**1. Introduction**

Malaria is a protozoan infection (Najera &Hempel, 2006 as cited in Okwa & Ibidapo, 2010) with protean manifestations in the human species (Mohaptra, 2002; Murthy, 2000; Talib, 1996) causing nearly one million deaths mainly in African children and decreasing gross domestic product by as much as 1.3% in countries with high disease rates (World Health Organization [WHO], 2010). Approximately half of the world's population is at risk of malaria and in 2008; malaria was present in 108 countries and territories of the world (WHO, 2010). The most specific at risk population groups include young children in stable transmission areas, non immune pregnant women, semi immune pregnant women irrespective of HIV status, people with HIV/AIDS, international travelers to malaria endemic from non endemic areas as well as immigrants from endemic areas and their children living in non endemic areas returning to their home countries to visit friends and relatives (WHO, 2010).

Making a diagnosis requires careful clinical examination and laboratory investigation. Whereas malaria could be over diagnosed in endemic areas, (Ammah et al, 1999; Gwer et al, 2007; Hussain et al, 2009; Rehlis & Kurczewska, 2001; Rougemont et al, 2003; Smith et al, 1994;) in the non endemic areas a high index of suspicion is usually required (Berrang –Ford et al, 2008). However, in the most vulnerable: neonates, under fives, (Dzeing-Ella et al 2005) pregnant women, the elderly and non immune( Sengoz inan et al,2010) who may develop potential life threatening complications of *falciparum* malaria, it is important in most cases to make a rapid, accurate diagnosis to ensure prompt treatment. WHO recommends that before giving treatment, clinical malaria should be confirmed by parasite –based diagnosis. Treatment given solely on the basis of symptoms (presumptive diagnosis and treatment) should only be considered when a parasitological diagnosis is not possible. In 2008, 33 of 43 malaria endemic countries in the African region and 45 out of 63 countries in other regions were reported to have developed a policy of parasitological testing of suspected malaria cases in persons of all ages. However, policy development has not matched actual practice. Parasitological test for suspected malaria cases is carried out in less than 20% of individuals living in 21 of the highest disease burden countries.(WHO, 2010).

Current Issues in Clinical and Laboratory Diagnosis in Malaria 163

methods, Kamwenedo et al (2002) also working in Malawi had demonstrated that cord blood genotypes were a subset of the maternal and placental blood. An issue of current interest relating to the integrity of the placenta has been the role of HIV infection. In several recent studies (Steketee,2004; Ticconi et al, 2003; van Ejik et al, 2003; Verhoeff et al, 1999;), it has been found that HIV impaired the ability of pregnant women to control malaria parasitemia. Results from these studies showed that HIV-infected women experienced consistently higher prevalence of peripheral and placental malaria (summary relative risk = 1.58 and 1.66, respectively), higher parasite densities, and more febrile illnesses, severe anemia, and adverse birth outcomes when compared with HIV-uninfected women, particularly in multigravidae. Thus, HIV alters the typical gravidity-specific pattern of malaria risk by shifting the burden from primarily primigravidae and secundigravidae to all

Maternal, especially perinatal malaria is a significant risk factor for congenital malaria. Intermittent preventive therapy (IpT) is recommended by WHO for pregnant women and infants in areas of high transmission in Sub Saharan Africa with stable malaria transmission who are particularly vulnerable to the consequences of malaria. This prophylaxis when adequately taken as two doses at least one month apart substantially reduces both maternal and neonatal morbidity and mortality related to malaria. However, the progress report on malaria prevention (WHO, 2010) indicates that coverage with Intermittent Preventive treatment for pregnant women ( IPTp) has remained far below the target levels. Thirty three out of 43 endemic countries in Africa had adopted the IPTp programme by 2009. Only 55% of all women attending antenatal clinics received the second dose of IPTp and since not all women attend antenatal clinic, household surveys in some countries report 2.4% to 62% with an overall weighted average of 12% received the second dose of therapy. This presents a picture that calls for urgent action to reduce maternal malaria and its consequences.

