Aegla expansa (Crustacea: Decapoda: Anomura: Aeglidae) �una especie microendémica del sur de Chile que �logra sobrevivir en humedales de alta intervención antrópica

Abstract

Los cangrejos de agua dulce de la familia Aeglidae están restringidos al sur de América del Sur (Chile, Brasil, Bolivia, Uruguay, Paraguay y Argentina) y todos pertenecen al género Aegla, que contiene más de 60 especies. En Chile los humedales de agua dulce, ribereños y lacustres, poseen 18 especies de crustáceos malacostráceos con un alto nivel de endemismo, que tendrían relaciones zoogeográficas arcaicas de tipo gondwánico. Aegla expansa fue descrita, a partir del exoesqueleto y el espécimen tipo (holotipo) de la especie provino de un sector al este de Hualqui en la región del Biobío en el sur de Chile. En una excursión a la terra típica (2000), solo se lograron recolectar dos juveniles de filiación incierta constatándose que el estero estaba prácticamente destruido por la actividad antrópica. La especie fue clasificada como Extinta en la Naturaleza en 2002 y años más tarde el holotipo y los dos juveniles no clasificados ya mencionados fueron destruidos por un incendio en la institución que los conservaba. No se volvieron a recolectar ejemplares, hasta que en una expedición en 2013 recolectamos A. expansa en esteros de la comuna de Hualqui, por lo que en 2014 se descartó la extinción y se clasificó como En Peligro, estatus legal actual en Chile. Los objetivos de este trabajo son caracterizar las nuevas localidades de registro, construir un mapa de distribución, caracterizar los hábitats de ocupación, estimar su densidad y proponer medidas de conservación de esta especie amenazada.

Full text

Crustaceana 93 (1) 33-39
CHROMOSOME COUNTS IN AEGLA EXPANSA JARA, 1992 (ANOMURA,
AEGLIDAE) A FRESHWATER CRAB FROM SOUTHERN CHILE
BY
PEDRO JARA-SEGUEL1,2), ANDRÉS MUÑOZ-PEDREROS1,3,7),
HERALDO V. NORAMBUENA4), CARLOS JARA5),
MARILYN GONZÁLEZ-URRUTIA4)and MARCELA GUERRERO6)
1)Núcleo de Estudios Ambientales (NEA), Temuco, Chile
2)Departamento de Ciencias Biológicas y Químicas, Facultad de Recursos Naturales, Universidad
Católica de Temuco, Chile
3)Departamento de Ciencias Ambientales, Facultad de Recursos Naturales, Universidad Católica
de Temuco, Chile
4)Centro de Estudios Agrarios y Ambientales (CEA), Valdivia, Chile
5)Universidad Austral de Chile, Valdivia, Chile
6)Facultad de Agronomía y Ciencias Forestales, Universidad de La Frontera, Temuco, Chile
ABSTRACT
The freshwater crab Aegla expansa Jara, 1992 (Aeglidae) is endemic to Chile and has a
geographical distribution restricted to one stream in the locality of Hualqui (37°S), Concepción
Province, Chile. A chromosome number 2n=154 is here described for somatic cells of A. expansa.
The chromosomes are large in number and small in size (<2μm). These cytogenetic data are the
first reported for a species of the genus Aegla.
Key words. — Chromosome number, Crustacea, Decapoda, Aegla expansa
RESUMEN
La pancora de agua dulce Aegla expansa Jara, 1992 (Aeglidae) es endémico de Chile y tiene
restringida distribución geográfica en estero de la localidad de Hualqui (37°S), provincia de
Concepción, Chile. Se describe un número cromosómico 2n=154 en células somáticas de A.
expansa. El número de cromosomas es alto siendo pequeños en tamaño (<2μm). Estos datos
citogenéticos son los primeros reportados para una especie del género Aegla.
Palabras clave. — Número cromosómico, crustáceos, decápodos, Aegla expansa
7)Corresponding author; e-mail: amunoz@uct.cl
©Koninklijke Brill NV, Leiden, 2020 DOI 10.1163/15685403-00003959
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34 PEDRO JARA-SEGUEL ET AL.
