A systematic review of antimalarial medicinal plants in Democratic Republic of the Congo

In Democratic Republic of the Congo (DRC), malaria is caused by a parasite called Plasmodium falciparum. Malaria is one of the country's major public health issues and responsible for the death of tens of thousands of people every year in both rural and urban environments. Antimalarial drugs are commonly used but some recorded cases of drug resistance are a major obstacle to controlling the spread of malaria. It is therefore essential to identify new bioactive molecules as an alternative. Many medicinal plants with different properties have been used as treatments for a variety of diseases in the DRC, including malaria. This study provides a systematic review of antimalarial plant resources in the DRC. From 28 papers on ethnomedicine published between 2001 and 2019, a total of 232 plant species belonging to 67 different plant families and identified in 13 provinces was reported in the treatment of malaria. A large number of these plant species belong to the Fabaceae, Asteraceae, Euphorbiaceae, Rubiaceae, and Apocyanaceae families. Species cited in more than three provinces include Cymbopogon citratus, Vernonia amygdalina, Rauvolfia vomitoria and Catharanthus roseus. Most of the species identified as antimalarial plants were tree species, with phanerophytes predominating. In addition, leaves were identified as the main ingredients for preparing remedies, most commonly by decoction administered orally. 
  
 


A systematic review of antimalarial medicinal plants in Democratic Republic of the Congo
In Democratic Republic of the Congo (DRC), malaria is caused by a parasite called Plasmodium falciparum. Malaria is one of the country's major public health issues and responsible for the death of tens of thousands of people every year in both rural and urban environments. Antimalarial drugs are commonly used but some recorded cases of drug resistance are a major obstacle to controlling the spread of malaria. It is therefore essential to identify new bioactive molecules as an alternative. Many medicinal plants with different properties have been used as treatments for a variety of diseases in the DRC, including malaria. This study provides a systematic review of antimalarial plant resources in the DRC. From 28 papers on ethnomedicine published between 2001 and 2019, a total of 232 plant species belonging to 67 different plant families and identified in 13 provinces was reported in the treatment of malaria. A large number of these plant species belong to the Fabaceae, Asteraceae, Euphorbiaceae, Rubiaceae, and Apocyanaceae families. Species cited in more than three provinces include Cymbopogon citratus, Vernonia amygdalina, Rauvolfia vomitoria and Catharanthus roseus. Most of the species identified as antimalarial plants were tree species, with phanerophytes predominating. In addition, leaves were identified as the main ingredients for preparing remedies, most commonly by decoction administered orally.
Keywords: antimalarial treatment, ethnomedicine, medicinal plants, systematic review, Democratic Republic of the Congo.

