Immunology and Infectious Disease Commons^™

Ectoparasites Of The Critically Endangered Green Sawfish Pristis Zijsron And Sympatric Elasmobranchs In Western Australia, Jack Ingelbrecht, Karissa O. Lear, Storm B. Martin, Alan J. Lymbery, Bradley M. Norman, Geoffrey A. Boxshall, David L. Morgan

Harold W. Manter Laboratory: Library Materials

This study reports the metazoan ectoparasite fauna of juvenile Critically Endangered green sawfish, Pristis zijsron, and sympatric elasmobranchs in Western Australia. Five parasite taxa were found on 76 screened P. zijsron: Caligus furcisetifer (Copepoda: Caligidae), Dermopristis pterophila (Monogenea: Microbothriidae), Branchellion plicobranchus and Stibarobdella macrothela (Hirudinea: Piscicolidae), and praniza larvae of an unidentified gnathiid isopod. Only C. furcisetifer and D. pterophila were common, exhibiting discrepant site-specificity, with C. furcisetifer occurring mostly on the head and rostrum, and D. pterophila around the pectoral and pelvic fins. Intensity of infection for C. furcisetifer and D. pterophila increased with host total length and …

Lepocreadiidae (Trematoda) Associated With Gelatinous Zooplankton (Cnidaria And Ctenophora) And Fishes In Australian And Japanese Waters, Thomas H. Cribb, Scott C. Cutmore, Nicholas Q.-X. Wee, Joanna G. Browne, Pablo Diaz Morales, Kylie A. Pitt

Harold W. Manter Laboratory: Library Materials

We examined gelatinous zooplankton from off eastern Australia for lepocreadiid trematode metacercariae. From 221 specimens of 17 species of cnidarian medusae and 218 specimens of four species of ctenophores, infections were found in seven cnidarian and two ctenophore species. Metacercariae were distinguished using cox1 mtDNA, ITS2 rDNA and morphology. We identified three species of Prodistomum Linton, 1910 [P. keyam Bray & Cribb, 1996, P. orientale (Layman, 1930), and Prodistomum Type 3], two species of Opechona Looss, 1907 [O. kahawai Bray & Cribb, 2003 and O. cf. olssoni], and Cephalolepidapedon saba Yamaguti, 1970. Two species were found in cnidarians …

New Hard Tick (Acari: Ixodidae) Reports And Detection Of Rickettsia In Ticks From Sierra Nevada De Santa Marta, Colombia, Ángel Oviedo, Miguel M. Rodríguez, Fernando S. Flores, Lyda R. Castro

Harold W. Manter Laboratory: Library Materials

The Sierra Nevada de Santa Marta (SNSM), located in northern Colombia, is considered a geographical island with high levels of biodiversity and endemism. However, little is known about tick species and their associated microorganisms at the SNSM. In this study we sampled host-seeking ticks in areas of the town of Minca within the SNSM. We collected 47 ticks identified as Amblyomma pacae, Amblyomma longirostre, Amblyomma ovale, Amblyomma mixtum, Haemaphysalis juxtakochi, Ixodes sp. cf. Ixodes affinis and Ixodes sp. Of these ticks, we tested for Rickettsia spp. by amplifying the gltA, SCA1, and 16S rRNA genes via PCR. Rickettsia amblyommatis …

Low Similarity Between Parasite Communities Of Ten Sympatric Carangid Species, Juan M. Osuna-Cabanillas, Emigdio Marín-Enríquez, Ana P. Martínez-Falc´On, Juan T. Timi, Francisco N. Morales-Serna

Harold W. Manter Laboratory: Library Materials

Host phylogeny and ecological convergence are two factors thought to influence the structure of parasite communities. The aims of this study were to determine the diversity of metazoan parasites of 10 sympatric fish species of the family Carangidae from the southeastern Gulf of California, and to analyze their similarity at infracommunity and component community levels, in order to determine if the host species, particularly those congeneric with similar ecological characteristics, exhibit similar assemblages of parasites. In total, 874 fish specimens were examined and 40 parasite species were identified. The component community was composed by 21 parasite species in Caranx caninus …