The major clinical features in congenital are fever within the first 24 hours of life, refusal to suck and anaemia (Orogade et al, 2008). Fever is the most consistent feature while some other studies have described hepatosplenomegaly, jaundice, irritability. These are indistinguishable from features commonly seen in neonatal septicaemia. The implication of this is that every febrile neonate born to a mother who is epidemiologically vulnerable for

There have been variable findings in prevalence of malaria parasitemia in this age group, ranging from 0% in children less than 3 months (Okwa, 2000) to 17.2 % in the first six months of life (Orogade, 2006) and 27.1% in a similar series by Afolabi et al (2001). Age specific parasite rates showed a sharp drop in frequency of parasitemia from first to second week of life (Orogade, 2006), but this gradually increases with age till the sixth month of life. This initial sharp drop has been attributed to spontaneous clearance of parasites which

malaria should be screened for malaria as well as other bacterial and viral infections.

pregnant women.

**2.2 Zero – Five months**

**2.1.3 Implication for malaria prophylaxis in pregnancy** 

**2.1.4 Major clinical features in congenital malaraia** 

**2.2.1 Trend of malaria parasitaemia with age** 

Moreover, the field of malaria diagnosis is rapidly expanding and bringing to fore hitherto unidentified issues both clinically and in the laboratory. Malaria, once thought to be rare in the newborn (Molyneux, 1989) has in recent times been increasingly reported (Falade et al, 2007; Ibanesebor, 1995; Kamwendo et al, 2002; Lamikanra,1993; Lehner & Andrews, 1988; Muktar et al, 2006; Olowu et al, 2000; Opare,2010; Runsewe –Abiodun et al, 2006; Sowunmi et al, 1996) Incidences range from 0.3 to 33%in these areas ( Fisher, 2003) and has brought into question the extent of protection for the baby in the face of maternal placental infections such as HIV. Malaria infection is also described in the first 6 months of life which may be clinically indistinguishable from common bacterial and viral infections (Orogade, 2006).The use of clinical algorithms proposed for malaria diagnosis and their role as predictors for morbidity and mortality are being investigated (Tabitha et al, 2005). With the introduction of rapid diagnostic tests (RDTs) for diagnosis in malaria, there should be less need for presumptive diagnosis based solely on clinical features in high burden areas. However, there is a growing need to monitor and assess the performance of the RDTs in face of varied availability of RDTs and their low specificity and sensitivity in mixed malaria parasite infections. This chapter reviews these current issues in malaria diagnosis and discusses their implication for prompt malaria identification and treatment which are key aspects of the WHO policy and strategy for global malaria control.

### **2. Clinical features in malaria**

#### **2.1 Newborn**

#### **2.1.1 Prevalence of congenital malaria**

Congenital malaria is defined as the presence of malaria parasites in the peripheral smear of the newborn within the first week of life (Mc Gregory, 1986). It had been thought to be rare due to the known effectiveness of placental barrier but recent reports from malaria endemic areas of incidence in small numbers were made. In a national multicenter study carried out to determine the epidemiology of congenital malaria in Nigeria (Falade et al, 2007), 1,875 babies were assessed within the first 4 hours of life for parasitemia using freshly prepared Giemsa kept at pH 7.2 to stain conventional thick and methanol fixed thin blood smears. A prevalence of 5.1% of patent parasitemia was obtained. It was observed that the mean parasite density in these neonates was low (8-200/µl) as was also reported in an earlier study by Mc Guiness et al (1998). In about two thirds of these babies with parasitemia observed in the first 4 hours of life there was spontaneous parasite clearance by the second day of life, while 33.7% of them persisted and babies became symptomatic within the first 3 days postpartum.

