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Haemogregarine Genetic Diversity in Captive African Rock Pythons from Nigeria Suggests a Geographical Pattern/Nijerya'daki Afrika Kaya Pitonlarindaki Hemogregarin Genetik Cesitlilik Cografik Bir Paterni Gostermektedir.


The phylum Apicomplexa is notable for its high medical and veterinary importance, particularly owing to the impacts of Plasmodium, Babesia, and Toxoplasma among others (1). Regardless of this, the knowledge on the taxonomical diversity of this phylum remains relatively limited. It has been estimated that only 0.1% of the Apicomplexan species have been described (2). Moreover, within this percentage, there is a bias in studies toward a few genera, being even less known regarding the parasites of wild hosts, including their identity, evolutionary relations, and ecology (3). Having this basic information could greatly improve the conservation efforts of hosts as well as increase the overall knowledge on Apicomplexa. Haemogregarines are geographically ubiquitous parasites that infect all groups of terrestrial vertebrates and an array of hematophagous arthropods (4). They present various life cycles, which usually involve an invertebrate vector as the definitive host and a vertebrate as the intermediate host but might also include other vertebrates as paratenic hosts upon whom the intermediate hosts prey. The use of prey--predator networks by these parasites has been shown by experimental transmission studies (5, 6) and supported by phylogenetic assessments (7). However, other factors, such as vertebrate host distribution, habits, and relatedness and their exposition to vectors have also been suggested to play a role in shaping the presence, frequency, and diversity of haemogregarines in host communities (7-9). Recent phylogenetic studies have also identified taxonomical incongruences, suggesting that the taxonomy of terrestrial haemogregarines should be rearranged and divided into four genera: Hepatozoon, Hemolivia, Karyolysus, and the newly created Bartazoon (10). Members from all these four genera have been found infecting reptiles; haemogregarines are one of the most commonly reported blood parasites, particularly in reptiles (1). Although haemogregarines are generally considered apathogenic in reptiles, the degree of the effect differs among studies, from no apparent effect to severe health effects (11). In this study, we screened African rock pythons from Nigeria for the presence of these parasites to assess their distribution, diversity, and phylogenetic relationships.



A total of 21 captive African rock pythons from 11 locations in the northern, central, and south-western parts of Nigeria across eight states were sampled between August 2016 and January 2017 (Figure 1a for the sampling map). Each snake was subjected to physical and clinical examinations, sexed, weighed, and measured. Blood was collected from the ventral coccygeal vein, as described by Lock (12), and stored in heparinized tubes for later use. Blood smears were examined for parasite detection using a light microscope. In ophidian hosts, haemogregarines are easily detected by observing their stages inside red blood cells (1). Photomicrographs were taken using a Magnus Fixed microscope adapter (FMA050 3MP) attached to a computer (MagnusPro 3.7 software). Blood from infected pythons was transferred to Whatman Flinders Technology Associates cards for molecular identification.

DNA Extraction and Amplification

DNA was extracted using standard high-salt methods (13). Polymerase chain reaction (PCR) was performed using primers specific for a 600-bp long region of the 18S rRNA gene, HepF300, and HepR900 (14). PCR was performed as described by Harris (15). Amplified products were purified and thereafter sequenced by an external company (Beckman Coulter Genomics, UK). Sequences were blasted in the GenBank database to confirm the identity of the amplified products. Sequences were corrected and aligned in Geneious v5.6.7, Biomatters Limited, New Zealand (16) using the MUSCLE algorithm (17). For phylogenetic analyses, GenBank sequences of other haemogregarines were added to the dataset, which resulted in a final dataset that included 145 sequences and was 584-bp long. The substitution model of evolution was chosen according to the Bayesian information criterion selected by jModelTest 2 (model 012030+G+F) (18). Phylogenetic relationships were estimated using the maximum likelihood (performed in PhyML 3.1, South of France bioinformatics platform, Montpellier, France) (19) and Bayesian inference (Mr. Bayes v.3.2.6 developed by Ronquist et al. (20) methods) (Figure 1b). For more details on parameters, see Tome (21).


