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Contents lists available at ScienceDirect

Veterinary Parasitology

jo u r nal homep age: www.elsev ier .com/ locate /vetpar

hort communication

olecular identification of Taenia serialis coenurosis in a wildthiopian gelada (Theropithecus gelada)

ndia A. Schneider-Creasea,∗,1, Noah Snyder-Macklera,1,ulie C. Jarveyb, Thore J. Bergmanc,d

Department of Evolutionary Anthropology, Duke University, Durham, NC 27708, United StatesSchool of Natural Resources and Environment, University of Michigan, Ann Arbor, MI 48109, United StatesDepartment of Ecology and Evolutionary Biology, University of Michigan, Ann Arbor, MI 48109, United StatesDepartment of Psychology, University of Michigan, Ann Arbor, MI 48109, United States

a r t i c l e i n f o

rticle history:eceived 24 May 2013eceived in revised form 13 August 2013ccepted 21 August 2013

a b s t r a c t

Since morphological identification of a larval Taeniid in geladas (Theropithecus gelada)has produced inconsistent results, genetic information is pivotal for species identifica-tion. Nuclear and mitochondrial DNA from a coenurus in a wild gelada were compared

eywords:aenia serialiseladaolecular phylogeny

to published sequences from multiple Taeniid species, confirming the identification of thisparasite as Taenia serialis. A demographic analysis finds age to be a strong predictor ofcoenuri. Tapeworms rarely employ primates as intermediate hosts, and the presence ofT. serialis in a wild gelada population may indicate a substantial ecological shift in thisparasite’s life cycle.

ulticeps

. Introduction

Taeniid parasites globally exploit mammalianredator–prey relationships, requiring a carnivorousefinitive host for the adult form and an herbivorous inter-ediate host for the larval form (Meyer, 1955). Although

rimates are not known to regularly act as intermediateosts, the intermediate stage of one Taeniid has beenescribed in the terrestrial and herbivorous Ethiopianelada (Theropithecus gelada) (Schwartz, 1927; Urbain andullier, 1935; Elek and Finkelstein, 1939; Rodhain andanson, 1954; Clark, 1969; Oshawa, 1979; Dunbar, 1980).

Please cite this article in press as: Schneider-Crease, I.A., et al.,

wild Ethiopian gelada (Theropithecus gelada). Vet. Parasitol. (201

Geladas become infected when they ingest tapewormggs shed in the feces of the definitive host. In interme-iate hosts, the oncospheres in the eggs are released into

∗ Corresponding author at: India Schneider-Crease, Department ofvolutionary Anthropology, Duke University, 104 Biological Sciencesuilding, Box 90383, Durham, NC 27708, United States.el.: +1 919 684 4124; fax: +1 919 660 7348.

E-mail address: [email protected] (I.A. Schneider-Crease).1 Co-first authors.

304-4017/$ – see front matter © 2013 Elsevier B.V. All rights reserved.ttp://dx.doi.org/10.1016/j.vetpar.2013.08.015

© 2013 Elsevier B.V. All rights reserved.

the intestinal tract, burrow through the intestinal mucosa,and settle in the connective tissue or intermuscular fas-cia. Each oncosphere develops into a coenurus, a mass ofmultiple invaginated scoleces in a fluid-filled membranethat expands through the branching and invagination ofendogenous daughter cysts. When a carnivore ingeststhe infected tissue, each scolex attaches to the intestinalmucosa to sprout the strobila, a series of hermaphroditicand self-fertilizing segments that constitute the tapewormbody (Meyer, 1955).

Morphological identifications of this parasite in cap-tive geladas have been inconsistent. Thus, the addition ofgenetic information to species identification is critical. Agenetic identification of the parasite and an analysis of thepatterns of disease in a wild population of geladas in theSimien Mountains National Park (SMNP), Ethiopia, are per-formed here.