#### **2.1.2 Relationship between malaria in pregnancy and congenital malaria**

Antepartum maternal and placental parasitemia have been identified as consistent risk factors for congenital malaria (Orogade et al 2004,2008). Neonates in these studies who had peripartum malaria parasitemia had a 20 fold increased risk of infection. In Papua New Guinea, Lehner et al(1990) observed that where there was a maternal antepartum parasitaemia of 29.4%, there were corresponding cord and neonatal blood parasitemia of 14.6% and 7.7%. They found a significant correlation between anti malarial IgG antibodies in paired maternal and cord blood which indicated transplacental transfer. Using the PCR

Moreover, the field of malaria diagnosis is rapidly expanding and bringing to fore hitherto unidentified issues both clinically and in the laboratory. Malaria, once thought to be rare in the newborn (Molyneux, 1989) has in recent times been increasingly reported (Falade et al, 2007; Ibanesebor, 1995; Kamwendo et al, 2002; Lamikanra,1993; Lehner & Andrews, 1988; Muktar et al, 2006; Olowu et al, 2000; Opare,2010; Runsewe –Abiodun et al, 2006; Sowunmi et al, 1996) Incidences range from 0.3 to 33%in these areas ( Fisher, 2003) and has brought into question the extent of protection for the baby in the face of maternal placental infections such as HIV. Malaria infection is also described in the first 6 months of life which may be clinically indistinguishable from common bacterial and viral infections (Orogade, 2006).The use of clinical algorithms proposed for malaria diagnosis and their role as predictors for morbidity and mortality are being investigated (Tabitha et al, 2005). With the introduction of rapid diagnostic tests (RDTs) for diagnosis in malaria, there should be less need for presumptive diagnosis based solely on clinical features in high burden areas. However, there is a growing need to monitor and assess the performance of the RDTs in face of varied availability of RDTs and their low specificity and sensitivity in mixed malaria parasite infections. This chapter reviews these current issues in malaria diagnosis and discusses their implication for prompt malaria identification and treatment which are key aspects of the

Congenital malaria is defined as the presence of malaria parasites in the peripheral smear of the newborn within the first week of life (Mc Gregory, 1986). It had been thought to be rare due to the known effectiveness of placental barrier but recent reports from malaria endemic areas of incidence in small numbers were made. In a national multicenter study carried out to determine the epidemiology of congenital malaria in Nigeria (Falade et al, 2007), 1,875 babies were assessed within the first 4 hours of life for parasitemia using freshly prepared Giemsa kept at pH 7.2 to stain conventional thick and methanol fixed thin blood smears. A prevalence of 5.1% of patent parasitemia was obtained. It was observed that the mean parasite density in these neonates was low (8-200/µl) as was also reported in an earlier study by Mc Guiness et al (1998). In about two thirds of these babies with parasitemia observed in the first 4 hours of life there was spontaneous parasite clearance by the second day of life, while 33.7% of them persisted and babies became symptomatic within the first 3

**2.1.2 Relationship between malaria in pregnancy and congenital malaria** 

Antepartum maternal and placental parasitemia have been identified as consistent risk factors for congenital malaria (Orogade et al 2004,2008). Neonates in these studies who had peripartum malaria parasitemia had a 20 fold increased risk of infection. In Papua New Guinea, Lehner et al(1990) observed that where there was a maternal antepartum parasitaemia of 29.4%, there were corresponding cord and neonatal blood parasitemia of 14.6% and 7.7%. They found a significant correlation between anti malarial IgG antibodies in paired maternal and cord blood which indicated transplacental transfer. Using the PCR

WHO policy and strategy for global malaria control.

**2. Clinical features in malaria**

**2.1.1 Prevalence of congenital malaria** 

**2.1 Newborn** 

days postpartum.

methods, Kamwenedo et al (2002) also working in Malawi had demonstrated that cord blood genotypes were a subset of the maternal and placental blood. An issue of current interest relating to the integrity of the placenta has been the role of HIV infection. In several recent studies (Steketee,2004; Ticconi et al, 2003; van Ejik et al, 2003; Verhoeff et al, 1999;), it has been found that HIV impaired the ability of pregnant women to control malaria parasitemia. Results from these studies showed that HIV-infected women experienced consistently higher prevalence of peripheral and placental malaria (summary relative risk = 1.58 and 1.66, respectively), higher parasite densities, and more febrile illnesses, severe anemia, and adverse birth outcomes when compared with HIV-uninfected women, particularly in multigravidae. Thus, HIV alters the typical gravidity-specific pattern of malaria risk by shifting the burden from primarily primigravidae and secundigravidae to all pregnant women.

