**3. Asymptomatic malaria as an important reservoir**

**Country/drug policy\* Year No. of malaria cases % of confirmed cases° No. of death cases**

**China**

Uncomplicated Pf: ART + NQ; AS + AQ; D-P

182 Towards Malaria Elimination - A Leap Forward

Severe malaria: AM; AS; pyronaridine *P. vivax:* CQ + PQ (8d)

**Cambodia** Uncomplicated Pf: AS + MQ, D-P Severe malaria: AM; AS; QN

**Laos**

AL

*P. vivax:* D-P + PQ (14d)

Uncomplicated Pf:

Severe malaria: AS + AL

**Myanmar** Uncomplicated Pf:

**Thailand**

D-P

Uncomplicated Pf:

Severe malaria: QN + doxycycline *P. vivax:* CQ + PQ (14d)

Severe malaria: AM; AS; QN

*P. vivax:* CQ + PQ (14d)

AL; AM; AS + MQ; D-P; PQ

*P. vivax:* CQ + PQ (14d)

**Pf Pv Others**

 3000 41.9 56.6 1.5 ≤100 240 8.2 91.8 — 0 ≤100 64.1 35.9 — 0 ≤100 10.7 89.3 — 0 ≤100 3.0 78.8 18.2 0 ≤10 0.0 100.0 — 0

 203,600 62.6 37.4 — 400 146,000 50.4 49.6 — 220 76,500 45.8 54.2 — 110 89,700 58.8 41.2 — 150 120,300 61.3 38.7 — 210 83,300 58.2 41.8 — 140

 42,800 92.7 7.1 0.2 ≤100 112,700 83.4 16.6 — 250 93,500 67.0 33.0 — 170 117,300 52.9 47.1 — 180 87,900 42.3 57.7 — 120 27,390 39.5 60.5 — ≤100

 1,506,000 68.4 31.6 — 2800 1,974,000 71.8 28.2 — 4000 585,000 70.4 29.6 — 1100 360,000 69.9 30.1 — 700 236,500 64.1 35.9 — 400 142,600 60.3 39.7 — 240

 24,900 40.5 59.5 0.1 ≤100 32,600 39.8 60.2 — ≤100 33,300 44.0 46.8 9.3 ≤100 37,900 37.8 54.1 8.1 ≤100 8000 41.7 58.0 0.2 ≤100 11,520 32.5 46.1 21.5 ≤100 It has long been held as conventional wisdom that asymptomatic infections would be much less frequent in low-endemicity settings because the level of exposure-related immunity to malaria in human populations may be low [25]. However, asymptomatic infections represent the vast majority of infections in all endemic settings [26]. The use of molecular tools is essential for identifying submicroscopic infections. For both *P. falciparum* and *P. vivax*, microscopy detects only 1/3–1/2 of the infections detected by regular PCR [27, 28]. As the sensitivity of detection methods increases (e.g., with the use of a larger blood volume or reverse transcriptase-PCR targeting the parasite 18S rRNA), greater proportions of asymptomatic infections are discovered, revealing larger pools of infections [29, 30]. In Western Thailand and other GMS regions, qPCR and large-volume ultrasensitive qPCR could detect as much as 20% of the villagers harboring malaria infections as compared to ~5% detected by microscopy [31, 32]. Although we still do not have a clear picture about how much these asymptomatic infections actually contribute to malaria transmission in these areas [33], studies in Western Thailand have clearly demonstrated mosquito infectivity of submicroscopic *P. falciparum* and *P. vivax* [34], albeit the asymptomatic parasite carriers were found to be much less infective to mosquitoes than acute cases [35]. Since asymptomatic individuals are unlikely to seek treatment, they are missed by passive case detection, and submicroscopic infections also are missed by microscopy-based active case detection. It is highly possible that these asymptomatic infections act as important silent reservoirs of transmission. Even under such low-endemicity settings, it is estimated that submicroscopic carriers may be the source of 20–50% of all human-to-mosquito transmission [36], underlining the significance of managing this population in the malaria elimination phase. Therefore, information about the prevalence and seasonal dynamics of the asymptomatic infections in the border regions and their contribution to transmission is required to guide the efforts of NMCPs to achieve malaria elimination.

