Johnsprentia copemani gen. nov., sp. nov. (Haemoproteidae), a parasite of the flying-fox, Pteropus alecto (Pteropidae) from Queensland Irene LANDAU Jean-Marc CHAVATTE Parasitologie comparee et modeles experimentaux, USM307, Museum National d’Histoire Naturelle, CP52, 61 rue Buffon, 75231 Paris Cedex 05, France. Email: [email protected] Ian BEVERIDGE Department of Veterinary Science, University of Melbourne, Parkville Vic 3052, Australia. Citation: Landau, I., Chavatte, J.-M. & Beveridge, I. 2012 02 17: Johnsprentia copemani gen. nov., sp. nov. (Haemoproteidae), a parasite of the flying-fox, Pteropus alecto (Pteropidae), from Queensland. Memoirs of the Queensland Museum - Nature 56(1): 61-66. Brisbane. ISSN 0079-8835. Accepted: 5 March 2010. ABSTRACT Black flying foxes (Pteropus alecto Temminck, 1837) captured in Townsville, Queensland, are parasitised by a new species of Haemoproteidae which differs morphologically and in its tissue localisation, having highly distinctive gametocytes and tissue forms in the lungs, which are described here. It cannot be accommodated in any of the genera of the Haemoproteidae known from mammals and is here named Johnsprentia copemani gen. nov., sp. nov. (cid:9633) Haemoproteidae, Pteropus alecto, Johnsprentia copemani, new genus, new species, Queensland. Knowledge of the haemoproteid parasites types of schizonts which differed in their size, their morphology and their localisation. of native Australian mammals is limited, with all those known to date being reported We describe here the gametocytes and schizonts by O'Donoghue & Adlard (2000). Studies on of one of these haemoproteid species. It cannot flying foxes are particularly limited although be accommodated in any of the known genera. one species, Hepatocystis pteropi (Breinl 1913), Here we describe and name it as Johnspreittia has been reported from a number of species of gen. nov. copemani sp. nov. after two noted flying foxes (Mackerras 1959). Australian parasitologists, J.F.A. Sprent and D.B. Copeman. The remaining two species will be We had the opportunity to examine the haemo¬ the subject of a later paper. proteid parasites of several black flying foxes, Pteropus alecto Temminck, 1837, over a period of time in captivity. It was apparent that, depending MATERIALS AND METHODS upon the individual, one to three species of haemoproteids were present, at differing levels, in Eleven Pteropus alecto Temminck (1837) captured their blood. Histological examination of internal in Townsville using a mist net and exhibiting organs using serial sections also revealed three a parasitaemia with Haemoproteidae, were tran- Memoirs of the Queensland Museum | Nature • 2012 • 56(1) • www.qm.qld.gov.au 61 Landau, Chavatte & Beveridge sported to the Museum National d'Histoire Johnsprentia copemani sp. nov. Naturelle, Paris, shortly after their capture, (Figs 1-2) arriving on the 15 December 1978 and the 07 Material. HOLOTYPE. Histological section of June 1979 respectively. Blood samples from schizont in lung of Pteropus alecto no. 409)(F each animal were collected by pricking the autopsied on 29/06/1979 in Paris (origin: radial vein and smeared onto a slide, air dried Townsville, Queensland, Australia, slide quickly, fixed with absolute methanol and deposited in QM No. G465432, illustrated in stained by Giemsa stain (8% in buffer phosphate, Fig. 2B). PARATYPES: a) One microgametocvte pH7.4). They were examined over a period marked on a blood smear from the same bat, collected on the same date, deposited in QM of several months. At autopsy, internal organs No. G465433; b) a blood smear and histological were fixed in Carnoy's fluid and serial sections sections of schizonts from the lung of Pteropus of each organ were stained by the giemsa- alecto no. 408XF and 20HD, deposited in MNHN colophonium method (Bray & Garnham 1962; PXX 201-208. Garnham 1966) and examined for tissue stages of the