Introduction
Malaria, a disease caused by the parasite Plasmodium falciparum, is one of the major public health problems in many tropical countries. This disease is spread by mosquitoes (Landis et al., 2009;Messina et al., 2011). According to the World Health Organization (WHO), about 228 million cases and more than 405 thousand deaths related to malaria have occurred in 2018 around the world, with the majority of deaths (93%) found in Africa (WHO, 2019).
In Democratic Republic of the Congo (DRC), reports indicated that, despite the efforts to prevent the spread of malaria, and alleviate its detrimental effects on the people's health nationwide (Swana et al., 2018), other forms of challenges, such as drug resistance of the pathogen was identified as obstacle to control efficiently the spread of the disease in the sub-Saharan Africa (Alker et al., 2008;Mobula et al., 2009;Mvumbi et al., 2015). To address this situation, there is a strong necessity to identify novel substances or bioactive molecules having the potential to overcome the drug resistance of the Plasmodium falciparum, eventually due to the recurrent use of similar molecules. Thus, medicinal plants could serve as an alternative to achieve this goal (Silva et al., 2011;Ntie-Kang et al., 2014), since they are relatively cost-effective and highly accessible (Madureira et al., 2002;Muganga et al., 2010).
The benefit of plants as bio-resources and their potentialities to treat different diseases has traditionally been highlighted by several research (Arshad et al., 2014;Zarei et al., 2017), and it is considered essential for human health care (Asadi-Samani et al., 2013). The worldwide use and distribution of bio-resources enclose a large potential to unveil the undescribed aspects of medicinal plants, yet undiscovered (Balima et al., 2018).
According to the WHO, around 80% of the population living in developing countries rely on traditional medicine for treating diseases (Kamatenesi-Mugisha and Oryem-Origa, 2005;Mahomoodally, 2013). Moreover, the extensive use of traditional medicines in Africa could be associated to the history and culture or customs, and economic environment (Mahomoodally, 2013). Medicinal plants still represent an important source of medical treatment in developing countries (Tabuti et al., 2003).
The Congo Basin region itself is megadiverse in plant species, including medicinal plants (Light et al., 2016). Screening studies for antimalarial medicinal plant species from Cameroon (Betti, 2002;Saotoing et al., 2011;Titanji et al., 2008;Kuete and Efferth, 2010;Betti et al., 2013a), Gabon (Betti et al., 2013b), the Central African Republic (Lakouéténé et al., 2009), Republic of Congo (Mbatchi et al., 2006;Nsonde-Ntandou et al., 2005) and Guinea Equatorial (Gomez Marín and Merino Cristóbal, 1990) have demonstrated the potential of traditional medicinal plants as source of antimalarial substances. Furthermore, the DRC, with about half of the African humid forests, ranks fifth in the world for the diversity of plant species (UNESCO, 2010;Masunda et al., 2019). It is estimated that the flora of the DRC comprises 377 families, 2,196 genera, and over 11,000 species (Mbala, 2003). Many plants species in the DRC have been studied and described for their beneficial impact for treating various diseases in numerous scientific papers. Thus, the purpose of this study is to develop a systematic review of scientific papers on ethnomedicine, particularly on the treatment of malaria.

Sources of information
Through literature review and data mining, previous published papers related to the field of ethnobotanyof medicinal plants in the DRC, were collected during December 2019, using available internet browsers.
The research was extended to the identification of references listed in retrieved articles. The major key words employed during the search, either in French or in English, were: "ethno-medicinal plants in DR of Congo" OR "ethnobotanical study in DR of Congo" OR "ethno pharmacology in DR of Congo" OR "phyto therapy in DR of Congo" OR "herbal treatments in DR of Congo" OR "anti-malaria plants in DR of Congo". In addition to the published scientific papers on the subjects of interest, all information found to be necessary for the study were included as well, collected from scientific works, such as books, academic theses (Masters and doctorate thesis dissertations), while considering the year of publication, the methodology, or the study language.

Analysis
The following ethnomedicinal information were collected from the literature: (i) plant organs used: leaves, stem, stem bark, root, root bark, flowers, fruits, seeds or grain, whole plant, upper (aboveground) part, underground part; (ii) mode of preparation: decoction, maceration, infusion, grinding, expression, extorsion, incineration, ash, etc.; (iii) way of administration: oral, enema, inhalation, bath, friction, chewing, tattoo, instillation; and (iv) the geographical location of the study. In the case of unavailability of data in one of the above groups, the corresponding field was marked "NR (not referenced)".
The dataset was taxonomically standardized (synonym and misspelling) and updated following guidelines in the online websites: the plant list 1 or/and African plant database 2 . The Angiosperm Phylogeny Group (APG III) was used to revise and update the family names (APG, 2009). The antimalarial plants species were characterized by morphology types: tree, shrub, sub-shrub, liana, herb. The life-form analysis focused on 223 species and was based on Raunkiaer's system: Phanerophytes, Chamaephytes, Hemicryptophytes, Geophytes or Cryptophytes, Therophytes, Aerophytes and Epiphytes.
The conservation status of each anti-malaria species was determined by the IUCN red list of threatened species (IUCN, 2019) and includes: Not Evaluated (NE), Data Deficient (DD), Least concern (LC), Near threatened (NT), Vulnerable (VU), Endangered (EN), Critically endangered (CR), Extinct in the Wild (EW) and Extinct (EX).