All pythons were apparently healthy at the time of blood collection based on physical examination. From the 21 blood smears analyzed under the microscope, 10 (47.6%) snakes were found to be infected, with haemogregarines being the suspected parasite in all cases (Figure 1c and d). Eight of the infected samples were genetically assessed; however, all attempts of PCR amplification failed in one of the samples. All the seven obtained sequences belong to the Bartazoon haemogregarine group, as identified by Karadjian (10), in a well-supported clade of haemogregarines from snake, gecko, and rodent hosts, but in two distinct lineages (Figure 1b). The haemogregarines infecting pythons from the southwestern region of Nigeria grouped with Hepatozoon ayorgbor, described from the ball python Python regius in Ghana, and a sequence retrieved from Hemidactylus alkiyumii and the Oman saw-scaled viper Echis omanensis, both from Oman. The remaining four positive samples were from pythons of the north and central parts of Nigeria. They were grouped with sequences from a white-lipped Herald snake Crotaphopeltis hotamboeia from Niger, a horseshoe whip snake Hemorrhois hippocrepis from Spain, the Arabian horned viper Cerastes gasperettii, and the diademed sand snake Lytorhynchus diadema from Oman.


Haemogregarines have been previously reported in two python species (14, 22-25) and other snakes from Africa (9, 24, 26, 27). For snakes of tropical Africa, prevalence appears to increase in captivity, with values of 47.6% (in the present study) and 78.2% in imported snakes from Ghana (22). In contrast, wild snakes were less commonly infected, presenting frequencies of approximately 14% (24, 26) and 16.7% (25). There is not yet a consensus on the effect of haemogregarines on their reptile hosts. While some authors have reported none or little effect, higher parasitemia levels have been reported to be associated with erythrocyte hemolysis, anemia, and mortality (11, 28-32). However, the patterns observed appear to suggest that captivity increases the predisposition to infection, and such information should be considered in captivity and conservation programs.

Several haemogregarine species have been described from python hosts [Smith (4) included a list of available species description from the bibliography, and more recently, Sloboda et al. (22) described H. ayorgbor]. Unfortunately, molecular data are only available for H. ayorgbor; therefore, it is not possible to determine whether the haemogregarine haplotypes detected here belong to any described species. However, because the haplotype infecting southern Nigerian rock pythons shares the same 18S rRNA sequence as that of H. ayorgbor, it can be suspected that it belongs to this species or at least to a very closely related species. Other haemogregarine haplotypes infecting pythons have also been reported. Rosado et al. (25) screened the samples of Python sebae from Mauritania, Senegal, and Mali and detected a distinct haemogregarine haplotype and Haklova-Kocikova et al. (24) sampled pythons (Python natalensis and P. regius) from Swaziland and found one to be infected. Unfortunately, the region of the 18S rRNA analyzed in this study did not overlap with the sequence retrieved by Haklova-Kocikova et al. (24), and, thus, it was not included in the present phylogenetic analysis. However, Haklova's haplotype belongs to the same clade of haemogregarine as that in our study and that of others identified from pythons. Thus, based on the information available to date, African pythons, specifically rock pythons, are infected by three (possibly four) haemogregarine haplo-types.

A striking pattern in our results is the geographical division in the distribution of the two detected haplotypes, with one haplotype occupying the northern and central parts of Nigeria and the other occupying the southern part. Notably, Nigeria presents a latitudinal climatic variation, with a dryer savannah north and a tropical south (33), which might correlate with the distribution pattern of the haemogregarines. This is also supported by the grouping of the northern haplotype with haemogregarines from snakes of Niger and Spain (both countries with dryer climates) and those of the southern haplotype with a haemogregarine species described from Ghana (a tropical country). In contrast, sequences of haemogregarines from snakes in Oman (9) also grouped with both haplotypes. This might happen for several reasons but contradicts the climatic explanation for the pattern (or at least as the only explanation). In this study, we did not seek to identify the definitive host of these haemogregarines. However, the tick Aponomma latum was found infecting some snakes, and a gamont and suspected sporocyst was observed in a slide preparation from one of these ectoparasites (but no developmental stages in the hemocoel) (Figure 1e). Conversely, Sloboda et al. (22) established the role of the mosquito Culex quinquefasciatus as a vector in the experimental transmission of H. ayorgbor. Because the haplo-types detected are closely related to this species, they might have the same type of definitive host. Only further experiments can confirm this, but additionally, the identity of the definitive host might offer clues to explain the geographical pattern in the distribution of these haemogregarines, along with other factors such as differences in diet and genetic structuration of the python hosts.