nations still use the lower total dose of PQ in fear of the possible harm to those with G6PD deficiency. Because evaluation of PQ efficacy in preventing relapses requires longer-term followup, the clinical efficacy of the current PQ regimen for radical cure of vivax malaria in the GMS is unknown. Even with longer follow-ups, it is still not possible to reliably determine whether a recurrent infection after day 28 is due to relapse or reinfection given that a relapse infection may be from reactivation of a different hypnozoite clone [64, 65]. For PQ efficacy, host factors also need to be considered. Recently, failures of the PQ radical cure have been linked to reduced activity of the hepatic cytochrome P450 (CYP) 2D6 [66], which mediates activation of PQ to its active metabolite(s) [67, 68]. Different CYP2D6 activities have differential effects on the pharmacokinetics of PQ [69]. CYP2D6 is involved in the metabolism of as many as 25% of drugs in clinical use and is also a member of the CYP450 family with the greatest prevalence and genetic polymorphism [70, 71]. About 70 CYP2D6 allelic variants have been found and grouped into 4 phenotypic classes of ultra-rapid, extensive, intermediate, and abolished protein activity [72]. The frequency of alleles with reduced function is as high as 50% in most Asian populations [73]. Thus, it is important to determine the extent by which reduced CYP2D6 activity is responsible

Malaria Elimination in the Greater Mekong Subregion: Challenges and Prospects

http://dx.doi.org/10.5772/intechopen.76337

185

The *G6PD* gene is extraordinarily polymorphic with more than 400 variants discovered based on biochemical diagnosis [75], among which 186 mutations are associated with G6PD deficiency [76]. The prevalence of G6PD deficiency and distribution of G6PD variants vary geographically [77]. In the GMS, G6PD deficiency is often highly prevalent among ethnic groups. Along the Thailand-Myanmar border, the prevalence of G6PD deficiency was above 10% [78–80], whereas in the Kachin ethnicity along the China-Myanmar border, it almost reached 30% [81]. In Thailand and Myanmar, the Mahidol variant (487G>A) is the most predominant and often accounts for ~90% of all mutations [79, 81–83]. According to the WHO classification, the Mahidol variant is a Class III mutation or mild-deficient variant with 60% enzyme activity [76]. However, this classification may not be accurate since patients with the Mahidol variant often had <1% of the normal G6PD activity [79, 84, 85]. Patients having the G6PD Mahidol variant (487G>A) rarely had acute hemolytic anemia after taking the normal dose of PQ [84, 86]. In contrast to the belief that PQ only induces mild hemolysis in patients with the Mahidol variant, there have been case reports showing that the normal dosage of 15 mg/kg/day for 3 days in vivax patients with this G6PD variant could lead to acute hemolytic anemia that required blood transfusion or even cause renal failure [87–89]. It is noteworthy that G6PD activity can vary substantially between individuals with the same variant and even within the same individual over time. Therefore, with the prevalence of vivax malaria in this region and the goal of malaria elimination, the deployment of point-of-care G6PD deficiency diagnostics is urgent [90]. In addition, there is a need to test whether weekly PQ of 0.75 mg/kg for 8 weeks, a dosage considered safe for the G6PD African

variant [91], could be prescribed in the GMS without prior testing for G6PD deficiency.

ACTs have played an indispensable role in reducing global malaria-associated mortality and morbidity. However, these achievements are threatened by the recent emergence of artemisinin resistance in *P. falciparum* in the GMS [92–94]. Artemisinin resistance is associated with a parasite clearance half-life of >5 h as compared to a normal value of ~2 h [94–96].

**5. Management of drug resistance in** *P. falciparum*

for PQ failures in radical cure of vivax malaria [74].

## **4. The burden of** *P. vivax* **malaria and G6PD deficiency**

Another characteristic of the rapidly evolving malaria epidemiology in the GMS is that the prevalence of *P. vivax* is increasing proportionally to *P. falciparum* [37] (**Table 1**). The resilience of vivax malaria to control efforts may be attributed to some intrinsic biological features of this parasite. First, *P. vivax* only invades reticulocytes, and thus the resulting parasitemia is normally far lower than that of *P. falciparum* malaria. This makes microscopy-based diagnosis and RDTs not sufficiently sensitive in detecting *P. vivax* infections [38–40]. Second, during blood-stage infections with *P. vivax*, gametocytes are formed before the manifestation of clinical symptoms, which allows transmission of the parasite before treatment. Third, *P. vivax* develops dormant hypnozoites in the liver of the human host, which awaken in the weeks and months following a primary attack and cause relapses. Finally, vivax malaria is often transmitted by outdoor biting mosquitoes, making the current insecticide-based control measures (LLIN and IRS) less effective. Because of these unique features, traditional malaria control efforts often fail to control *P. vivax* transmission. In addition, containment of *P. falciparum* has been prioritized in the GMS, partially because of the emerging artemisinin resistance. As a result, *P. falciparum* prevalence has decreased, while the proportion of *P. vivax* has increased.