INTRODUCTION
The chromosome number (CN) is the most basic feature of the genome of a
species, and is also technically the easiest to obtain (Peruzzi et al., 2012). In many
groups of animals and plants, the CN has been a fundamental feature for analysing
genetic variation at inter-specific level (Lee, 1999; Méndez et al., 2001; Park et al.,
2002; Jara-Seguel et al., 2010; Peruzzi et al., 2011, 2012) and resolving taxonomic
conflicts (Lopes et al., 1998; Windham & Yatskievych, 2003; Guerra, 2012;
Gavrilov-Zimin, 2017). In addition, cytological and evolutionary studies have
included the CN as a fundamental character in addition to karyotype morphology,
chromosome banding, DNA C-values, and gene sequencing (Spotorno et al., 2001;
Souza et al., 2015). Chromosome counts have also been an initial step in the
discovery of polyploidy in mammals, thus disproving the prevailing dogma that
diploidy would be a stable feature within the class (Contreras et al., 1990; Gallardo,
1999).
There are relatively few cytogenetic studies of crustaceans; they are difficult
to perform due to the large number of chromosomes in these species (Radwan
et al., 2014). For this reason, cytogenetic information is only available for ca.
280–300 species across different orders of crustaceans worldwide (Makino, 1956;
Niiyama, 1959a,b; Nakamura et al., 1988; Coluccia et al., 2001; Scalici et al.,
2010; Mansouri et al., 2012; Radwan et al., 2014; Vilar et al., 2014; Jeffery et al.,
2018) representing ca. 0.42% of the total number of species (ca. 67 000 species).
Anomura is an infraorder of the Decapoda for which information on chromosome
number is only available for six species belonging to five families (Niiyama,
1959a,b), i.e., no more than 0.24% of all species of this infraorder worldwide (ca.
2500 species).
Aegla is a freshwater genus belonging to Anomura; it includes 78 species, all
inhabiting South America. The genus has a wide geographical distribution from
20°S in Brazil to 50°S in Chile, and species of Aegla play an important role in
lake, river, and stream environments. Aegla is the only living genus within the
Aeglidae, which also includes the two extinct genera, Protaegla and Haumuriaegla
(Bueno et al., 2016). A. expansa is an endemic Chilean species of Aegla, with a
limited geographical distribution in the south of the country. It is endangered, and
its biological aspects are only poorly known (Jara, 1992).
In this work we describe observations of the chromosome number of A. expansa.
This is the first chromosome count reported for the genus Aegla, thus contributing
with a basic genomic character, that in the future may be applied to understanding
other biological aspects of these freshwater crustaceans.
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CHROMOSOMES OF AEGLA EXPANSA 35
MATERIAL AND METHODS
Five specimens of Aegla expansa were collected in a stream called San Onofre,
Puente Negro, Hualqui, Concepción Province, southern Chile (37°0026S
72°5437W) (fig. 1) using a fine mesh handheld net (approx. 5 mm) of 1 m2.
Live samples were kept in a tank for 24 hours under laboratory conditions. Three
male and two female specimens, with lengths between 20 mm and 35 mm, were
injected intraperitoneally with a 0.01% colchicine solution (0.2 ml per specimen).
The injected specimens were allowed to swim in a well aerated aquarium for 2 h,
then killed humanely and their gills were cut out. The gill tissue was kept for 1 h at
room temperature under hypotonic treatment in potassium chloride (KCl) 0.75%,
then transferred to Carnoy’s fixative solution (Ethanol-Glacial acetic acid 3 : 1, v/v)
with three changes at intervals of 15 min. Mitotic metaphases were obtained by the
drop method, placing small portions of tissue previously suspended and macerated
in Glacial acetic acid 50% (v/v) on slides heated to 40°C on a heating plate. The
slides were then air-dried, stained with 4% Giemsa in Sorenson’s phosphate buffer
for 10 min, and differentiated in distilled water. The chromosomes were examined
and photographed by means of an Olympus CX31 microscope connected to an
Olympus C5050 digital camera. A chromosome count was carried out on enlarged
printed images of four metaphases for each individual (20 metaphases in total).
Fig. 1. Map showing the location of the San Onofre stream, in the locality of Hualqui, Biobío Region,
Chile. A, Habitat of the San Onofre stream; and, B, specimen of Aegla expansa Jara, 1992, collected
in the field.
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36 PEDRO JARA-SEGUEL ET AL.
RESULTS AND DISCUSSION
Fifty metaphases were obtained in total from the gill tissue of five specimens of
Aegla expansa, although only 20 metaphases with no superimposed chromosomes
were selected for the chromosome count. Chromosome number and size of the
chromosomes are two cytogenetic aspects discussed here for A. expansa.
The chromosome counts carried out in A. expansa showed a modal number
of 2n=154 (12 metaphases) in a range between 141 and 164 chromosomes.