To our knowledge, this is the first study on the presence and molecular assessment of blood parasites infecting snakes from Nigeria. Two haemogregarine haplotypes infecting Nigerian rock pythons genetically identified as part of the Bartazoon group, which appear to be differently distributed across the country, were detected in this study. Further studies are required to identify the possible definitive hosts of the haemogregarine parasites and to expand the sampling to other hosts and localities to assess whether the uncovered geographical pattern is due to climatic differences, particular of the host group, or distorted by the captivity.

Ethics Committee Approval: Ethics committee approval was received for this study from the University of Ibadan Animal Care and Use Research Committee.

Peer-review: Externally peer-reviewed.

Author Contributions: Concept--H.O.J., O.O.T., B.T.; Design--H.O.J., O.O.T., B.T., A.P.; Supervision--O.O.T., A.P.; Resources--H.O.J., B.T., O.O.T., A.P.; Materials--H.O.J., B.T., O.O.T., A.P.; Data Collection and/or Processing--H.O.J., B.T., O.O.T., A.P.; Analysis and/or Interpretation --B.T., H.O.J., O.O.T., A.P.; Literature Search--H.O.J., B.T.; Writing Manuscript--B.T., H.O.J.; Critical Review--O.O.T., A.P.

Conflict of Interest: No conflict of interest was declared by the authors.

Financial Disclosure: BT is funded by a Fundacao para a Ciencia e Tecnologia (FCT) PhD scholarship (PD/BD/52601/2014). AP is funded through a FCT contract (IF/01257/2012) under the Programa Operacional Potencial Humano--Quadro de Referencia Estrategico Nacional from the European Social Fund and Portuguese Ministerio da Educacao e Ciencia. Molecular work was funded by IF exploratory project (IF/01257/2012) to AP.

Etik Komite Onayi: Bu calisma icin etik komite onayi Ibadan Universitesi'nden alinmistir.

Hakem Degerlendirmesi: Dis bagimsiz.

Yazar Katkilari: Fikir--H.O.J., O.O.T., B.T.; Tasarim--H.O.J., O.O.T., B.T., A.P.; Denetleme--O.O.T., A.P.; Kaynaklar--H.O.J., B.T., O.O.T., A.P.; Malzemeler--H.O.J., B.T., O.O.T., A.P.; Veri Toplanmasi ve/veya Islemesi--H.O.J., B.T., O.O.T., A.P.; Analiz ve/veya Yorum--B.T., H.O.J., O.O.T., A.P.; Literatur Taramasi--H.O.J., B.T.; Yaziyi Yazan--B.T., H.O.J.; Elestirel Inceleme--O.O.T., A.P.

Cikar Catismasi: Yazarlar cikar catismasi bildirmemislerdir.

Finansal destek: CT, Bilim ve Teknoloji Vakfi (Fundacao para a Ciencia e Tecnologia (FCT)) doktora bursu ile karsilandi (PD/BD/52601/2014). AP, Avrupa Sosyal Fonu ve Portuguese Ministerio da Educacao e Ciencia, Programa Operacional Potencial Humano--Quadro de Referencia Estrategico Nacional altindaki bir FCT anlasmasi ile desteklendi (IF/01257/2012). Molekuler calisma IF arastirma projesi tarafindan finanse edildi (IF/01257/2012).


(1.) Telford SR. Hemoparasites of the Reptilia: color atlas and text. CRC Press of the Taylor & Francis Group; 2009.

(2.) Morrison DA. Evolution of the Apicomplexa: Where are we now? Trends Parasitol 2009; 25: 375-82.

(3.) Perkins SL, Martinsen ES, Falk BG. Do molecules matter more than morphology? Promises and pitfalls in parasites. Parasitology 2011; 138: 1664-74. [CrossRef]

(4.) Smith TG. The genus Hepatozoon (Apicomplexa: Adeleina). J. Parasitol 1996; 82: 565-85.

