Global Ecology and Conservation 4 (2015) 146–149

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Global Ecology and Conservation

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Original research article Gastro-intestinal helminths of goliath frogs (Conraua goliath) from the localities of Loum, and Nkondjock in the Littoral Region of Cameroon Nguiffo Nguete Daniel, Wabo Pone Josue ∗, Mpoame Mbida Laboratory of Biology and Applied Ecology, Department of Animal Biology, Faculty of Science, University of Dschang, PO Box 067 Dschang, Cameroon article info a b s t r a c t

Article history: The gastro-intestinal helminth parasites of goliath frog (Conraua goliath) from the Littoral Received 10 June 2015 Region of Cameroon in the Localities of Loum, Yabassi and Nkondjock were surveyed. Out Received in revised form 26 June 2015 of the 30 goliath frogs examined (13 males and 17 females) between April and May 2013, Accepted 30 June 2015 26 (85%) contained at least one helminth parasite and yielded a total of nine hundred and Available online 7 July 2015 seventy three (973) helminths comprising: Nematodes (90.5%), Trematodes (9.4%) and Pen- tastomids (0.1%). Nematodes included: Africana taylori (60%), Oswaldocruzia perreti (0.2%), Keywords: Aplectana sp. (21%), Gendria sp. (7.1%), Amphibiophilus sp. (0.2%), Strongyluris sp. (0.1%), Gastro-intestinal helminths Conraua goliath Physalopteroides sp. (1.6%), and Oxyuridae gen. sp. (0.3%). Trematodes comprised: Meso- Species richness coelium sp. (7.3%) and Diplodiscus subclavatus (2.1%). Pentastomids were represented by Diversity Sebekia sp. (0.1%). The mean species richness and diversity were 1.97±2.12 and 0.41±0.04 Cameroon respectively. The intensity of parasite infection was correlated with host body weight, pos- itively for Africana taylori, Aplectana sp., Diplodiscus subclavatus, and Mesocoeliumsp. Infec- tion rates were influenced by land-use pattern. Thus higher prevalences were observed in Loum (intensive agricultural area). © 2015 The Authors. Published by Elsevier B.V. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).

1. Introduction

Cameroon has a diverse Anuran fauna with at least 200 different species (Depierre, 1978). The goliath frog is the largest known frog with a maximum weight of 3.3 kg (Boulenger, 1906) and is endemic in Cameroon and Equatorial Guinea. These frogs are classified as endangered species by International Union for Conservation Nature (IUCN) and as protected species of class ‘A’ by the Wildlife Law in Cameroon. Little information is available on the helminth parasites of amphibians from Cameroon (Dollfus, 1929, Gassmann, 1975, Durette-Desset and Vaucher, 1979, McAllister et al., 2010). To the best of our knowledge, the parasites of goliath frogs have not been studied. The objective of this study therefore was to establish an initial helminth list for C. goliath and also determine their helminth richness and diversity.

2. Materials and methods

The goliath frogs were collected from 3 localities of the Littoral Region of Cameroon: Loum (4°42′58′′N, 9°44′11′′E) with 300 m of altitude, Yabassi (4°45′N, 9°96′67E) and Nkondjock (4°87′36′′N, 10°24′86′′E). Mean annual precipitation in this

∗ Corresponding author. E-mail address: [email protected] (W.P. Josue). http://dx.doi.org/10.1016/j.gecco.2015.06.009 2351-9894/© 2015 The Authors. Published by Elsevier B.V. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/ licenses/by-nc-nd/4.0/). N.N. Daniel et al. / Global Ecology and Conservation 4 (2015) 146–149 147

Table 1 Infection pattern of Conraua goliath by gastro-intestinal tract helminths from Loum, Yabassi and Nkondjock in the Littoral Region in Cameroon. Helminths Sites of infection Localities Loum Yabassi Nkondjock Prévalence Mean Prévalence Mean Prévalence Mean intensity n(%) intensity of n(%) intensity of n(%) of infection infection infection