Molecular identification of Taenia serialis coenurosis in a3), http://dx.doi.org/10.1016/j.vetpar.2013.08.015

2. Materials and methods

A multiloculated cyst from a dead 13-year-old malegelada in the SMNP was procured in November 2011. One

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Fig. 1. Microscope view (10×) of a scolex with a branching endogenous

Fig. 2. Phylogenetic relationships between four species of Taenia (T. seri-alis, T. multiceps, T. crassiceps, T. pisiformis) and the sample obtained forthis study (“sample from gelada”). The tree is based on maximum par-simony using published partial 12S and ITS-2 sequences obtained fromGenBank (T. pisiformis: ITS-2 JX317674, 12S DQ104230; T. crassiceps: ITS-

stood to be synonymous, and Taenia is the preferred

daughter cyst from a wild Ethiopian gelada. Photograph by Dr. JamesFlowers.

of the authors (J.C.J.), extracted tissue from a protuberantcoenurus on the left ventral forelimb (Fig. 1), and storedit in RNAlater (Applied Biosystems/Ambion, Austin, TX,U.S.A.).

DNA was extracted from the cestodes using the Qia-gen DNeasy Blood and Tissue Kit (Qiagen). 347 bp of themitochondrial rDNA (12S) region and 434 bp of the secondinternal transcribed spacer of nuclear rDNA (ITS-2) wereamplified and sequenced using the primers and PCR pro-tocol described in Padgett et al. (2005). Sequences werealigned using Sequencher v5.0 (Gene Codes Corporation,Ann Arbor, MI) and blasted against the NCBI nucleotide databank.

For the phylogenetic analysis, two nucleotide sub-stitution models were selected using AIC and BIC asimplemented in jmodeltest (Darriba et al., 2012). Bothmodels were run using the Bayesian program BEAST v1.7.5(Drummond et al., 2012). The MCMC chain length was setto 108 and the state was recorded every 103, resultingin 105 trees. The effective sample size for all parame-ters was greater than 130. After discarding the first 102

trees as burn-in, the remaining trees were summarizedinto one consensus tree using TreeAnnotater v1.7.5, whichwas imported into FigTree v1.4 (http://tree.bio.ed.ac.uk/)for visualization (Fig. 3).

To measure the disease prevalence in the SMNP,researchers surveyed 291 geladas for coenuri. The Uni-versity of Michigan Gelada Research Project has collectedbehavioral data on this population for seven years, andresearchers are trained to recognize individuals with100% accuracy based on morphological traits. Researchersrecorded the presence/absence, size, and location ofcoenuri for each known individual. Recorded cysts wereconfirmed by two independent observers.

The prevalence rates given are of obvious signs of dis-ease, not infection, since infected individuals may notexhibit coenuri. To ascertain patterns of coenuri across ages

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wild Ethiopian gelada (Theropithecus gelada). Vet. Parasitol. (20

and sexes, Pearson’s chi-squared tests were run using JMPstatistical software (JMP, Version 7. SAS Institute Inc., Cary,NC, 1989–2007).

2 DQ099564, 12S EU219547; T. multiceps: ITS-2 FJ886762, 12S JQ710642;T. serialis: ITS-2 DQ099575, 12S DQ104236). Numbers above branchesrepresent the posterior support (max of 1) for each branch.

3. Results

The BLAST search revealed a strong match betweenthe sample sequences (ITS-2: GenBank ID KF414738; 12S:GenBank ID KF414739) and published sequences for Tae-nia serialis ITS-2 (99% nucleotide identity; GenBank IDDQ099575) and 12S (99% nucleotide identity; GenBank IDDQ104236) regions (Jia et al., 2010). The next most closelyrelated species was Taenia multiceps, with which the sam-ple had 95% and 90% nucleotide identity at 12S and ITS-2,respectively (GenBank IDs: GQ228818, FJ886762) (Padgettet al., 2005).

Phylogenetic reconstruction with published TaeniidITS-2 and 12S sequences confirms that this sample is moreclosely related to T. serialis than to any other Taeniid species(Fig. 2). AIC and BIC supported two different models, K81 + Iand K80 + G; however, both models supported the samephylogeny as implemented in BEAST v1.7.5 (Drummondet al., 2012).

The prevalence rate of disease for the population was4.8% (Table 1). The prevalence rate was 9.9% (13/131)among adults (over ∼3 years old), and 0.9% among juve-niles (approx. 1.5–3 years old) (1/110). Coenuri were notobserved in infants (0–1.5 years old) (n = 50). Adults weresignificantly more likely to display coenuri than juvenilesand infants (Pearson’s �2 = 13.661, P = 0.001, d.f. = 2). Theprevalence rate for adult females was 10.8% (10/93), and formales was 7.9% (3/38). There was no significant differencein the disease rate between males and females (Pearson’s�2 = .247, P = 0.620, d.f.=1).