Selected antimalarial related studies
In the perspective of using only studies providing useful data and information related to the use of medicinal plants as a treatment for malaria, an initial selection was performed. As a result, all studies with no relevant data on antimalarial plants were discarded, and only 41 full-text studies were evaluated in order to verify the evidence of the antimalarial plants. In addition, another 13 studies falling either into the category of review article or in vitro studies were excluded. Consequently, after the screening and selection process, 28 publications and academic reports (Masters and Doctorate Theses) were retained for further investigations (figure 1). Of this number, the earliest article was published in 2001 but about 92.9% of the publications were released between 2010-2019. Studies were conducted on 13 different provinces in the DRC as illustrated in figure 2. Moreover, the majority of plant species (185 species, 79.7%) was reported to be used in at least one province, and 27 species (11.6%) were listed in at least two provinces (table I). Meanwhile, 20 species (8.6%) were mentioned in more than two provinces. An illustration of the top 20 plant species widely distributed across the country (used in more than 2 provinces) is given in table II.

Diversity and morphology of antimalarial plants
A total of 232 species of medicinal plants, distributed in 181 genera, and belonging to 67 families were mentioned as being involved in the treatment of malaria in different parts of the DRC (figure 2; table III). Among these families, about 82% are dicots, and nearly 14% are monocots, and only 3% are gymnosperms and pteridophytes. The dicotyledonous group is represented by 55 families and 169 genera, while the monocotyledonous group is represented by 10 families and 10 genera. The investigation on the life-form of the antimalarial plants showed that Phanerophytes represent 68.3% of the species cited, followed by Therophytes (13.4%), Chamaephytes (12.9%), Geophytes (4.5%), and Hemycryptophytes (0.9%) (table III). Regarding the morphological type, figure 3 shows that woody plants (66.8%) were the dominant type (trees: 38.8%, shrubs: 16.8%, sub-shrub: 3.02%, and liana: 8.2%). About half of the reported antimalarial plant species across the country (48.3%) belong to the following families: Fabaceae (30 species), Asteraceae (27 species), Rubiaceae (15 species), Euphorbiaceae (14 species), Apocynceae (10 species), Annonaceae (8 species) and Meliaceae (8 species). Around 49.2% of the families contributed with only one species to the antimalarial plant species, while 145 genera are represented by a single antimalarial plant species. Euphorbia (Euphorbiaceae) and Strychnos (Loganiaceae) were represented by 4 plant species each. In addition to Euphorbia and Strychnos, Acacia, Afromomum, Aloe, Chenopodium, Combretum, Dalbergia, Ficus, Landolphia, Morinda, Senna and Ziziphus have 3 species each (appendix 1).

Parts used, mode of preparation, and route of administration
The utilization of plant parts and their mode of preparation is subjected to their accessibility and the knowledge of indigenous people (Umair et al., 2019). The results of the analysis of the plant parts used show that the leaves were the most used parts of plant by traditional healers, which accounted for about 60% of the total parts of plants used, followed by roots (32.7%), and stems/ bark (22.4%). A few plants were harvested for their fruits or seeds/grains (6.5%) or used whole (5.2%). In 3.4% of cases, the parts of the plant used were not referenced (figure 4). Sixteen methods were used to prepare plant materials. Among these, the majority used decoction (169 reported), followed by maceration (48), infusion (27), grinding and sap (4 each), powder (3) expression, extorsion, and spray (2 each); chewing, ash, incineration, milling, paste, pounding, and leaves roasted in palm oil (1 each); and 19 were not referenced (NR) (figure 5). The majority (156) of antimalarial remedies were taken orally, followed by enema/anal/suppository (12), bath, inhalation, and instillation (7 each). In rare instances, treatment was administered by rubbing leaves all over the body (2) and by tattoo (1). Sixty-five reports failed to indicate the mode of administration of the plant medicines.