In the GMS, the first-line therapy for vivax malaria remains chloroquine (CQ) and primaquine (PQ) (**Table 1**) [41]. Reports of clinical CQ resistance in many regions of the world and falling efficacy of PQ are of great concern for vivax malaria control [42–45]. Although some studies indicated that *P. vivax* in the GMS remained sensitive to CQ [46–51], others clearly documented CQ-resistant *P. vivax* [52–55]. In Myanmar, sporadic CQ-resistant *P. vivax* cases were first reported more than 20 years ago [52, 53]. A later report of 34% treatment failures in Dawei of Southern Myanmar suggests an increase of CQ resistance [55]. More recent studies identified both early and late treatment failures in Myawaddy of the Kayin State and Kawthaung of the Tanintharyi Region, Myanmar [56]. In northeastern Myanmar bordering China, a recent study showed 5.2% cumulative incidence of recurrent parasitemia during a 28-day follow-up of 587 *P. vivax* treated with CQ/PQ [57], suggesting sensitivity to CQ may also be deteriorating in this region. This reduced sensitivity of *P. vivax* to CQ requires close surveillance and potential implementation of more effective treatment measures such as ACTs [58].

Studies from Papua New Guinea suggest that 80% of the vivax infections may be attributed to relapses. A modeling approach predicts that as much as 96% of clinical attacks by *P. vivax* in Thailand are due to relapses [60]. For radical cure, WHO recommends a dose of 0.25–0.5 mg/kg of PQ daily for 14 days. However, the lower dose (total of 3.5 mg/kg) fails to prevent relapses in many different endemic sites [61]. Because of the potential risk of severe hemolysis that this drug could cause in patients with glucose-6-phosphate dehydrogenase (G6PD) deficiency, PQ is not widely prescribed [43, 62, 63]. In routine practice, G6PD status is not screened; the GMS nations still use the lower total dose of PQ in fear of the possible harm to those with G6PD deficiency. Because evaluation of PQ efficacy in preventing relapses requires longer-term followup, the clinical efficacy of the current PQ regimen for radical cure of vivax malaria in the GMS is unknown. Even with longer follow-ups, it is still not possible to reliably determine whether a recurrent infection after day 28 is due to relapse or reinfection given that a relapse infection may be from reactivation of a different hypnozoite clone [64, 65]. For PQ efficacy, host factors also need to be considered. Recently, failures of the PQ radical cure have been linked to reduced activity of the hepatic cytochrome P450 (CYP) 2D6 [66], which mediates activation of PQ to its active metabolite(s) [67, 68]. Different CYP2D6 activities have differential effects on the pharmacokinetics of PQ [69]. CYP2D6 is involved in the metabolism of as many as 25% of drugs in clinical use and is also a member of the CYP450 family with the greatest prevalence and genetic polymorphism [70, 71]. About 70 CYP2D6 allelic variants have been found and grouped into 4 phenotypic classes of ultra-rapid, extensive, intermediate, and abolished protein activity [72]. The frequency of alleles with reduced function is as high as 50% in most Asian populations [73]. Thus, it is important to determine the extent by which reduced CYP2D6 activity is responsible for PQ failures in radical cure of vivax malaria [74].

low-endemicity settings, it is estimated that submicroscopic carriers may be the source of 20–50% of all human-to-mosquito transmission [36], underlining the significance of managing this population in the malaria elimination phase. Therefore, information about the prevalence and seasonal dynamics of the asymptomatic infections in the border regions and their contribution to trans-

Another characteristic of the rapidly evolving malaria epidemiology in the GMS is that the prevalence of *P. vivax* is increasing proportionally to *P. falciparum* [37] (**Table 1**). The resilience of vivax malaria to control efforts may be attributed to some intrinsic biological features of this parasite. First, *P. vivax* only invades reticulocytes, and thus the resulting parasitemia is normally far lower than that of *P. falciparum* malaria. This makes microscopy-based diagnosis and RDTs not sufficiently sensitive in detecting *P. vivax* infections [38–40]. Second, during blood-stage infections with *P. vivax*, gametocytes are formed before the manifestation of clinical symptoms, which allows transmission of the parasite before treatment. Third, *P. vivax* develops dormant hypnozoites in the liver of the human host, which awaken in the weeks and months following a primary attack and cause relapses. Finally, vivax malaria is often transmitted by outdoor biting mosquitoes, making the current insecticide-based control measures (LLIN and IRS) less effective. Because of these unique features, traditional malaria control efforts often fail to control *P. vivax* transmission. In addition, containment of *P. falciparum* has been prioritized in the GMS, partially because of the emerging artemisinin resistance. As a result, *P. falciparum* prevalence has decreased, while the proportion of *P. vivax* has increased.

