1Dipartimento di Biologia Animale e dell’Uomo, Universita di Roma �La Sapienza�, Rome, Italy; 2Biology Department, Addis AbabaUniversity, Addis Ababa, Ethiopia; 3Pest Management Centre, Sokoine University of Agriculture, Morogoro, Tanzania;4Department of Zoology, Kenyatta University, Nairobi, Kenya

Chromosomal diversity in the genus Arvicanthis (Rodentia, Muridae) from EastAfrica: a taxonomic and phylogenetic evaluation

R. Castiglia1, A. Bekele2, R. Makundi3, N. Oguge

4 and M. Corti1

AbstractIn this paper we discuss the contribution of cytogenetics to the systematics of Arvicanthis in East Africa, by reviewing all the known chromosomalcytotypes of the genus in the area. We also provide G- and C-banding comparisons for two recently described karyotypes, provisionally namedANI-5 (2n ¼ 56, NFa ¼ 62) and ANI-6 (2n ¼ 60, NFa ¼ 72). This, therefore, brings the total number of known cytotypes in this area to 10. Fiveof these correspond to the species recognized by the latest rodent checklist, i.e. A. nairobae (2n ¼ 62, NFa ¼ 78), A. neumanni (2n ¼ 52–53,NFa ¼ 62), A. blicki (2n ¼ 48, NFa ¼ 62), A. abyssinicus (2n ¼ 62, NFa ¼ 64) and A. niloticus (2n ¼ 62, NFa ¼ 60–62). The taxonomic statusof the remaining five cytotypes (A. cf. somalicus, 2n ¼ 62 NFa ¼ 62–63; ANI-5, 2n ¼ 56, NFa ¼ 62; ANI-6/6a 2n ¼ 60, NFa ¼ 72/76; ANI-7,2n ¼ 56, NFa ¼ 78; and ANI-8, 2n ¼ 44, NF ¼ 72) is discussed. Finally, we reconstruct the phylogenetic relationships among all the knownkaryotypes on the basis of banding data available for the genus in Africa and show the occurrence of two main clades, each characterized bydifferent types of chromosomal rearrangements. The times of the cladogenetic events, inferred by a molecular clock, indicate that karyotypeevolution has accomplished almost all the dichotomic events from the end of the Miocene to the present day. The discovery of a largechromosomal differentiation between populations showing low genetic distances and intrapopulation chromosomal polymorphism suggests thatthe process of chromosomal differentiation in Arvicanthis is still ongoing and may possibly be responsible for speciation.

Key words: Arvicanthis – cytogenetics – cytotaxonomy – karyotype evolution – chromosomal phylogeny

A taxonomic overview of the genus Arvicanthis

The correct evaluation of species distribution and their

taxonomic status is a fundamental prerequisite in assessingbiodiversity in any geographical region. This constitutes themain challenge prior to any conservation planning and/or

biodiversity management. For some groups this objective isproblematical due to the occurrence of cryptic species andtherefore taxonomic uncertainty. In rodents this is commonbecause of the high morphological uniformity compared to

important genetic diversity (Taylor 2000; Corti 2001; Granjonand Dobigny 2003). The high frequency of cryptic speciesoccurring in syntopy or in allopatry, makes the classical

morphological approach inadequate to determine the entirerodent diversity in many geographical areas. Morphology onits own, therefore, cannot form the basis of future biodiversity

studies. In the last 30 years, however, molecular and cyto-genetic techniques have greatly improved the taxonomicresolution for many genera (Corti 2001). These studies haveshown that species recognition requires a multidisciplinary

approach based on cytogenetics (see for example Taylor 2000),the estimate of genetic divergence and DNA taxonomy(Blaxter 2004) and morphometrics (Loy et al. 2004).

The African Arvicanthine rats of genus Arvicanthis Lesson,1842 represent an emblematic example of this kind of problem.Several species are widely distributed in all the sub-Saharan

African savannahs, along the Nile valley and down to theZambezi. Their opportunistic and generalized diet makes themvery common in agricultural fields and particularly in staple

crops where they cause serious pre- and post-harvest damage(Stenseth et al. 2003).

The systematics of the genus was recently improved bymolecular (Ducroz et al. 1998), cytogenetic (Capanna et al.

1996; Volobouev et al. 2002b; Castiglia et al. 2003) andgeometric morphometric investigations (Fadda and Corti

2001), so that now there are seven recognized species in the

most recent checklist (Musser and Carleton, 2006) versus thefive which had previously been accepted by the same authors(Musser and Carleton 1993).This increase in species number depends on the studies

carried out recently in West Africa. In fact, throughcytogenetics and molecular genetics (Volobouev et al. 1987;Civitelli et al. 1995; Ducroz et al. 1997, 1998) four distinct

species, chromosomally well differentiated, have been identi-fied which were previously included in the single speciesA. niloticus (Desmarest, 1822). These karyotypic variants

have been reviewed by Volobouev et al. (2002b) and assignedto A. niloticus (2n ¼ 62; autosomal fundamental number,NFa ¼ 62–64; Niger, Chad, Mali, Senegal, Burkina Faso,

Mauritania), A. rufinus (Temminck, 1853) (2n ¼ 62;NFa ¼ 74; Benin), A. ansorgei Thomas, 1910 (2n ¼ 62;NFa ¼ 74–76; Burkina Faso, Mali, Senegal). In addition,another unidentified species has been found in the Central

African Republic, which has been provisionally named ANI-2(2n ¼ 58; NFa ¼ 72).The taxonomic resolution for the East African species is,

however, more complex. Musser and Carleton (2006) recog-nized the occurrence of five species in this region, namelyA. nairobae J. A. Allen, 1909, A. neumanni (Matschie, 1894)