(5.) Johnson EM, Panciera RJ, Allen KE, Sheets ME, Beal JD, Ewing SA, et al. Alternate Pathway of Infection with Hepatozoon americanum and the Epidemiologic Importance of Predation. J Vet Intern Med 2009; 23: 1315-8. [CrossRef]

(6.) Sloboda M, Kamler M, Bulantova J, Votypka J, Modry D. Rodents as intermediate hosts of Hepatozoon ayorgbor (Apicomplexa: Adeleina: Hepatozoidae) from the African ball python, Python regius? Folia Parasitol 2008; 55: 13-6.

(7.) Tome B, Maia JPMC, Harris DJ. Hepatozoon infection prevalence in four snake genera: influence of diet, prey parasitemia levels, or parasite type? J Parasitol 2012; 98: 913-7.

(8.) Barta JR, Ogedengbe JD, Martin DS, Smith TG. Phylogenetic position of the Adeleorinid Coccidia (Myzozoa, Apicomplexa, Coccidia, Eucoccidiorida, Adeleorina) inferred using 18S rDNA sequences. J Eukaryot Microbiol 2012; 59: 171-80. [CrossRef]

(9.) Maia JP, Harris DJ, Carranza S, Gomez-Diaz E. Assessing the diversity, host-specificity and infection patterns of apicomplexan parasites in reptiles from Oman, Arabia. Parasitology 2016; 143: 1730-47. [CrossRef]

(10.) Karadjian G, Chavatte JM, Landau I. Systematic revision of the adeleid haemogregarines, with creation of Bartazoon n. g., reassignment of Hepatozoon argantis Garnham, 1954 to Hemolivia, and molecular data on Hemolivia stellata. Parasite 2015; 22: 31. [CrossRef]

(11.) Madsen T, Ujvari B, Olsson M. Old pythons stay fit; effects of haematozoan infections on life history traits of a large tropical predator. Oecologia 2005; 142: 407-12. [CrossRef]

(12.) Lock BA, Wellehan J. Ophidia (Snakes) In Fowler's Zoo and wild animal medicine 8th edition, Eds, Fowler, M. and Miller, R.E. Missouri: WB Saunders Co; 2015, p. 64.

(13.) Sambrook J, Fritsch EF, Maniatis T. Molecular Cloning: A Laboratory Manual. Cold Spring Harbour Press, New York; 1989.

(14.) Ujvari B, Madsen T, Olsson M. High prevalence of Hepatozoon spp. (Apicomplexa, Hepatozoidae) infection in water pythons (Liasis fuscus) from tropical Australia. J Parasitol 2004; 90: 670-2. [CrossRef]

(15.) Harris DJ, Maia JPMC, Perera A. Molecular characterization of Hepatozoon species in reptiles from the Seychelles. J. Parasitol 2011; 97: 106-10. [CrossRef]

(16.) Kearse M, Moir R, Wilson A, Stones-Havas S, Cheung M, Sturrock S, et al. Geneious Basic: An integrated and extendable desktop software platform for the organization and analysis of sequence data. Bioinformatics 2012; 28: 1647-9. [CrossRef]

(17.) Edgar RC. MUSCLE: multiple sequence alignment with high accuracy and high throughput. Nucleic Acids Res 2004; 32: 1792-7. [CrossRef]

(18.) Darriba D, Taboada GL, Doallo R, Posada D. jModelTest 2: more models, new heuristics and parallel computing. Nat Methods 2012; 9: 772. [CrossRef]

(19.) Guindon S, Gascuel O. A simple, fast, and accurate algorithm to estimate large phylogenies by maximum likelihood. Syst. Biol 2003; 52: 696-704.

(20.) Ronquist F, Teslenko M, Van Der Mark P, Ayres DL, Darling A, Hohna S, et al. MrBayes 3.2: efficient Bayesian phylogenetic inference and model choice across a large model space. Syst Biol 2012; 61: 539-42. [CrossRef]

(21.) Tome B, Rato C, Perera A, Harris DJ. High diversity of Hepatozoon spp. in geckos of the genus Tarentola. J Parasitol 2016; 102: 476-80.