Chromosome loss in metaphases with 2nnumbers lower than the modal num-
ber of 154 was possible due to a technical artefact during dropping of cells
(6 metaphases), whereas in metaphases with 2nnumbers higher than the modal
number (2 metaphases), chromosomes were superimposed on those of nearby cells.
The somatic chromosome number of A. expansa described here using gill-cell
drops was higher than that described using histological sections in a testicle of
Cervimunida princeps Benedict, 1902, which showed 2n=109 chromosomes
in spermatogonial division and with a chromosome formula of primary spermato-
cytes n=53 +X1+X2+Y(Niiyama, 1959b). In the case of A. expansa,sex
chromosomes were cytologically indistinguishable in the somatic metaphases anal-
ysed (fig. 2). C. princeps is a marine species belonging to the family Galatheidae,
which is close to Aeglidae according phylogenies based on DNA sequences (Pérez-
Losada et al., 2002). Such differences in CN between A. expansa and C. princeps
are to be expected, because of their family-level separation, although they may be
a consequence of chromosome re-arrangement, e.g., chromosome fusions, which
may have occurred during the evolution of the species within each family, resulting
Fig. 2. Giemsa-stained metaphase of gill cells of Aegla expansa Jara, 1992, 2n=154. Scale bar =
10 μm.
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CHROMOSOMES OF AEGLA EXPANSA 37
TABLE I
Chromosome number for Aegla expansa Jara, 1992, and for various
other Anomura species, taken from Niiyama (1959a,b)
Family/species 2n
AEGLIDAE
Aegla expansa Jara, 1992 154
COENOBITIDAE
Coenobita rugosa H. Milne Edwards, 1837 230
GALATHEIDAE
Cervimunida princeps Benedict, 1902 109
LITHOIDAE
Paralithodes camtschaticus (Tilesius, 1815) 208
P. platypus (J. F. Brandt in Von Middendorf, 1851) 206
PAGURIDAE
Eupagurus ochotensis (Brandt, 1851) 254
in some cases in a reduction in 2nnumbers. However, this assumption is still spec-
ulative for Aegla species, and for Anomura in general, because of the scarcity of
studies on karyotype morphology carried out to date. Several studies in decapods
using modern cytogenetic methods (e.g., chromosome banding and fluorescence)
have revealed the occurrence of chromosome fusions during karyotype evolution
in members of the families Astacidae, Parastacidae (Mlinarec et al., 2016), and
Palaemonidae (Torrecilla et al., 2017), so similar chromosome change mechanisms
cannot be excluded for Anomura. Table I summarizes the family variation in CN
observed within Anomura, showing evidence that may support these ideas.
A. expansa presented small chromosomes, not exceeding 2 μm in size. This
makes their separation difficult in many of the preparations obtained. Unfortu-
nately, the lack of data currently available on chromosome size in species be-
longing to Anomura makes comparisons between species difficult. Within the de-
capods, chromosome size has been described for some Astacidae species belong-
ing to the genus Austropotamobius (2n=176) where the largest chromosome
pair does not exceed 1.7 μm in size (Mlinarec et al., 2016). In general, the chro-
mosome size in decapods varies between 1 and 5.4 μm, as described by several
authors (Lécher et al., 1995; Coluccia et al., 2001; Jeffery et al., 2016).
Following these preliminary cytogenetic data reported for A. expansa, the next
step for future research will be to study meiosis, karyotype morphology, and
DNA C-values by classical methods. Modern techniques that have previously been
used in other crustacean groups to evaluate chromosome re-arrangements and sex
chromosomes (Vilar et al., 2014; Mlinarec et al., 2016; Torrecilla et al., 2017)
could also be performed. The cytogenetic data obtained in these ways may increase
knowledge of the genome of A. expansa,aswellasofthatofotherAegla species,
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38 PEDRO JARA-SEGUEL ET AL.
and this may have multiple applications for understanding aspects of the biology
of the representatives of this genus.
ACKNOWLEDGEMENTS
The authors are grateful to the project “Information collection of species from
the research, generation and/or systematization of information for the conservation
of biodiversity program”, funded by the Ministerio del Medio Ambiente, Chile. We
would also like to acknowledge the Núcleo de Estudios Ambientales (NEA) of the
Facultad de Recursos Naturales, Universidad Católica de Temuco, and the Centro
de Estudios Agrarios y Ambientales (CEA), Valdivia, Chile.
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First received 7 August 2019.
Final version accepted 21 November 2019.
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