Nematoda Gendria sp. Large intestine 61(9.6) 8.71 ± 6.6 1(0.3) 1 ± 0.00 5(11.3) 1.66 ± 1.15 Aplectana sp. Large intestine 161(25.4) 32.2 ± 47.27 40(15) 9.8 ± 7.66 3(6.8) 3 ± 0.00 Africana taylori Large intestine 368(58.1) 90 ± 130.83 182(61.3) 28.71 ± 21.91 34(77.3) 4.85 ± 3.02 Amphibiophilus sp. Large intestine 2(0.3) 2 ± 0.00 0(0) 0 ± 0 0(0) 0 ± 0 Strongyluris sp. Large intestine 0(0) 0 ± 0 1(0.3) 1 ± 0 0(0) 0 ± 0 Physalopteroides sp. Large intestine 0(0) 0 ± 0 16(4.9) 16 ± 0 0(0) 0 ± 0 Oxyuridae gen. sp. Large intestine 0(0) 0 ± 0 3(0.9) 3 ± 0 0(0) 0 ± 0 Oswaldocruzia perreti Small intestine 2(1.3) 8 ± 0.00 0(0) 0 ± 0 0(0) 0 ± 0 Trematoda Mesocoelium sp. Small intestine 13(2) 0 ± 0 57(17.3) 19 ± 18.73 1(2.3) 1 ± 0.00 Diplodiscus Large intestine 20(3.2) 2.6 ± 2.6 0(0) 0 ± 0 0(0) 0 ± 0 subclavatus Pentastomids Sebekia sp. (nymph) Large intestine 0(0) 0.0 ± 0.00 0(0) 0 ± 0 1(2.3) 1 ± 0.00 Species richness 2.36 ± 2.76 2.37 ± 2.44 1.18 ± 2.13 Diversity 0.46 ± 0.05 0.46 ± 0.05 0.31 ± 0.03 n = intensity.

Region is about 3909.7 mm with annual temperature ranging between 20 and 27 °C(Rouche, 2012). The predominant vegetation in the study area is forest (semi deciduous and biafrean). A total of 30 goliath frogs (13 males and 17 females) were sampled as follows: Loum 11, Yabassi 8 and Nkondjock 11. They were collected between April and May 2013 by hand and peach net during the night. Frogs were transported to the laboratory and anesthetized with benzocaine. Postmortem examination was carried out within 18 h of collection. The gastro-intestinal tract (GIT) divided into small and large intestine was opened using the small scissors and the exudate examined. After successive washings and sedimentation of the intestinal contents, helminths were collected using a pair of tongs (Meyer and Olsen, 1980; Euzeby, 1982). Nematodes were fixed in hot (65 °C) AFA (alcohol formalin acetic acid) solution and thereafter conserved in vials containing fresh 70% alcohol. Nematodes were cleared in pure glycerin. Trematodes and Pentastomids were also conserved in vials with fresh 70% alcohol. Before examination they were dehydrated by enhanced concentration of alcohol (35%, 70%, 80%, 95%, and 100%) for 1 h in each concentration, colored by Semichon Carmine acetic acid and cleared by xylene (Meyer and Olsen, 1980; Euzeby, 1982). Prevalence is defined as the percentage of frog infected with a helminth species and mean intensity as the mean number of a helminth species in infected frog. Species richness is the number of helminth species per host (Bush et al., 1997). The diversity was calculated by Shannon’s Weaver index accounts for both abundance and evenness of the species present. The proportion of species i relative to the total number of species (pi) was calculated, and then multiplied by the natural logarithm of this proportion (ln pi). The resulting product was summed across species, and ′  multiplied by −1. (H = − Pi log2 Pi.) Helminths were identified by observation and measurement of morphological characteristics based on descriptions from literature (Dollfus, 1929, Popova, 1965, Maeder, 1973, Gassmann, 1975, Chabaud, 1978, Durette-Desset and Vaucher, 1979, Baker, 1981, Schmidt and Robert, 1981, Soulsby, 1982, Euzeby, 1982, Durette-Desset, 1983, Smyth, 1996, Aisien et al., 2009 and Imasuen and Aisien, 2012). Determination of the genera and species was confirmed by Charles Bursey of the Pennsylvania State University.