In the GMS, the first-line therapy for vivax malaria remains chloroquine (CQ) and primaquine (PQ) (**Table 1**) [41]. Reports of clinical CQ resistance in many regions of the world and falling efficacy of PQ are of great concern for vivax malaria control [42–45]. Although some studies indicated that *P. vivax* in the GMS remained sensitive to CQ [46–51], others clearly documented CQ-resistant *P. vivax* [52–55]. In Myanmar, sporadic CQ-resistant *P. vivax* cases were first reported more than 20 years ago [52, 53]. A later report of 34% treatment failures in Dawei of Southern Myanmar suggests an increase of CQ resistance [55]. More recent studies identified both early and late treatment failures in Myawaddy of the Kayin State and Kawthaung of the Tanintharyi Region, Myanmar [56]. In northeastern Myanmar bordering China, a recent study showed 5.2% cumulative incidence of recurrent parasitemia during a 28-day follow-up of 587 *P. vivax* treated with CQ/PQ [57], suggesting sensitivity to CQ may also be deteriorating in this region. This reduced sensitivity of *P. vivax* to CQ requires close surveillance and potential

Studies from Papua New Guinea suggest that 80% of the vivax infections may be attributed to relapses. A modeling approach predicts that as much as 96% of clinical attacks by *P. vivax* in Thailand are due to relapses [60]. For radical cure, WHO recommends a dose of 0.25–0.5 mg/kg of PQ daily for 14 days. However, the lower dose (total of 3.5 mg/kg) fails to prevent relapses in many different endemic sites [61]. Because of the potential risk of severe hemolysis that this drug could cause in patients with glucose-6-phosphate dehydrogenase (G6PD) deficiency, PQ is not widely prescribed [43, 62, 63]. In routine practice, G6PD status is not screened; the GMS

implementation of more effective treatment measures such as ACTs [58].

mission is required to guide the efforts of NMCPs to achieve malaria elimination.

**4. The burden of** *P. vivax* **malaria and G6PD deficiency**

184 Towards Malaria Elimination - A Leap Forward

The *G6PD* gene is extraordinarily polymorphic with more than 400 variants discovered based on biochemical diagnosis [75], among which 186 mutations are associated with G6PD deficiency [76]. The prevalence of G6PD deficiency and distribution of G6PD variants vary geographically [77]. In the GMS, G6PD deficiency is often highly prevalent among ethnic groups. Along the Thailand-Myanmar border, the prevalence of G6PD deficiency was above 10% [78–80], whereas in the Kachin ethnicity along the China-Myanmar border, it almost reached 30% [81]. In Thailand and Myanmar, the Mahidol variant (487G>A) is the most predominant and often accounts for ~90% of all mutations [79, 81–83]. According to the WHO classification, the Mahidol variant is a Class III mutation or mild-deficient variant with 60% enzyme activity [76]. However, this classification may not be accurate since patients with the Mahidol variant often had <1% of the normal G6PD activity [79, 84, 85]. Patients having the G6PD Mahidol variant (487G>A) rarely had acute hemolytic anemia after taking the normal dose of PQ [84, 86]. In contrast to the belief that PQ only induces mild hemolysis in patients with the Mahidol variant, there have been case reports showing that the normal dosage of 15 mg/kg/day for 3 days in vivax patients with this G6PD variant could lead to acute hemolytic anemia that required blood transfusion or even cause renal failure [87–89]. It is noteworthy that G6PD activity can vary substantially between individuals with the same variant and even within the same individual over time. Therefore, with the prevalence of vivax malaria in this region and the goal of malaria elimination, the deployment of point-of-care G6PD deficiency diagnostics is urgent [90]. In addition, there is a need to test whether weekly PQ of 0.75 mg/kg for 8 weeks, a dosage considered safe for the G6PD African variant [91], could be prescribed in the GMS without prior testing for G6PD deficiency.
