This study aimed to analyze a case of a high level of infection with acanthocephalans in order to

understand if the infection pattern is related to biological characteristics of the hosts. Acanthocephalans of

33 specimens of Rhinella marina (Linnaeus, 1758) from Huanuco, Peru, were collected and identified;

Spearmans' rank test (r) was then used to calculate possible correlations between the host body size and

parasite abundance, and between parasite body size and parasite abundance. Helminths were identified as

Pseudoacanthocephalus lutzi (Hamann, 1891). The infection prevalence was 97.1%. A total of 874

helminths was recovered, with a mean abundance of 24.1 ± 4.7 and mean intensity of infection of 24.8 ±

4.8. The host body size and parasite body size were not significantly correlated with parasite abundance.

The present study, apart from contributing to our knowledge of the helminth fauna of amphibians in Peru,

has shown that R. marina has a high rate of helminth infection with P. lutzi in that region.

ISSN Versión impresa 2218-6425 ISSN Versión Electrónica 1995-1043

ORIGINAL ARTICLE / ARTÍCULO ORIGINAL

1UNESP - Univ Estadual Paulista, Campus de Botucatu, Instituto de Biociências, Departamento de Parasitologia, Botucatu,

São Paulo, Brazil.

2UFPI - Universidade Federal do Piauí, Piauí, Brazil.

3URP - Universidad Ricardo Palma, Facultad de Ciencias Biológicas, Lima, Perú.

4UNFV – Universidad Nacional Federico Villarreal, Facultad de Ciencias Naturales y Matemática, Lima, Perú.

5Wabash College, Lima, Peru.

6UNHEVAL - Universidad Nacional HermilioValdizán, Huanuco, Perú.

*Correspondence author: gisatoledo@hotmail.com

ABSTRACT

Key words: Helminth – Huanuco – Infection pattern – Parasite abundance

Neotropical Helminthology

405

Neotropical Helminthology, 2017, 11( ), j - : 2 ul dic 405-411

1 2 3,4

Gislayne de Melo Toledo *; Mariluce Gonçalves Fonseca ; Jose Iannacone ;

5 6 1

Jorge Manuel Cárdenas Callirgos ; Carlos Pineda Castillo & Reinaldo José da Silva

INFECTION WITH PSEUDOACANTHOCEPHALUS LUTZI (HAMANN, 1891) (ACANTHOCEPHALA:

ECHINORHYNCHIDAE) IN RHINELLA MARINA (LINNAEUS, 1758) (AMPHIBIA: BUFONIDAE) IN

PERU

INFECCIÓN CON PSEUDOACANTHOCEPHALUS LUTZI (HAMANN, 1891) (ACANTHOCEPHALA:

ECHINORHYNCHIDAE) EN RHINELLA MARINA (LINNAEUS, 1758) (AMPHIBIA: BUFONIDAE) EN

PERU

INTRODUCTION

406

RESUMEN

Palabras clave: abundancia de parásitos – helmintos – Huanuco – patrón de infección

Este estudio tuvo como objetivo analizar un caso de alto nivel de infección con acantocéfalos para

comprender si el patrón de infección está relacionado con las características biológicas de los hospederos.

Se recolectaron acantocéfalos de 33 ejemplares de Rhinella marina (Linnaeus, 1758) de Huanuco, Perú, y

luego se utilizó la prueba de rango de Spearmans (r ) para calcular posibles correlaciones entre el tamaño

del cuerpo del hospedero y la abundancia de parásitos, y el tamaño del cuerpo del parásito y la abundancia

de parásitos. Los helmintos fueron identificados como Pseudoacanthocephalus lutzi (Hamann, 1891). La

prevalencia de infección fue del 97,1%. Se recuperó un total de 874 helmintos, con una abundancia media

de 24,1 ± 4,7 y una intensidad media de infección de 24,8 ± 4,8. El tamaño del cuerpo del huésped y el

tamaño del cuerpo del parásito no se correlacionaron significativamente con la abundancia del parásito. El

presente estudio, además de contribuir a nuestro conocimiento de la fauna helmíntica de anfibios en Perú,

ha demostrado que R. marina tiene una alta tasa de infección por helmintos con P. lutzi en esa región.

1995; Gutierrez et al., 2005; Arredondo & Gil de

Pertierra, 2009), Uruguay (Cordero, 1933), Peru

(Naupay, 1973; Tantaleán, 1976; Barrera et al.,

1988; Tantaleán et al., 2005), Paraguay, and Brazil

(Smales, 2007).

During a study on the helminth fauna of

amphibians from Peru, an interesting case was

observed in the municipality of Huanuco, in which

a population of toads R. marina presented a great

amount of acantocephalans in their intestines. The

aim of the present study is to analyze this

acantocephalan infection case in order to

understand if the infection pattern is related to the

biological characteristics of the hosts.

A total of 35 specimens of the toad R. marina were

colleted in April 2014, in Estanque La Pedroza,

Centro Poblado La Esperanza, Amarilis, Huanuco,

Peru (09°58'30" LS and 76°14'25" LW, 1,910

mols).