parasites. Type material has been deposited Ety mology. Named after our colleague the late Dr. D. Bruce Copeman who helped capture flying foxes in in the Queensland Museum, Brisbane (QM) and his garden in Townsville. the Museum National d'Histoire Naturelle Description. Gametocytes. Young forms or (MNHN), Paris. those which have just reached maturity are described from an animal (408XF) parasitised SYSTEMATICS only by Jolmsprentia (Fig. 1A-1H). This animal, on its arrival in Paris, had only ring-forms Phylum: Apicomplexa (Sporozoa) and trophozoites. It was examined daily until autopsy 21 days later. The immature parasites Class: Coccidea developed slowly and on the day of autopsy, several fully-developed gametocytes were observed. More mature gametocytes were found Order: Haemosporida in other animals in which additional species of parasite were present (Fig. 1I-M). The young Family: Haemoproteidae trophozoites are round or oval with a thin, arcuate nucleus along one side, sometimes Jolmsprentia gen. nov. with two unequal masses of chromatin. In young trophozoites, there are frequently fine Definition. Haemoproteidae with elongated cytoplasmic projections (Fig. 1B-D), which schizonts, in the lungs of Megachiroptera, are absent in older forms. The cytoplasm is not producing colloid, gametocytes in blood clear, pale-blue and with numerous poorly- films with a nucleus apparently adherent to defined vacuoles, some vacuoles being larger the margin of the parasite and with peripheral and better defined with fine grains of pigment. regions denser than the centre. When the gametocyte reaches two-thirds the volume of the erythrocyte, it is spherical, clear Type Species, jolmsprentia copemani sp. nov. in the centre, denser around the periphery, Etymology. Named after the late Professor John F.A. with the cytoplasm and most of the pigment Sprent, formerly Professor of Parasitology at the displaced towards the periphery. The centre University of Queensland. has scattered inconspicuous vacuoles and fine, 62 Memoirs of the Queensland Museum | Nature • 2012 • 56(1) Johnsprentia copemani gen. nov., sp. nov. (Haemoproteidae) FIG. 1. Drawings of the gametocytes of Johnsprentia copemani sp. nov. stained with Giemsa. A, E, F, young gametocytes; B-D, young gametocytes with fine cytoplasmic prolongations; G-I, K, microgametocytes; J, old microgametocyte; L-M, macrogametocytes; N, O, normal red blood cell. scattered grains of pigment. At this stage, it always peripheral, forming an elongated arc is difficult to differentiate microgametocytes around the border. The macrogametocytes and macrogametocytes (Fig. 1E-G). The fully (Fig. 1M, N) are generally clearer than the developed microgametocytes are highly microgametocytes, with a smaller nucleus. The chromophilic, purple, with a dark, dense red globular surrounding layer is of normal periphery (Fig. 1H, I, K). This is not due to size, rarely slightly larger, of usual colour and overstaining as young forms in the same film is always visible around the parasite. do not stain in the same manner. As with the younger forms, small, scarcely visible vacuoles Schizonts. The schizonts are described from bat are scattered through the cytoplasm and the number 408 XF (Fig. 2). They occurred exclusively fine pigmentation is found mainly around the in the lungs where they were numerous. They periphery. Old microgametocytes are often are elongate, between the alveoli, probably surrounded by a reddish ring. The nucleus is in endothelial cells. They are sinuous with a Memoirs of the Queensland Museum | Nature • 2012 • 56(1) 63 Landau, Chavatte & Beveridge FIG 2. Photomicrographs of schizonts of Johnsprentia copemani sp. nov. in the lungs of Pteropus alecto. A-C, immature schizonts; D, detail of 'C' showing pseudosepta; E-F, immature schizont in consecutive sections showing the sinuous nature of the schizont; G, almost mature schizont with clear cytoplasm and punctiform nuclei; H, transverse section of young schizont showing large, dense, well defined nuclei. 