives
complementary information to strengthen the findings of in vivo studies carried
out in areas with a high level of transmission.



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*Source: 781865-2009-05-27.xml*