4. Discussion

The contribution of a genetic component to the iden-tification of T. serialis in geladas is imperative becausemorphological approaches have produced inconsistentresults. The first studies on this parasite classified it underthe genus Multiceps; now Multiceps and Taenia are under-

Molecular identification of Taenia serialis coenurosis in a13), http://dx.doi.org/10.1016/j.vetpar.2013.08.015

nomenclature (Meyer, 1955; Benger et al., 1981). While fivestudies used scolex arrangement and the size and numberof rostellar hooks for identification (Schwartz, 1927; Urbain

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Table 1Prevalence of T. serialis coenuri in geladas across age-sex classesa.

Sex AgeInfant Juvenile Adult All

Female 0% (0/17) 1.8% (1/55) 10.8% (10/93) 6.7% (11/164)5)

/110)

class.

aWt1cR

FBJ

Male 0% (0/33) 0% (0/5Both 0% (0/50) 0.9% (1

a Number of individuals with coenuri/total individuals in that age-sex

nd Bullier, 1935; Elek and Finkelstein, 1939; Rodhain andanson, 1954; Clark, 1969), others used the morphology of

he hook guards (Schwartz, 1927) or hook blades (Meyer,

Please cite this article in press as: Schneider-Crease, I.A., et al.,

wild Ethiopian gelada (Theropithecus gelada). Vet. Parasitol. (201

955). Other parasitologists assert that no morphologicalriterion reliably diagnoses Taenia species (Beveridge andickard, 1976).

ig. 3. A Female gelada with facial coenurus. Photograph by Dr. Jacintaeehner. B Female gelada with mammary coenurus. Photograph by Dr.

acinta Beehner.

7.9%(3/38) 2.4% (3/126)9.9% (13/131) 4.8% (14/291)

The demographic analysis shows that T. serialis coenuriare significantly more likely to be found in adult thanin subadult or infant geladas, corroborating two previousstudies of this population (Oshawa, 1979; Dunbar, 1980).The chronic physiological and social stresses of adulthoodmay have immunosuppressive effects that increase adultsusceptibility (Muehlenbein and Bribiescas, 2005). Alter-natively, the time necessary for development of noticeablecysts may exceed the geladas’ juvenile period.

T. serialis coenuri can displace viscera and exert mechan-ical pressure on nerves and arteries, causing spastic limbparalysis, muscle atrophy, increased vulnerability to pre-dation, and death (Scott, 1926; Elek and Finkelstein, 1939).Coenuri may also impact fitness by stimulating an ener-getically costly immune response, requiring an individualto prioritize feeding and resting over reproductive behav-iors (Oshawa, 1979). The presence and consequences of T.serialis in this population will continue to be monitored.

Other diseases may present similarly to T. serialiscoenuri, and, if so, were erroneously included in this anal-ysis. However, given the history of cysts whose grossmorphology in geladas is similar to those observed at thissite (Fig. 3) and which have been identified morphologi-cally as Taenia or Multiceps serialis, we are confident thatthe cysts reported are indicative of T. serialis infection. Thisinterpretation is strengthened by the genetic confirmationof T. serialis from a characteristic cyst in this population.

5. Conclusion

A genetic analysis confirms the presence of T. seri-alis in geladas, and a demographic analysis shows thatadults are more likely to exhibit coenuri than non-adults.However, the biology of this T. serialis life cycle is notyet entirely understood. We speculate that the defini-tive host is the predatory spotted hyena (Crocuta crocuta)or the scavenging black-backed jackal (Canis mesomelas)based on previous reports of T. serialis in these species(Hürni and Stiefal, 2003). Future work should identify thedefinitive host and elucidate the risks for T. serialis infec-tion in other primates. Anthropogenic habitat change mayincrease geladas’ vulnerability to parasites, and changesin host-parasite dynamics may put humans at risk foremerging infectious diseases. The ecology of parasite-hostinteractions in this region must be understood in order topredict and control the spread of disease.

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Conflict of interest

No personal or financial relationships influenced thisresearch.

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Acknowledgements

We thank the Ethiopian Wildlife Conservation Author-ity, the Simien Mountains National Park, and the Universityof Michigan Gelada Research Project. We thank Alison VonStriver, Caitlin Barale, Charles Nunn, Jacinta Beehner, JamesFlowers, Jenny Tung, Leslie Digby, and Paul Durst for theirhelp in the field, the laboratory, and during manuscriptpreparation, and the three anonymous reviewers for theirhelpful comments. Research was approved by the Univer-sity Committee on Use and Care of Animals (UCUCA no.09554) at the University of Michigan, the University ofPennsylvania Institutional Animal Care and Use Committee(IACUC no. 802996), and adhered to the laws and guidelinesof Ethiopia.