Application Of Dna Barcodes In The Genetic Diversity Of Hard Ticks (Acari: Ixodidae) In Kazakhstan, Ziwei Zheng, Weixin Zeng, Suwen Wang, Wenbo Tan, Xiaobo Lu, Kenesbay Kairullayev, Ligu Mi, Wurelihazi Hazihan, Gang Liu, Meihua Yang, Yuanzhi Wang

Harold W. Manter Laboratory: Library Materials

Forty-five tick species have been recorded in Kazakhstan. However, their genetic diversity and evolutionary relationships, particularly when compared to ticks in neighbouring countries, remain unclear. In the present study, 148 mitochondrial cytochrome c oxidase subunit I (COI) sequence data from our laboratory and NCBI (National Center for Biotechnology Information; https://www.ncbi.nlm.nih.gov/) data were used to address this knowledge gap. Phylogenetic analyses showed that i) Hyalomma anatolicum anatolicum (Koch, 1844) ticks from Jambyl Oblast (southeastern Kazakhstan) and Gansu Province (northwestern China) constituted a newly deviated clade; and ii) Dermacentor reticulatus (Fabricius, 1974) ticks from South Kazakhstan Oblast were closer …

Molecular Species Delimitation Of Marine Trematodes Over Wide Geographical Ranges: Schikhobalotrema Spp. (Digenea: Haplosplanchnidae) In Needlefishes (Belonidae) From The Pacific Ocean And Gulf Of Mexico, Gerardo Pérez-Ponce De León, Brenda Solórzano-García, Daniel C. Huston, Berenit Mendoza-Garfias, Jhonatan Cabañas-Granillo, Scott C. Cutmore, Thomas H. Cribb

Harold W. Manter Laboratory: Library Materials

Geographical distribution plays a major role in our understanding of marine biodiversity. Some marine fish trematodes have been shown to have highly restricted geographical distributions, while some are known to occur over very wide ranges; however, very few of these wide distributions have been demonstrated genetically. Here, we analyse species of the genus Schikhobalotrema (Haplosplanchnidae) parasitizing beloniforms from the tropical west Pacific, the eastern Pacific and the Gulf of Mexico (GoM). We test the boundaries of these trematodes by integrating molecular and morphological data, host association, habitat of the hosts and geographical distribution, following a recently proposed and standardized delineation …

Intraspecific Variation Of Morphological Traits Backed Up With Molecular Evidence Votes For Re‑Appraisal Of Hitherto Distinguished Balaustium Species—A Case Study Of Balaustium Murorum (Acariformes: Parasitengona, Erythraeidae), Joanna Mąkol, Magdalena Felska

Harold W. Manter Laboratory: Library Materials

Molecular examination of representatives of Balaustium from several populations in SW Poland, performed using the sequence data from the mitochondrial cytochrome c oxidase subunit I, confirmed their common specific affiliation and identity with Balaustium murorum. The potential presence of distinct species in the studied material, preliminarily inferred from the discovery of clusters as a result of Principal Component Analysis exploring the metric data sets, was rejected due to the finding of only one haplotype, at intra- and inter-population sampling. An insight into meristic traits in larvae, focused on chaetotaxy of legs, revealed wider variation than hitherto recognized for the …

Leucocytozoon Cariamae N. Sp. And Haemoproteus Pulcher Coinfection In Cariama Cristata (Aves: Cariamiformes): First Mitochondrial Genome Analysis And Morphological Description Of A Leucocytozoid In Brazil, Lis Marques De C. Vieira, Pedro Henrique O. Pereira, Daniel Ambrózio Da Rocha Vilela, Irène Landau, M. Andreína Pacheco, Ananias A. Escalante, Francisco C. Ferreira, Érika Martins Braga

Harold W. Manter Laboratory: Library Materials

The distribution of avian haemosporidians of the genus Leucocytozoon in the Neotropics remains poorly understood. Recent studies confirmed their presence in the region using molecular techniques alone, but evidence for gametocytes and data on putative competent hosts for Leucocytozoon are still lacking outside highland areas. We combined morphological and molecular data to characterize a new Leucocytozoon species infecting a non-migratory red-legged seriema (Cariama cristata), the first report of a competent host for Leucocytozoon in Brazil. Leucocytozoon cariamae n. sp. is distinguished from the Leucocytozoon fringillinarum group by its microgametocytes that are not strongly appressed to the host cell …