(22.) Sloboda M, Kamler M, Bulantova J, Votypka J, Modry D. A new species of Hepatozoon (Apicomplexa: Adeleorina) from Python regius (Serpentes: Pythonidae) and its experimental transmission by a mosquito vector. J. Parasitol 2007; 93: 1189-98. [CrossRef]

(23.) Sumrandee C, Baimai V, Trinachartvanit W, Ahantarig A. Hepatozoon and Theileria species detected in ticks collected from mammals and snakes in Thailand. Ticks Tick Borne Dis 2015; 6: 309-15. [CrossRef]

(24.) Haklova-Kocikova B, Hiznanova A, Majlath I, Racka K, Harris DJ, Foldvari G, et al. Morphological and molecular characterization of Karyolysus--a neglected but common parasite infecting some European lizards. Parasit Vectors 2014; 7: 555. [CrossRef]

(25.) Rosado D, Harris DJ, Brito JC. Molecular screening of Hepatozoon (Apicomplexa: Adeleorina) infections in Python sebae from West Africa using 18 S rRNA gene sequences. Herpetol Notes 2015; 8: 461-3.

(26.) Tome B, Maia J, Salvi D, Brito J. Patterns of genetic diversity in Hepatozoon spp. infecting snakes from North Africa and the Mediterranean Basin. Syst Parasitol 2014; 87: 249-58. [CrossRef]

(27.) Abdel-Baki AA, Al-Quraishy S, Zhang JY. Redescription of Haemogregarina garnhami (Apicomplexa: Adeleorina) from the blood of Psammophis schokari (Serpentes: Colubridae) as Hepatozoon garnhami n. comb. based on molecular, morphometric and morphologic characters. Acta Parasitol 2014; 59: 294-300.

(28.) Oppliger A, Vernet R, Baez M. Parasite local maladaptation in the Canarian lizard Gallotia galloti (Reptilia: Lacertidae) parasitized by haemogregarian blood parasite. J Evol Biol 1999; 12: 951-5. [CrossRef]

(29.) Salakij J, Salakij C, Narkkong N, Chanhome L, Rochanapat N, Suthunmapinunta P. Hematozoa of snakes in Queen Saovabha Memorial Institute. Kasetsart J (Nat Sci) 2001; 35: 149-56.

(30.) Brown GP, Shilton CM, Shine R. Do parasites matter? Assessing the fitness consequences of haemogregarine infection in snakes. Can J Zool 2006; 676: 668-76. [CrossRef]

(31.) Saggese MD. Clinical approach to the anemic reptile. J Exot Pet Med 2009; 18: 98-111. [CrossRef]

(32.) Jegede HO, Omobowale TO, Okediran BS, Adegboye AA. Hematological and plasma chemistry values for the African rock python (Python sebae). Int J of Vet Sci Med 2017; 5: 181-6. [CrossRef]

(33.) Ogunsote OO, Prucnal-Ogunsote B. Defining climatic zones for architectural design in Nigeria: a systematic delineation. J. Environ. Technol 2002; 1: 1-14.

Henry Olanrewaju Jegede (1) [iD], Beatriz Tome (2,3) [iD], Ana Perera (2) [iD], Temidayo O. Omobowale (4) [iD]

(1) Veterinary Teaching Hospital, University of Ilorin, Ilorin, Nigeria

(2) CIBIO, InBIO - Research Network in Biodiversity and Evolutionary Biology, Universidade do Porto, Campus de Vairao, Rua Padre Armando Quintas, Vairao, Portugal

(3) Departamento de Biologia, Faculdade de Ciencias da Universidade do Porto, Rua do Campo Alegre, Porto, Portugal

(4) Department of Veterinary Medicine, University of Ibadan, Ibadan, Nigeria

Cite this article as: Jegede HO, Tome B, Perera A, Omobowale TO. Haemogregarine Genetic Diversity in Captive African Rock Pythons from Nigeria Suggests a Geographical Pattern. Turkiye Parazitol Derg 2018; 42:28-32.

Address for Correspondence / Yazisma Adresi: Temidayo O. Omobowale E.mail:

Received: 26.10.2017 Accepted: 11.12.2017

Gelis Tarihi: 26.10.2017 Kabul Tarihi: 11.12.2017

DOI: 10.5152/tpd.2018.5608
No portion of this article can be reproduced without the express written permission from the copyright holder.
Copyright 2018 Gale, Cengage Learning. All rights reserved.

Article Details
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Title Annotation:Original Investigation / Ozgun Arastirma
Author:Jegede, Henry Olanrewaju; Tome, Beatriz; Perera, Ana; Omobowale, Temidayo O.
Publication:Turkish Journal of Parasitology
Date:Mar 1, 2018
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