References

Benger, A., Rennie, R.P., Roberts, J.T., Thornley, J.H., Scholten, T., 1981. Ahuman coenurus infection in Canada. Am. J. Trop. Med. Hyg. 30 (3),638.

Beveridge, I., Rickard, M., 1976. The development of Taenia pisi-formis in various definitive host species. Int. J. Parasitol. 5,633–639.

Clark, J.D., 1969. Coenurosis in a Gelada Baboon (Theropithecus gelada). J.Am.Vet. Med. Assoc. 155 (7), 1258–1263.

Darriba, D., Taboada, G.L., Doallo, R., Posada, D., 2012. jModelTest: more

Please cite this article in press as: Schneider-Crease, I.A., et al.,

wild Ethiopian gelada (Theropithecus gelada). Vet. Parasitol. (20

models, new heuristics and parallel computing. Nat. Methods 9 (8),772.

Drummond, A.J., Suchard, M.A., Xie, D., Rambaut, A., 2012. Bayesian phy-logenetics with BEAUti and the BEAST 1.7. Mol. Biol. Evol. 29 (8),1969–1973.

PRESSarasitology xxx (2013) xxx– xxx

Dunbar, R.I.M., 1980. Demographic and life history variables of a pop-ulation of gelada baboons (Theropithecus gelada). J. Anim. Ecol. 49,485–506.

Elek, S.R., Finkelstein, L.E., 1939. Multiceps serialis infestation in a baboon.Report of a case exhibiting connective tissue cystic masses. Zoologica24, 323–328.

Hürni, H., Stiefel, S.L., 2003. Report on a Mission to the Simien Moun-tains National Park World Heritage Site, Ethiopia. University of Berne,Switzerland, Report for NCCR North-South and the East & SouthernAfrica Partnership Programme of the Centre for Development andEnvironment.

Jia, W.Z., Yan, H.B., Guo, A.J., Zhu, X.Q., Wang, Y.C., Shi, W.G., Chen, H.T.,Zhan, F., Zhang, S.H., Fu, B.Q., Littlewood, D.T., Cai, X.P., 2010. Completemitochondrial genomes of Taenia multiceps, T. hydatigena and T. pisi-formis: additional molecular markers for a tapeworm genus of humanand animal health significance. BMC Genomics 11, 447.

Meyer, M.C., 1955. Coenuriasis in varying hare in Maine, with remarkson the validity of Multiceps serialis. Trans. Am. Microsc. Soc. 74 (2),163–169.

Muehlenbein, M.P., Bribiescas, R.G., 2005. Testosterone-mediatedimmune functions and male life histories. Am. J. Hum. Biol. 17 (5),527–558.

Oshawa, H., 1979. The local gelada population and environment of theGish area. In: Kawai, M. (Ed.), Ecological and Sociological Studies ofGelada Baboons. Karger Publishers, Tokyo, pp. 4–45.

Padgett, K.A., Nadler, S.A., Munson, L., Sacks, B., Boyce, W.M., 2005. Sys-tematics of Mesocestoides (Cestoda: Mesocestoididae): evaluation ofmolecular and morphological variation among isolates. J. Parasitol. 91(6), 1435–1443.

Rodhain, J., Wanson, M., 1954. Un nouveau cas de coenurose chez lebabouin, Theropithecus gelada RUPPELL. Riv. Parasitol. 15 (4), 613–620.

Schwartz, B., 1927. A subcutaneous tumor in a primate caused by tape-worm larvae experimentally reared to maturity in dogs. J. Agric. Res.,

Molecular identification of Taenia serialis coenurosis in a13), http://dx.doi.org/10.1016/j.vetpar.2013.08.015

U.S. Dept. Agric. 35 (5), 471–480.Scott, H.H., 1926. Report on deaths occurring in the Society’s Gardens

during the year 1925. Proc. Zool. Soc. Lond., 231–244.Urbain, A., Bullier, P., 1935. Un cas de cenurose conjonctive chez un Gelada.

Bull. Acad. Vét. France 8 (6), 322–324.