Distribution Of The Soft Tick Carios Vespertilionis In Lowlands And Low Mountain Regions Of Germany, Anja Fritzsche, Stefan Zaenker, Jannis Gottwald, Renate Keil, Christian Zaenker, Michael Bröker, Lidia Chitimia-Dobler

Harold W. Manter Laboratory: Library Materials

In Germany, the knowledge about ticks infesting bats is limited, and is restricted only to a few studies, most of them dating back decades. To further improve our knowledge on ticks parasitising bats, healthy and sick bats in central Germany were examined for ticks. In total 519 larvae and one nymph of Carios vespertilionis were collected from nine bat species: Eptesicus nilssonii, Eptesicus serotinus, Myotis daubentonii, Myotis myotis, Nyctalus leisleri, Pipistrellus nathusii, Pipistrellus pygmaeus, Pipistrellus pipistrellus, and Vespertilio murinus. Either the presence of C. vespertilionis was new for some areas or it was confirmed in some federal states in …

The Complete Mitochondrial Genome Of Eulaelaps Huzhuensis (Mesostigmata: Haemogamasidae), Hui-Juan Yang, Zhi-Hua Yang, Tian-Guang Ren, Wen-Ge Dong

Harold W. Manter Laboratory: Library Materials

Some mites of the family Haemogamasidae can transmit a variety of zoonotic diseases and have important public health and safety implications. Currently, however, little attention has been paid to molecular data of Haemogamasidae species, limiting our understanding of their evolutionary and phylogenetic relationships. In this study, the complete mitochondrial genome of Eulaelaps huzhuensis was determined for the first time, and its genomic information was analyzed in detail. The mitochondrial genome of E. huzhuensis is 14,872 bp in length with 37 genes and two control regions. The base composition showed a distinct AT preference. Twelve protein-coding genes have a typical ATN …

Rickettsia Africae And Other Unclassified Rickettsia Species Of The Spotted Fever Group In Ticks Of The Western Ghats, India, N. Naren Babu, Anup Jayaram, Amogh Milind Auti, Yuvraj Bhandari, Ujwal Shetty, Govindakarnavar Arunkumar

Harold W. Manter Laboratory: Library Materials

The spotted fever group (SFG) of Rickettsia are zoonotic disease-causing pathogens, commonly transmitted by hard ticks to a wide range of hosts, including humans. Rickettsia conorii is the common SFG recognised in India, whereas most of the infections due to other group species go undifferentiated at the species level. Hence, this study was conducted to screen host-seeking ticks in the Western Ghats region, India, for the DNA of SFG Rickettsia. The ticks were collected from Kerala, Goa, and Maharashtra states of India during a survey conducted between November 2017 and January 2018. In total, 288 tick pools were screened …

Atlas Of Ticks (Acari: Argasidae, Ixodidae) In Germany: 1st Data Update, Franz Rubel, Stefan Zaenker, Alexander Weigand, Dieter Weber, Lidia Chitimia-Dobler, Olaf Kahl

Harold W. Manter Laboratory: Library Materials

The first data update of the atlas of ticks in Germany published in 2021 is presented here. This atlas provides maps based on georeferenced tick locations of 21 species endemic in Germany as well as three tick species that are regularly imported to Germany. The data update includes the following numbers of newly georeferenced tick locations: 17 Argas reflexus, 79 Carios vespertilionis, 2 Dermacentor marginatus, 43 Dermacentor reticulatus, 4 Haemaphysalis concinna, 3 Haemaphysalis punctata, 3 Hyalomma rufipes, 3 Ixodes apronophorus, 9 Ixodes arboricola, 1 Ixodes ariadnae, 30 Ixodes canisuga, …

Antimalarial Chemoprophylaxis For Forest Goers In Southeast Asia: An Open-Label, Individually Randomised Controlled Trial, Rupam Tripura, Lorenz Von Seidlin, Siv Sovannaroth, Thomas J. Peto, James J. Callery, Meas Sokha, Mom Ean, Chhouen Heng, Franca Conradis-Jansen, Wanassanan Madmanee, Pimnara Peerawaranun, Naomi Waithira, Panarasri Khonputsa, Monnaphat Jongdeepaisal, Kulchada Pongsoipetch, Paphapisa Chotthanawathit, Ung Soviet, Christopher Pell, Jureeporn Duanguppama, Huy Rekol, Joel Tarning, Mallika Imwong, Mavuto Mukaka, Nicholas J. White, Arjen M. Dondorp, Richard J. Maude

Harold W. Manter Laboratory: Library Materials

Summary

Background

Malaria in the eastern Greater Mekong subregion has declined to historic lows. Countries in the Greater Mekong subregion are accelerating malaria elimination in the context of increasing antimalarial drug resistance. Infections are now increasingly concentrated in remote, forested foci. No intervention has yet shown satisfactory efficacy against forest-acquired malaria. The aim of this study was to assess the efficacy of malaria chemoprophylaxis among forest goers in Cambodia.