Threat status
The unsustainable exploitation of plant species has resulted in a high vulnerability for certain species (Raj et al., 2018). It was found that 49 plant species used as a remedy for treating malaria in the DRC were reported in the IUCN's Red List (table IV). According to the IUCN list, one species was classified as "data deficient", 43 as "least concern", three as "vulnerable", one as "near threatened", and one as "critically endangered". The other species were not included yet in the list. The international trade in a few of the species studied is also banned by the CITES treaty 3 : Aloe christianii, Aloe dawei, Dalbergia

Discussion
Two hundred thirty-two medicinal plants associated with the treatment of malaria in DRC that distributed across 181 genera and 67 families were identified. From these families, Fabaceae, Asteraceae, Euphorbiaceae, and Rubiaceae had the highest number of antimalarial plants species. These findings are similar to those obtained by Asase et al. (2010), Traore et al. (2013), and Taek et al. (2018) who reported that the Fabaceae had a high number of antimalarial plants species compared to other families. In a converse approach, Iyamah and Idu (2015) indicated that Fabaceae counts the highest number of antimalarial plants in Southern Nigeria, followed by Asteraceae. The predominance of medicinal plants from Fabaceae, Asteraceae ad Rubia-ceae is due to the highest number of species disseminated throughout the DRC (table V) (Bakwaye et al., 2013). Similar to observations from other countries (Adekunle, 2008;Tabuti, 2008;Kodi et al., 2017), woody plants constituted the largest source of antimalarial medicinal plants.
From an ecological perspective, the life-forms profile of the plant allows a better appreciation of ecological conditions in which they live (Kami Kanda et al., 2019). The high percentage of phanerophytes in antimalarial plants could be due to permanence of those plants throughout year (Mamadou et al., 2019). The predominance of woody species and phanerophytes shows a forest physiognomy (Kikufi et al., 2017;Lassa et al., 2019), while the therophytes correspond to the ruderal strategy (Kikufi and Lukoki, 2008   Although several plant parts were used in the Congo, leaves were the most commonly used in malaria treatment. This is similar to other studies from African countries that have demonstrated leaves to be the most frequently used plant part in plant remedies (Saotoing et al., 2011;Adia et al., 2014;Anywar et al., 2016). This contrasts with the findings of Ngarivhume et al. (2015) in Zimbabwe where roots were the most commonly used plant part. The preference for the utilization of leaves is justified by their effortless to collect, to store, and to process, but also, their action in photosynthesis and their bioactive compounds (Kayani et al., 2014;Bibi et al., 2015;Vijayakumar et al., 2015;Amjad et al., 2017;Faruque et al., 2019). Further, cutting leaves is less harmful to the plant development and growth (Alalwan et al., 2019).
It was found in this study that the most commonly used mode of preparation of antimalarial plants was decoction. This affirmation corroborates the reports from the other African countries (Koudouvo et al., 2011;Yetein et al., 2013;Alebie et al., 2017;Okello and Kang, 2019) and Asian countries (Bora et al., 2007;Ong et al., 2018). Decoction is largely used because of it is easy to prepare by mixing herbs with water, tea, or soup (Umair et al., 2019).
The primary administration route is oral. These results are consistent with the observations reported by other countries (Bora et al., 2007;Tor-anyiin et al., 2003;Idowu et al., 2010).
Among the recorded plant species, some have already been studied in vitro by Congolese Scientists, however, the phytochemical study of many antimalarial plant species has not yet been documented in vitro and/or in vivo.

Conclusion
This systematic review of medicinal plants provides a comprehensive insight into the existing antimalarial plants species in the Democratic Republic of the Congo (DRC). The 28 ethnobotanical studies published in the last two decades used in the study, highlighted the diversity of commonly used plant species with pharmacological effects, and their spatial distribution across the DRC (cultures and provinces), and represent an alternative mean for malaria prevention and a remedy for its treatment in the DRC. In addition, plants remain the major therapeutic remedy for malaria. Nevertheless, there are likely more sources of traditional knowledge and articles not published online that may contain precious information in the Phyto pharmacopeia against malaria that could serve as a basis for future studies.