64 Memoirs of the Queensland Museum | Nature • 2012 • 56(1) Johnsprentia copemani gen. nov., sp. nov. (Haemoproteidae) pink border measuring 1.0-1.5 ^im in thickness. have not been reported in any other species Because of their sinuous form it is difficult described from Pteropus or any other bats, and to determine their precise size. The largest have not been observed in other haemoproteids schizont in section (Fig. 2A) measured 160 from mammals. In addition, the schizonts are }im long, 28 ^im in maximum width and was exclusively pulmonary, elongated and devoid of seen in 14 sections 5 [im in thickness. Very colloid. young schizonts (Fig. 2H) have a dark blue They are differentiated from the other genera cytoplasm, with large, dense chromatin masses of haemoproteids by the following characters: which fragment progressively as the schizont Hepatocystis (Laveran 1899) parasitic in matures. Individual nuclei are initially relatively numerous groups of mammals (primates, large with dense chromatin and a well-defined bats, Sciuridae, hippopotamus, tragulids): the outline (Fig. 2E, F). The centre of the nucleus classical merocysts of Hepatocystis develop in becomes clear and pink, the chromatin forms hepatocy tes, are generally very large (can attain 2 aggregations of variable shape around the clear mm in diameter in primates), frequently expand or pink centre. In some schizonts, pseudosepta into neighbouring tissues and secrete a colloidal are seen (Fig. 2B-D). A fully-developed substance inside or around the schizont; the schizont (Fig. 2G) measured 140 \xm long schizonts of Johnsprentia are localised in the and 26 jam wide, the cytoplasm was a clear lungs and are much smaller, are compact and blue, finely granular with small punctiform, elongate and do not secrete colloid. uniformly distributed nuclei. There was no Nycteria Garnham & Heisch (1953), parasites histomacrophagocytic reaction around the of micro-bats in which the schizonts localise in parasite. No schizonts were found in hundreds hepatocytes and are rounded or lobed, while of sections of other organs of this bat (liver, those of Johnsprentia localise in the lungs and spleen, kidney). are elongate. DISCUSSION Polychromophilus Dionisi (1899), the schizonts are, (as with those of Johnsprentia) pulmon¬ In cases of polyparasitism, such as is ob¬ ary, but by contrast, those of Polychromophilus served in flying foxes in Queensland, it can occur equally frequently in other organs be difficult to link the tissue stages with the such as the kidney, spleen, liver and even the corresponding gametocytes and it is necessary adrenals. The schizonts of Polychromophilus are to connect histological observations on the tissues ellipsoidal and surrounded by a thick, bright¬ with those made on the erythrocytes. In the case ly pink capsule, their cytoplasm is poorly of the species described here, we were fortunate chromophilic and their nuclei are small, even to be able to study one bat at the commencement in young forms. The schizonts of Johnsprentia of an infection with only one type of gametocyte are exclusively pulmonary, are elongated and and schizont. Their morphology was uniform and botuliform, their host cell is poorly visible, their therefore we are convinced that they belong to limiting membrane is thin, the cytoplasm of a single species. the immature stages stains intensely blue with Giemsa and their nuclei are relatively large. The distinctive features of this parasite are primarily those of the gametocyte, in which Bioccala Landau et al. (1980), parasites of the nucleus is elongated along the periphery microbats. The small schizonts of Bioccala (Mer of the cell and part of the cytoplasm is also & Goldblum 1947; Landau et al. 1980) are concentrated at the periphery where it forms a disseminated