Methods

We conducted an open-label, individually randomized controlled trial in Cambodia, which recruited participants aged 16–65 years staying overnight in forests. Participants were randomly allocated 1:1 to antimalarial chemoprophylaxis, a 3-day …

Worldwide Host Associations Of The Tick Genus Ixodes Suggest Relationships Based On Environmental Sharing Rather Than On Co-Phylogenetic Events, Agustín Estrada-Peña, Alberto A. Guglielmone, Santiago Nava

Harold W. Manter Laboratory: Library Materials

Abstract

Background

This study aims to capture how ticks of the genus Ixodes gained their hosts using network constructs. We propose two alternative hypotheses, namely, an ecological background (ticks and hosts sharing environmentally available conditions) and a phylogenetic one, in which both partners co-evolved, adapting to existing environmental conditions after the association took place.

Methods

We used network constructs linking all the known pairs of associations between each species and stage of ticks with families and orders of hosts. Faith’s phylogenetic diversity was used to evaluate the phylogenetic distance of the hosts of each species and changes occurring in the …

Descriptive Dna Barcoding Of Argas (Persicargas) Arboreus And Argas (Persicargas) Persicus Ticks (Ixodida: Argasidae) Infesting Birds In Egypt, Enas H. Ghallab, Ayat Yousery, Mona G. Shaalan

Harold W. Manter Laboratory: Library Materials

Argas ticks are primary parasites of birds with veterinary importance. Nevertheless, these ticks have received little attention regarding molecular identification studies. DNA barcoding is a powerful technique for identifying tick species besides traditional morphological identification. The present work is a first effort to divulge DNA sequences of Argas (Persicargas) arboreus from Egypt and worldwide. We used cytochrome c oxidase subunit I (COI) from A. arboreus infesting herons, and from the fowl tick Argas (Persicargas) persicus. Our results pointed out another success for the Folmer primers that are widely used in DNA barcoding, permitting the discrimination …

The First Complete Genome Of The Simian Malaria Parasite Plasmodium Brasilianum, Marko Bajic, Shashidhar Ravishankar, Mili Sheth, Lori A. Rowe, M. Andreina Pacheco, Dhruviben S. Patel, Dhwani Batra, Vladimir Loparev, Christian Olsen, Ananias A. Escalante, Fredrik Vannberg, Venkatachalam Udhayakumar, John W. Barnwell, Eldin Talundzic

Harold W. Manter Laboratory: Library Materials

Naturally occurring human infections by zoonotic Plasmodium species have been documented for P. knowlesi, P. cynomolgi, P. simium, P. simiovale, P. inui, P. inui-like, P. coatneyi, and P. brasilianum. Accurate detection of each species is complicated by their morphological similarities with other Plasmodium species. PCR-based assays offer a solution but require prior knowledge of adequate genomic targets that can distinguish the species. While whole genomes have been published for P. knowlesi, P. cynomolgi, P. simium, and P. inui, no complete genome for P. brasilianum has …

Morphometric And Molecular Characterization Of An Unpigmented Haemosporidian Parasite In The Neotropical Turnip-Tailed Gecko (Thecadactylus Rapicauda), Nubia E. Matta, Leydy P. González, Mario Vargas-Ramírez, Gediminas Valkiūnas, Ananias A. Escalante, M. Andreína Pacheco

Harold W. Manter Laboratory: Library Materials

Morphological traits from blood stages have been the gold standard for determining haemosporidian parasite species. However, the status of some taxa and the value of such traits in parasites from reptiles remain contentious. The scarce sampling of these species worsens the situation, and several taxa lack molecular data. A survey was performed in the Magdalena Department in Colombia, where 16 species of reptiles were captured. A peculiar haemosporidian parasite was found in the Turnip-tailed gecko Thecadactylus rapicauda. This haemosporidian does not show malarial pigment in blood stages under light microscopy; thus, it fits the Garnia genus’s characters belonging to …