The amphibians were sampled by active search at

night and transported live to the laboratory where

they were euthanized with sodium thiopental

solution, in accordance with Resolution No. 301

Acanthocephala is a group poorly known in

amphibians and reptiles of the South American

(Campião et al., 2014). Adult acanthocephalans

parasitize the intestine of vertebrates and use

invertebrates as intermediate hosts, where larval

development occurs (Tantaleán et al., 2005; Santos

& Amato, 2010). Acanthocephalans do not have

free-living larval stages, and a paratenic host can be

present in the life cycle of some species (Santos &

Amato, 2010). According to Bush et al. (2001) and

Kennedy (2006), vertebrates are the known

paratenic hosts, which harbor the cystacanth larval

stage in an extra-intestinal locationand usually

differ from known definitive hosts (Al-Jahdali et

al., 2015).

Amin (2013) listed 24 families, 157 genera, and

1,298 species of acanthocephalans. The genera

Pseudoacanthocephalus Petrochenko, 1958

e n c o m p a s s 1 8 d e s c r i b e d s p e c i e s .

Pseudoacanthocephalus lutzi (Hamann, 1891) was

originally described as Echinorhynchus lutzi

Hamann, 1891 from the cane toad Rhinella marina

(Linnaeus, 1758) in Brazil (Hamann, 1891). This

worm was transferred to Acanthocephalus

Koelreuther, 1771 by Mey er (1 932 ).

Pseudoacanthocephalus lutzi has been reported

infecting various amphibians and reptiles in

Argentina (Lajmanovich & Martinez de Ferrato,

Neotropical Helminthology, 2017, 11(2), jul-dic

MATERIALS AND METHODS

Toledo et al.

Description

Pseudoacanthocephalus lutzi (Fig. 1)

Syns.: Echinorhynchus lutzi Hamman, 1891;

Acanthocephalus saopaulensis Smales, 2007.

Based on 10 specimens (5 females and 5 males).

General: Trunk and all shared structures larger in

females than in males; trunk about twice as large in

females. Proboscis cylindrical, with 14–16

longitudinal and regularly alternating rows of 5 to 7

hooks each. Hook length increases from apex to

middle part of proboscis. Roots about half as long

as blades, simple, spatulate, directed posteriorly.

Proboscis receptacle cylindrical, double-walled

with cellular elements associated with retractor

muscles just exterior to its posterior tip. Proboscis

retractor muscles inserted at base of proboscis

receptacle and at anterior third of the trunk.

Lemnisci claviform extending shortly past

posterior end of receptacle.

Males (based on five mature specimens): Trunk

8.35–10.00 (9.39) mm long and 1.35–1.85 (1.533)

mm wide. Proboscis, 519–649 (564) long, with

14–18 hook rows, each with 5–7 hooks each.

Proboscis receptacle 838–1,153 (1,021) long.

Lemnisci 870–1,063 (945) long. Testes ovoid, in

tandem, contiguous; anterior testis 657–734 (686)

long, 392–512 (434) wide; posterior testis 626–846

(708) long, 380–564 (446) wide. Cement glands in

compact cluster, in number 4. Reproductive system

completely post-equatorial. Posterior end sigmoid-

shaped. Genital pore nearly terminal (Figs. 1A-C).

Female (based on five mature specimens): Trunk

12.58–14.99 (13.68) mm long and 1.54-1.68 (1.60)

mm wide. Proboscis, 442–647 (535) long, with

14–18 hook rows, each with 5–7 hooks each.

Proboscis receptacle 958–1,073 (1,015) long.

Lemnisci 816–1,211 (998) long. Genital pore

ventral, sub-terminal (Figs. 1D-E).

The prevalence of infection was 97.1% for 35 frogs

examined. A total of de 842 helminths were

recovered, with a mean abundance of 24.1 ± 4.7

and a mean intensity of infection of 24.8 ± 4.8. The

acanthocephalans were found in the small intestine

and large intestine.

and Ordinance No 148/2012 of the Conselho

Federal de Biologia – CFBio. The snout–vent

length (SVL) in mm and body mass in g were

recorded. Then they were necropsied and the

gastrointestinal tract was examined for the

presence of helminths. The collection was

authorized by Director's Resolution N°024-2014-

SERFOR-DGGSPFFS.

Acanthocephalans found in the intestine were

collected and placed plates containing distilled

water, and kept in the refrigerator until the

proboscis evertion for 24 h of exposure. After they

were fixed 100% ethyl alcohol and then preserved

in 70% ethyl alcohol. Some specimens were

stained with carmine and cleared in creosote for

identification procedure.

For parasite identification, morphological and

morphometric were performed in a computerized

system for image analysis LAS V3 (Leica

Application Suite) adapted in microscope

DM5000B with differential interference contrast

(DIC). All measurement of parasites is in u at less

was indicated otherwise.