throughout the body and do not dense, chromophilic band. These characteristics resemble the current parasite in any way. Memoirs of the Queensland Museum | Nature • 2012 • 56(1) 65 Landau, Chavatte & Beveridge Biguetiella Landau et al. 1984, again has schiz- ACKNOWLEDGEMENTS onts that are very small and intra-hepatocytic, Animals were collected and exported with bearing no resemblance to the current parasite. appropriate State and Federal collection and The schizonts of Rayella (Dasgupta 1967), export permits form rounded, intrahepatocytic groups, very different from the isolated, elongated pul¬ LITERATURE CITED monary schizonts of Johnsprentia. Bray, R.S. & Garnham, P.C.C. 1962. The Giemsa- Dionisia Landau et al. 1980. The schizonts of Colophonium method for staining protozoa in tissue section. Indian Journal of Malariolovy Dionisia are rounded or oval, small in size and 16:153-155. localise in the lumen of the hepatic vasculature, Dasgupta, B. 1967. A new malarial parasite of the in a host cell which is hypertrophic and surround¬ flying squirrel. Parasitology 57(3): 467-474. ed by a thick capsule. The schizonts of Johnsprentia Garnham, RC.C. 1966. Malaria parasites and other do not possess any of these characters. Haemosporidia. (Blackwell: Oxford) Garnham, RC.C. & Heisch, R.B. 1953. On a new blood In fact, the most similar schizonts morph¬ parasite of insectivorous bats. Transactions of the ologically are those of Parahaemoproteus of birds, Royal Society of Tropical Medicine and Hygiene 47: 357-363. such as those described from the musculature Landau, I., Baccam, D., Ratanaworabhan, N., Yenbutra, of Psittacula roseata in Thailand, and which S., Boulard, Y. & Chabaud A.G. 1984. Nouveaux are transmitted by Culicoides (Miltgen et al. Haemoproteidae parasites de Chiropt£res en 1981). Although larger, as they can attain a size of Thailande. Annales de Parasitologic Humaine et 900 gm, they are similar to those of Pteropus in Comparee 59: 437-447. Landau, I., Chabaud, A.G., Miltgen, F. & Baccam, D. being elongated, botuliform, highly chromophilic, 1980. Dionisia bimoi n. gen., n. sp. Haemoproteidae with pseudosepta, have a thin outer membrane parasite du Microchiroptere Hipposideros cyclops and their host cell is scarcely visible. There is, in au Gabon. Annales de Parasitologic Humaine et Comparee 55: 271-280. neither species, an accumulation of macrophages prior to the rupture of the schizont. Landau, I., Chabaud, A.G., Miltgen, F., Hugot, J.R, Leger, N., Beveridge, I. & Baccam, D. 1980. Sur The obvious similarity between the schizonts le genre Polychromophilus (Haemoproteidae, parasite de Microchiroptdres). Annales de of Johnsprentia and those of Parahaemoproteus Parasitologic Humaine et Comparee 55:13-32. desseri could indicate a close relationship, one Mackerras, M.J. 1959. The Haematozoa of Australian parasitic in mammals, the other in birds. Such a mammals. Australian Journal of Zoology 7:103-135. phenomenon is possible since we know that, in Mer, G.G & Goldblum, N. A haemosporidian of bats. the case of the haemosporidians, the ancestral 1947. Nature (London) 159: 444. host is an invertebrate, probably Culicoides in Miltgen, F., Landau, I., Ratanaworabhan, N. & Yenbutra, S. 1981. Parahaemoproteus desseri n. sp. this case, and not the vertebrate host. However, Gametogonie et schizogonie chez 1'hote naturel: this hypothesis is unlikely as the morphology Psittacula roseata de Thailande et sporogonie of the gametocytes is of greater phylogenetic experimentale chez Culicoides nubeculosus. Annales de Parasitologic Humaine et Comparee 56:123-130. value than that of the schizonts, which, in O' Donoghue, P.J. & Adlard, R.D. 2000. Catalogue this case, suggests that the two taxa are quite of protozoon parasites recorded in Australia. different phylogenetically. Memoirs of the Queensland Museum 45:1-163. 66 Memoirs of the Queensland Museum | Nature • 2012 • 56(1)