Great-Tailed Grackles (Quiscalus Mexicanus) As A Tolerant Host Of Avian Malaria Parasites, M. Andreína Pacheco, Francisco C. Ferreira, Corina J. Logan, Kelsey B. Mccune, Maggie P. Macpherson, Sergio Albino Miranda, Diego Santiago-Alarcon, Ananias A. Escalante

Harold W. Manter Laboratory: Library Materials

Great-tailed Grackles (Quiscalus mexicanus) are a social, polygamous bird species whose populations have rapidly expanded their geographic range across North America over the past century. Before 1865, Great-tailed Grackles were only documented in Central America, Mexico, and southern Texas in the USA. Given the rapid northern expansion of this species, it is relevant to study its role in the dynamics of avian blood parasites. Here, 87 Great-tailed grackles in Arizona (a population in the new center of the range) were screened for haemosporidian parasites using microscopy and PCR targeting the parasite mitochondrial cytochrome b gene. Individuals were caught …

Why Plasmodium Vivax And Plasmodium Falciparum Are So Different? A Tale Of Two Clades And Their Species Diversities, Ananias A. Escalante, Axl S. Cepeda, M. Andreína Pacheco

Harold W. Manter Laboratory: Library Materials

The global malaria burden sometimes obscures that the genus Plasmodium comprises diverse clades with lineages that independently gave origin to the extant human parasites. Indeed, the differences between the human malaria parasites were highlighted in the classical taxonomy by dividing them into two subgenera, the subgenus Plasmodium, which included all the human parasites but Plasmodium falciparum that was placed in its separate subgenus, Laverania. Here, the evolution of Plasmodium in primates will be discussed in terms of their species diversity and some of their distinct phenotypes, putative molecular adaptations, and host–parasite biocenosis. Thus, in addition to a current …

Maps Of Ticks (Acari: Argasidae, Ixodidae) For Austria And South Tyrol, Italy, Franz Rubel, Katharina Brugger

Harold W. Manter Laboratory: Library Materials

A first compilation of georeferenced tick locations in Austria and South Tyrol, Italy, is presented here. This allows the tick fauna to be examined in the various climatic regions of the European Alps. The dataset comprises 424 tick locations of Austria and 48 tick locations of South Tyrol, which were digitized from literature and visualized in the form of geographical maps. The tick fauna of Austria includes two species of Argasidae in the genera Argas and Carios and 15 species of Ixodidae in the genera Dermacentor, Haemaphysalis, and Ixodes, altogether 17 tick species. In addition, two species …

Larval Habitats, Species Composition And Distribution Of Malaria Vectors In Regions With Autochthonous And Imported Malaria In Roraima State, Brazil, Nathália Coelho Vargas De Almeida, Jaime Louzada, Maycon Sebastião Alberto Santos Neves, Thiago M. Carvalho, Júlio Castro‑Alves, Teresa Fernandes Silva‑Do‑Nascimento, Ananias A. Escalante, Joseli Oliveira‑Ferreira

Harold W. Manter Laboratory: Library Materials

Background: Malaria control requires local action. Assessing the vector diversity and abundance provides information on the local malariogenic potential or risk of transmission. This study aimed to determine the Anopheles species composition, habitats, seasonal occurrence, and distribution in areas with autochthonous and imported malaria cases in Roraima State.

Methods: A longitudinal study was conducted from January 2017 to October 2018, sampling larvae and adult mosquitoes in three municipalities of Roraima State: Boa Vista, Pacaraima and São João da Baliza. These areas have different risks of malaria importation. Four to six mosquito larval habitats were selected for larval sampling …

Role Of Three Bird Species In The Life Cycle Of Two Sarcocystis Spp. (Apicomplexa, Sarcocystidae) In The Czech Republic, Ondřej Máca, David González-Solís