3. Results

The GIT helminth communities of goliath frogs consisted of 11 species (Table 1): 8 Nematodes, (Africana taylori, Oswal- docruzia perreti, Aplectana sp., Gendria sp., Amphibiophilus sp., Strongyluris sp., Physalopteroides sp., and Oxyuridae gen. sp.). 2 Trematodes (Mesocoelium sp. and Diplodiscus subclavatus) and 1 larval Pentastomids (Sebekia sp.). 85.95% of goliath frogs were infected with one or more helminth species (100% in Loum, 90% in Yabassi and 67.85% in Nkondjock). The infection pattern in different localities is presented in Table 1. Some species were characteristic of localities like O. perreti, Amphibio- philus sp. and D. subclavatus which were confined in Loum whereas Physalopteroides sp., Strongyluris sp. and Oxyurid were restricted to Yabassi, nymph of Sebekia sp. found only in Nkondjock. Other helminths like Mesocoelium sp., Gendria sp. and Africana taylori occurred in the 3 localities. The trematodes such as Mesocoelium sp. were more prevalent in Yabassi (17.3%) and Nkondjock (2.3%). 148 N.N. Daniel et al. / Global Ecology and Conservation 4 (2015) 146–149

The mean helminth richness was 2.36 ± 2.76; 2.37 ± 2.44 and 1.18 ± 2.13 respectively in Loum, Yabassi and Nkondjock. However, the mean overall richness was 1.97 ± 2.12. The mean helminth diversity was the same at Loum and Yabassi (H = 0.46) and lower at Nkondjock (H = 0.31). Concerning infection in relation to host body weight, correlations between species were positive and not significant (P > 0.05) for Mesocoelium sp. (r = 0.19) and significant (P ≤ 0.05) for A. taylori (r = 0.7), Aplectana sp. (r = 0.75) and D. subclavatus (r = 0.79).

4. Discussion

The helminth species found in goliath frogs in the present study are most similar to those observed amongst the amphibian hosts in other African countries except for the nymph of Sebekia sp. which has several fish species as intermediate host (Christoffersen and De Assis, 2013). The occurrence of this nymph as observed during this study could be due to the fact that the goliath frogs shared the same habitat with crocodiles (definitive host) and fishes (intermediate host). Frogs from Loum and Yabassi had the highest overall helminth diversity with species richness of 2.36 and 2.37 respec- tively whereas those from Nkondjock had the lowest diversity and richness. This observation shows the importance of vari- ation in host habitat due to the degree of practicing agricultural activities, deforestation and poaching in different localities. Nguessu(2013) examined the liver weight of goliath frogs (increase with accumulation of toxic product) in same localities and recorded a significant difference at Loum where intensive agriculture is practiced compared to Yabassi and Nkondjock. The helminths that have direct life cycles may be an important factor contributing to species richness and community complexity. In the present study, 5 helminths species (45%) all of which are nematodes (Gendria sp, Aplectana sp., A. taylori, O. perreti, Amphibiophilus sp., Strongyluris sp., and Oxyuridae gen. sp.) are believed to have direct life cycles (Anderson, 2000). Gendria sp., Aplectana sp. and A. taylori composing 27% of all helminths found in this study were frequent in their occurrence (Loum, Yabassi and Nkondjock) and contribute to the complexity of helminth communities of goliath frogs. Seven species (O. perreti, Amphibiophilus sp., Strongyluris sp., Physalopteroides sp., Oxyuridae gen. sp., D. subclavatus, Sebekia sp.) were found only at one locality. The absence of helminths in C. goliath from one or more localities may be due to the lack of a favorable environment for growth by either the parasite, an appropriate host, or both. The occurrence of O. perreti in frogs only in Loum may be due to the fact that Loum is an agricultural zone and it is highly deforested. Aisien et al.(2011) and McKenzie(2007) had observed Oswaldocruzia sp. only in agricultural zones and pasture environments respectively. It is likely that, this parasite is more frequent in agricultural and polluted environments. Regarding the helminth that occurred in frogs at all locations, the degree to which each habitat (locality) was preferred by these species, as indicated by prevalence and mean intensity, did vary considerably. This again indicates the importance of local factors. In conclusion goliath frogs showed a wide variation in helminthic fauna. Observations from this study are indicative of some relationship between the pattern of parasitism and land use by goliath frogs in Littoral Region of Cameroon. The ecology of goliath frog (habitat and diet in particular) and helminth transmission dynamics are important in shaping helminth community structure. Moreover anthropogenic land use effects and their associated impact on water bodies are important co-factors related to the abundance of helminths in goliath frogs like it was the case in Loum.

Acknowledgments

We are very grateful to Dr. Charles Bursey (Pennsylvania State University), USA for his assistance in helping us to identify the isolated helminths; to the Ministry of Forestry and Wildlife who permitted us to work on this frog and to Dr. Kana Raphael and Nguessu Charles (University of Dschang) for their assistance. We also thank Dr. Khan Payne of the Animal Biology Department for reading and correcting the manuscript.

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