## **2.1.3 Implication for malaria prophylaxis in pregnancy**

Maternal, especially perinatal malaria is a significant risk factor for congenital malaria. Intermittent preventive therapy (IpT) is recommended by WHO for pregnant women and infants in areas of high transmission in Sub Saharan Africa with stable malaria transmission who are particularly vulnerable to the consequences of malaria. This prophylaxis when adequately taken as two doses at least one month apart substantially reduces both maternal and neonatal morbidity and mortality related to malaria. However, the progress report on malaria prevention (WHO, 2010) indicates that coverage with Intermittent Preventive treatment for pregnant women ( IPTp) has remained far below the target levels. Thirty three out of 43 endemic countries in Africa had adopted the IPTp programme by 2009. Only 55% of all women attending antenatal clinics received the second dose of IPTp and since not all women attend antenatal clinic, household surveys in some countries report 2.4% to 62% with an overall weighted average of 12% received the second dose of therapy. This presents a picture that calls for urgent action to reduce maternal malaria and its consequences.

#### **2.1.4 Major clinical features in congenital malaraia**

The major clinical features in congenital are fever within the first 24 hours of life, refusal to suck and anaemia (Orogade et al, 2008). Fever is the most consistent feature while some other studies have described hepatosplenomegaly, jaundice, irritability. These are indistinguishable from features commonly seen in neonatal septicaemia. The implication of this is that every febrile neonate born to a mother who is epidemiologically vulnerable for malaria should be screened for malaria as well as other bacterial and viral infections.

## **2.2 Zero – Five months**

### **2.2.1 Trend of malaria parasitaemia with age**

There have been variable findings in prevalence of malaria parasitemia in this age group, ranging from 0% in children less than 3 months (Okwa, 2000) to 17.2 % in the first six months of life (Orogade, 2006) and 27.1% in a similar series by Afolabi et al (2001). Age specific parasite rates showed a sharp drop in frequency of parasitemia from first to second week of life (Orogade, 2006), but this gradually increases with age till the sixth month of life. This initial sharp drop has been attributed to spontaneous clearance of parasites which

Current Issues in Clinical and Laboratory Diagnosis in Malaria 165

The major clinical symptoms apart from fever were cough, diarrhea generalized rash, excessive crying and vomiting ( Afolabi, 2001; Orogade, 2006). All these symptoms are non specific but had about one half to two fold occurrence in children with malaria than in others without malaria parasitemia. Fever however seems to be the most constant factor even in the older infant and children. The babies had a low mean temperature of 37.7°C ± 0.58 and respiratory distress with respiratory rate of 45.66 ± 21.6 cycles/minute. Dehydration, palmar pallor, jaundice and hepatomegaly were the commonest signs though not of significant relationship. Palmar pallor is not a good indicator of anaemia χ2 = 1.24, p= 0.264. The children with parasitemia had relatively low grade fever with mean temperatures of 37.7 ± 0.58 and tachycardia, mean pulse rate of 137.67 ± 9.94..Overall most of the clinical features were non-specific and could not be attributable to only malaria. Malaria in the children aged 0-5 months though the prevalences and parasite densities are lower, yet it produces significant morbidity in the children. Prevention of malaria in the pregnant women by chemoprophylaxis is still an area that requires focus and advocacy and public enlightenment on the use of insecticide treated nets for more widespread use is needed.

In the areas of stable malaria transmission, this group of children are the most affected in morbidity and mortality. Reporting the findings of severe malaria in Gabonese children, children Dzeing-Ella et al (2005) observed that most children with severe malaria are under 5 years old. Commonest features were anaemia, respiratory distress, cerebral malaria hypoglycaemia. Anaemia was commoner in children under 18 months of age, while cerebral malaria was commoner above 18 months. Poor prognostic factors were coma, hyperlactaemia and hypoglycaemia. Another study reporting uncomplicated malaria in febrile under 5 years children ( Ikeh & Teclaire, 2008) showed prevalences of about 52.2% and the most common presentation was fever. Most of the children within this age group at are various levels of developing immunity and yet have parasite rates that could range from