Helminth specimens will be deposited at the

Coleção Helmintológica do Instituto de

Biociências de Botucatu (CHIBB) at the

Universidade Estadual Paulista, São Paulo state,

Brazil, and at the Colección Helmintológica y de

Invertebrados Relacionados del Museo de Historia

Natural (UNMSM) at the Universidad Nacional

Mayor de San Marcos, Lima, Peru.

Prevalence, mean intensity of infection, and mean

abundance were calculated according to Bush et al.

(1997). All values corresponding to the mean of

any variable are accompanied by the respective

standard error.

Spearmans' rank test (r) was used to calculate

posssible correlations between the host body sizes

and parasite abundances, and between parasite

body sizes and parasite abundances. For this

analysis, the body measurements of 33 specimens

of R. marina were used. Regarding the parasites,

875 specimens of acanthocephalans were

measured for length and width; and the means were

obtained for each host specimen. The Mann-

Whitney U test was used to test for possible

differences between abundance and host sex.

407

Neotropical Helminthology, 2017, 11(2), jul-dic

RESULTS

Pseudoacanthocephalus lutzi in Rhinella marina

The total lengths of toad ranged 60-100 (86.5 ± 8.6)

mm, and their weights ranged 33-76 (50.2 ± 11.9)

g, including 14 females and 19 males. The host

body size was not significantly correlated with the

parasites abundance (lengths: r= 0.16, n=33, p=

0.35; weights: r= 0.14, n = 33, p= 0.43), as well as

the parasite body size showed no correlation with

parasite abundance (lengths: r= 0.15, n = 32, p=

0.41; width: r= 0.18, n = 32, p= 0.31). The parasite

abundance did not show difference between the

sexes of hosts (U = 119; p = 0.61).

408

Neotropical Helminthology, 2017, 11(2), jul-dic

D E

Figure 1. Pseudoacanthocephaluslutzistained with carmine from Rhinella marina:(A) Male – overview; (B) Anterior end

ofmale; (C) Posterior end of male; (D) Anterior end of female; (E) Posterior end of female.

Toledo et al.

obtain food from the few permanent water bodies

(Evans & Lampo, 1996). This suggests that

aquatic insects may act as intermediate hosts for P.

lutzi (Arredondo & Pertierra, 2009).

The present study, apart from contributing to our

knowledge of the helminth fauna ofamphibians in

Peru, has shown that R. marina has a high rate of

helminth infection for P. lutziin that region. Our

results also show that there is no influence of host

body size and sex on the parasite abundance for this

parasite species of R. marina.

G. M. Toledo thanks the Conselho Nacional de

DesenvolvimentoCientífico e Tecnológico (CNPq)

by thePhD.doctoral scholarship (140301/2013-5).

R. J. Silva is grateful to Pro-Reitoria de Pós-

graduação da UNESP (PROPG-UNESP), CNPq

(3 07808 /20 14- 9) an d CNPq -PR OTA X

(440496/2015-2) / FAPESP 2016/50377-1.

409

Neotropical Helminthology, 2017, 11(2), jul-dic

Helminths found in the intestine were identified as

P. lutzi, according to Smales (2007) and Amin &

Heckmann (2014). Specimens of P. lutzi show

marked intraspecific morphological variations

inrows number (12–18) and hooks number per row

(5–7), size and extension of lemnisci, number of

cement glands (4–6), and size of eggs in various

host species and geographical locations (Amin &

Heckmann, 2014).

The abundance and community structure of

amphibian parasites is influenced mainly by the

probability of individual hosts acquiring the

species of parasites, which in turn is influenced by

the ecological requirements of host species

(Janovy et al., 1992). It has been suggested that the

size of the body of amphibians influences the

number of helminth species and the abundance of

parasites (Muzzal, 1991; McAlpine, 1997; Bolek

& Coggins, 2000; Yoder & Coggins, 2007;

Hamann et al., 2010, 2012, 2013; Toledo et al.,

2015). However, in the present study, the size

(length and weight) of R. marina was not a

determinant factor for the abundance of P. lutzi

parasites, not corroborating with the hypothesis

that the larger size can harbour higher infections,

leading to higher parasitism index values.

Acanthocephalan adult sizes correlated positively

with host mass, but also this correlation depends of

cystacanth size and egg size (Poulin et al., 2003).

In the current study, parasite body size showed no

correlation with parasite abundance of P. lutzi.

Relationships between sex and helminth infection

show no clear patterns in amphibians. In some

cases, the levels of parasitism by sex depended on

the studied species (Comas et al., 2014). In this

research, parasite abundance of P. lutzi did not

show differences between sexes of R. marina.

The type host of P. lutzi is R. marina, a native toad

of Central and South America. It lives in a wide

variety of habitats including savannah woodlands,

forests, scrub lands, and arid and semiarid areas.

This species mainly feeds on terrestrial insects

such as ants and beetles (Weber, 1938), but it also is

an opportunist with a highly diversified diet (Zug &

Zug, 1979). The aquatic odonata larvae are prey

items in arid areas, indicating that R. marina may

DISCUSSION

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Pseudoacanthocephalus lutzi in Rhinella marina