Harold W. Manter Laboratory: Library Materials

Birds are one of the groups involved in the development of Sarcocystis Lankester (1882), serving either as intermediate or definitive hosts. The white-tailed sea eagle Haliaeetus albicilla (Linnaeus, 1758), red kite Milvus milvus (Linnaeus, 1758) (both Accipitriformes) and common starlings Sturnus vulgaris Linnaeus, 1758 (Passeriformes) were examined to elucidate their participation in the development of Sarcocystis, as well as to determine the specific identity of the parasites based on morphological and especially molecular analyses. In 2020–2021, one white-tailed eagle, one red kite and five common starlings were parasitologically examined for the presence of Sarcocystis using flotation centrifugation coprological method …

Genomic Approaches To Uncovering The Coevolutionary History Of Parasitic Lice [Review], Kevin P. Johnson

Harold W. Manter Laboratory: Library Materials

Next-generation sequencing technologies are revolutionizing the fields of genomics, phylogenetics, and population genetics. These new genomic approaches have been extensively applied to a major group of parasites, the lice (Insecta: Phthiraptera) of birds and mammals. Two louse genomes have been assembled and annotated to date, and these have opened up new resources for the study of louse biology. Whole genome sequencing has been used to assemble large phylogenomic datasets for lice, incorporating sequences of thousands of genes. These datasets have provided highly supported trees at all taxonomic levels, ranging from relationships among the major groups of lice to those among …

Circling The Drain: The Extinction Crisis And The Future Of Humanity, Rodolfo Dirzo, Gerardo Ceballos, Paul R. Ehrlich

Harold W. Manter Laboratory: Library Materials

Humanity has triggered the sixth mass extinction episode since the beginning of the Phanerozoic. The complexity of this extinction crisis is centered on the intersection of two complex adaptive systems: human culture and ecosystem functioning, although the significance of this intersection is not properly appreciated. Human beings are part of biodiversity and elements in a global ecosystem. Civilization, and perhaps even the fate of our species, is utterly dependent on that ecosystem’s proper functioning, which society is increasingly degrading. The crisis seems rooted in three factors. First, relatively few people globally are aware of its existence. Second, most people who …

Diversity And Phylogenetic Relationships Of ‘Tetraphyllidean’ Clade 3 (Cestoda) Based On New Material From Orectolobiform Sharks In Australia And Taiwan, Janine N. Caira, Kirsten Jensen

Harold W. Manter Laboratory: Library Materials

In an effort to expand knowledge of Clade 3—one of the ten clades that compose the non-monophyletic order ‘Tetraphyllidea’ all current members of which parasitise orectolobiform sharks—we targeted species of orectolobiform sharks that had not previously been examined for ‘tetraphyllidean’ cestodes. That work led to the discovery of three new species off Australia and Taiwan. Ambitalveolus gen. n. was erected to accommodate these species. Ambitalveolus costelloae gen. n. et sp. n., Ambitalveolus kempi sp. n., and Ambitalveolus penghuensis sp. n. differ from one another in scolex size, genital pore position, and number of marginal loculi, proglottids, and testes. Among ‘tetraphyllideans’, …

Malaria Resilience In South America: Epidemiology, Vector Biology, And Immunology Insights From The Amazonian International Center Of Excellence In Malaria Research Network In Peru And Brazil, Kathrine Torres, Marcelo U. Ferreira, Marcia C. Castro, Ananias A. Escalante, Jan E. Conn, Elizabeth Villasis, Maisa Da Silva Araujo, Gregorio Almeida, Priscila T. Rodrigues, Rodrigo M. Corder, Anderson R.J. Fernandes, Priscila R. Calil, Winni A. Ladeia, Stefano S. Garcia-Castillo, Joaquin Gomez, Lis Ribeiro Do Valle Antonelli, Ricardo T. Gazzinelli, Douglas T. Golenbock, Alejandro Llanos-Cuentas, Dionicia Gamboa, Joseph M. Vinetz

Harold W. Manter Laboratory: Library Materials

The 1990s saw the rapid reemergence of malaria in Amazonia, where it remains an important public health priority in South America. The Amazonian International Center of Excellence in Malaria Research (ICEMR) was designed to take a multidisciplinary approach toward identifying novel malaria control and elimination strategies. Based on geographically and epidemiologically distinct sites in the Northeastern Peruvian and Western Brazilian Amazon regions, synergistic projects integrate malaria epidemiology, vector biology, and immunology. The Amazonian ICEMR’s overarching goal is to understand how human behavior and other sociodemographic features of human reservoirs of transmission— predominantly asymptomatically parasitemic people—interact with the major Amazonian malaria …