**2.3.2 Use of clinical algorithms and predictors for malaria morbidity and mortality** 

Development of clinical algorithms were initially done as guidelines to ensure that the young child at risk of potentially fatal diseases were identified and received prompt attention and commenced some management at the community level. In this guideline, the Integrated Management of Childhood Diseases [IMCI], children under 5 years of age, living in areas of high malaria endemicity were to be treated for malaria if they presented to a health facility with fever, temp >37. 5C. Studies by Tabitha et al (2005) found in a study that using a set of symptoms and signs with highest sensitivity and specificity and comparing these to parasitemia, a significant proportion of patients would have been sent home untreated. This tendency increased with increasing age. In situations as alluded to by the report of Khan et al (2005), the other issue was that often there were co-morbidities in febrile illness in some communities. Simply using algorithms for treatment of malaria might also lead to delay in treatment of other equally life threatening infections like Enteric fever which

**2.2.2 Clinical effects of parasitemia** 

**2.3 Six months – Five years** 

**2.3.1 Clinical features of complicated and uncomplicated malaria** 

80-90%. This explains their potential to develop severe malaria.

occurs in some babies at this time. It is noteworthy that about one fifth of all parasitemia occurs in the first month of life. The neonatal age group in this study formed a large burden of the disease prevalence. Mothers who were gravida 1-2, who did not utilize chemoprophylaxis in pregnancy and had education ranging from none to primary school level were important as risk factors for malaria parasitemia in this age group. These same risk factors have also been identified especially in malaria endemic regions where women in their first two pregnancies who develop maternal parasitemia have been identified as high risk for maternal anaemia and fetal complications such as fetal wastages, still births, premature deliveries and low birth weight in the newborn babies. In this study parasitemia documented in the first week of life would likely have resulted from congenital acquisition as prevalence of 9.31% obtained closely compares to the recent congenital malaria reports

There is an average low mean parasite density rate of 64.82 ± 50.61/μL (Orogade, 2006)and this does not vary with age groups, χ2 ( Bartlett's test) = 6.09, p=0.29. The mean Haematocrit is also significantly lower than controls low at 35.62 ± 7.09.%. Analysis of variants revealed a significant positive correlation between the mean haematocrit and malaria parasitemia.

 Use of malaria prevention was 93.6% and this had a significant impact on reduction of parasitemia. Of the methods used for malaria prevention, window/door net screening and insecticides sprays were the most common. Less than half of mothers used bednets for their babies and only 7(2.8% )of these were insecticide treated bednets. Other non conventional methods used were local chemicals (*ota pia-pia*) sprayed on floors and walls in the rooms where the children slept. This apparently significantly reduced parasitemia where it was used one and half fold. Of all the babies seen only 47.4% were being exclusive breast fed.

Beyond the neonatal period, the use of malaria prevention methods in particular *ota pia-pia* were significant for protection against malaria. *Ota pia-pia* which was used in 13.37% of cases seems to have been quite effective. This is a locally prepared chemical which when sprayed on floors and walls in the rooms are effective anti vector agents. However, the exact chemical compositions of these chemicals are yet to be fully analyzed and the safety of their use is not documented. Moreso, their potential harmful effects to the newborn and infant are unknown. Insecticide treated bednets have been introduced in Nigeria as an effective measure of malaria prophylaxis. However Orogade (2006) revealed a very poor utilization of this proven effective measure. Bednets, which has been a preventive measure of long standing use was utilized for less than half of the babies. There was rather more common use of window/door net screens as well as insecticide sprays which have also been known and in use for several years but which are not as effective.

There was low educational status in more than a third of the mothers and this affects how well informed the mothers would be especially about health preventive matters. This less popular use of bednets as well as the level of health information the mothers may comprehend could explain the poor use of treated bednets. Socioeconomic status of families, though not investigated in this study has been found to be a reflection of the choice of malaria preventive measures, in this region. Socioeconomic status determines the economic power to sustain the preventive measure chosen, so cheaper, locally made and available means would be more acceptable.

## **2.2.2 Clinical effects of parasitemia**

164 Malaria Parasites

occurs in some babies at this time. It is noteworthy that about one fifth of all parasitemia occurs in the first month of life. The neonatal age group in this study formed a large burden of the disease prevalence. Mothers who were gravida 1-2, who did not utilize chemoprophylaxis in pregnancy and had education ranging from none to primary school level were important as risk factors for malaria parasitemia in this age group. These same risk factors have also been identified especially in malaria endemic regions where women in their first two pregnancies who develop maternal parasitemia have been identified as high risk for maternal anaemia and fetal complications such as fetal wastages, still births, premature deliveries and low birth weight in the newborn babies. In this study parasitemia documented in the first week of life would likely have resulted from congenital acquisition as prevalence of 9.31% obtained closely compares to the recent congenital malaria reports

There is an average low mean parasite density rate of 64.82 ± 50.61/μL (Orogade, 2006)and this does not vary with age groups, χ2 ( Bartlett's test) = 6.09, p=0.29. The mean Haematocrit is also significantly lower than controls low at 35.62 ± 7.09.%. Analysis of variants revealed a significant positive correlation between the mean haematocrit and malaria parasitemia.

 Use of malaria prevention was 93.6% and this had a significant impact on reduction of parasitemia. Of the methods used for malaria prevention, window/door net screening and insecticides sprays were the most common. Less than half of mothers used bednets for their babies and only 7(2.8% )of these were insecticide treated bednets. Other non conventional methods used were local chemicals (*ota pia-pia*) sprayed on floors and walls in the rooms where the children slept. This apparently significantly reduced parasitemia where it was used one and half fold. Of all the babies seen only 47.4% were being

Beyond the neonatal period, the use of malaria prevention methods in particular *ota pia-pia* were significant for protection against malaria. *Ota pia-pia* which was used in 13.37% of cases seems to have been quite effective. This is a locally prepared chemical which when sprayed on floors and walls in the rooms are effective anti vector agents. However, the exact chemical compositions of these chemicals are yet to be fully analyzed and the safety of their use is not documented. Moreso, their potential harmful effects to the newborn and infant are unknown. Insecticide treated bednets have been introduced in Nigeria as an effective measure of malaria prophylaxis. However Orogade (2006) revealed a very poor utilization of this proven effective measure. Bednets, which has been a preventive measure of long standing use was utilized for less than half of the babies. There was rather more common use of window/door net screens as well as insecticide sprays which have also been known

There was low educational status in more than a third of the mothers and this affects how well informed the mothers would be especially about health preventive matters. This less popular use of bednets as well as the level of health information the mothers may comprehend could explain the poor use of treated bednets. Socioeconomic status of families, though not investigated in this study has been found to be a reflection of the choice of malaria preventive measures, in this region. Socioeconomic status determines the economic power to sustain the preventive measure chosen, so cheaper, locally made and available

and in use for several years but which are not as effective.

means would be more acceptable.

exclusive breast fed.

The major clinical symptoms apart from fever were cough, diarrhea generalized rash, excessive crying and vomiting ( Afolabi, 2001; Orogade, 2006). All these symptoms are non specific but had about one half to two fold occurrence in children with malaria than in others without malaria parasitemia. Fever however seems to be the most constant factor even in the older infant and children. The babies had a low mean temperature of 37.7°C ± 0.58 and respiratory distress with respiratory rate of 45.66 ± 21.6 cycles/minute. Dehydration, palmar pallor, jaundice and hepatomegaly were the commonest signs though not of significant relationship. Palmar pallor is not a good indicator of anaemia χ2 = 1.24, p= 0.264. The children with parasitemia had relatively low grade fever with mean temperatures of 37.7 ± 0.58 and tachycardia, mean pulse rate of 137.67 ± 9.94..Overall most of the clinical features were non-specific and could not be attributable to only malaria. Malaria in the children aged 0-5 months though the prevalences and parasite densities are lower, yet it produces significant morbidity in the children. Prevention of malaria in the pregnant women by chemoprophylaxis is still an area that requires focus and advocacy and public enlightenment on the use of insecticide treated nets for more widespread use is needed.