(A. somalicus in Musser and Carleton 1993), A. blicki Frick,1914, A. abyssinicus (Ruppell, 1842) and A. niloticus (Table 1).There are however uncertainties regarding the taxonomy

and systematics of this last species in the region: in fact,

Yalden et al. (1976, 1996) maintained that an additionalspecies occurs in Ethiopia on the basis of morphologicaldifferences, i.e. A. dembeensis (Ruppell, 1842), which in sub-

sequent work was included in A. niloticus on the basis ofchromosomal (Corti et al. 1996) and molecular homogeneity(Ducroz et al. 1998). Moreover, Orlov et al. (1992) and

� 2006 The AuthorsJournal compilation � 2006 Blackwell Verlag, Berlin

Accepted on 31 December 2005JZS doi: 10.1111/j.1439-0469.2006.00356.x

JZS (2006) 44(3), 223–235

Bulatova et al. (2002) showed that under the �A. niloticus–dembeensis� complex there are at least three cryptic taxa inEthiopia when considering the karyotype and morphology of

spermatozoa (Baskevich and Lavrenchenko 1995). Given that,there still remains some taxonomic uncertainty among theA. niloticus–dembeensis complex, we included the two taxa in

A. niloticus (following Corti et al. 1996; Volobouev et al.2002b) while waiting for an eventual clarification fromadditional molecular and morphological data.Molecular-based phylogenies have also greatly improved

systematics of the genus. Two well-supported clades havebeen identified (Ducroz et al. 1998): clade 1, with speciesoccurring in East (A. nairobae) Central (ANI-2) and West

Africa (A. rufinus and A. ansorgei); and clade 2, includingEast African species only, i.e. A. niloticus, A. neumanni,A. abyssinicus. The genetic differentiation between these two

clades is high and their separation occurred about 5.3 Ma(Ducroz et al. 1998). These two clades are also supported bycytogenetics (Volobouev et al. 2002b): all the members of

clade 1 share a reciprocal translocation between chromoso-mal pairs 4 and 7. This is a kind of chromosomal mutationwhich usually causes severe impairment during gametogenesisin structural heterozygotes (Koleva and Benova 1992; King

1993).In Arvicanthis, chromosomal diversification is often accom-

panied by mtDNA divergence. For example, the sister taxa

A. ansorgei/A. rufinus and A. niloticus/A. abyssinicus diverge,respectively, by 8.5–11.1% and 12.8–15.1% in the sequence ofthe complete cytochrome b (Ducroz et al. 1998). However,

inter- and intra-specific values of divergences overlap; forexample, ANI-2 and A. rufinus diverge by 5.4–5.7% only, avalue that falls within the levels of divergence found in

intraspecific and in interspecific comparison (Ducroz et al.1998; Volobouev et al. 2002a). For this reason both karyotypeand molecular genetics analysis are fundamental in delimitingspecies of Arvicanthis.

Because of the importance of karyotype assessment for boththe systematics and taxonomy of the genus, we discuss in thispaper all the known karyotype variability from East Africa

and provide banding data on the karyotypes from threelocalities in Kenya and one locality in Ethiopia, two of whichare new and have recently been described on the basis of

standard techniques only (Corti et al. 2005). Moreover, weattempt to infer phylogenetic relationships among all theknown karyotypes of Arvicanthis for which banding data areavailable. Finally, we discuss the contribution of cytogenetics

in the systematics of Arvicanthis in this part of Africa andprovide a taxonomical evaluation of the chromosomal cyto-type found in the area.

Chromosomal methods and taxonomical evaluationof cytotypes

The new data provided in this paper have been obtainedthrough the analysis of 13 specimens from one locality in

Ethiopia and three localities in Kenya (Fig. 1, Table 2).Species identification was based on morphological com-parisons (Yalden et al. 1976, 1996; W. Verheyen and

J. Hulselmans, personal communication) and our previouschromosomal analysis (Corti et al. 1996; Volobouev et al.2002b; Castiglia et al. 2003).

The chromosomal species/cytotypes that did not correspond

to recognized species have been named using the acronym ANIand a number, following Volobouev et al. (2002b) nomencla-ture for West Africa.

Specimens were transported to the Kenyatta University ofNairobi, Kenya, and the University of Addis Ababa, Ethiopia,for chromosome preparation. Chromosome metaphases were

obtained from bone marrow following Hsu and Patton (1969).Cells suspensions in fixative were then transported to theUniversity of Roma �La Sapienza�, Italy, where light-micro-scope preparations were made. These were stained by the

Giemsa standard method (pH7). G-bands were enhanced withtrypsin following the protocol of Seabright (1971). Theheterochromatic portion of the genome was identified by C-

banding using barium hydroxide 5% (Sumner 1972). Picturesof metaphases were collected using the Photometrics Sensys1600 digital camera (Roper Scientific Photometrics, Tucson,

AZ). Numeration of chromosomes follows that of A. niloticus,considered the ancestral karyotype of the genus (Volobouevet al. 2002b) (Fig. 2).

All chromosomal data for the East Africa localities werereviewed from literature and our findings. Localities (Table 2)are mapped on the physical map shown in Fig. 1. These includesamples fromTanzania,Kenya, Ethiopia,Uganda and Somalia.

Given the importance of chromosomal evolution for thetaxonomy of the genus, we propose a taxonomical evaluationof all the undescribed East African Arvicanthis taxa, identified

by distinct cytotypes.First of all we considered chromosomal differences because

they are believed to act as a post-mating isolation mechanism

which depends on the type and the number of rearrangementsinvolved (reviewed in King 1993). However, this approach hassome limitations because a single hybrid individual betweencytotypes has yet to be identified in Arvicanthis. For this

reason, we take into account the level of chromosomalheterozygosity in putative hybrids between pairs of cytotypesand the associate level of infertility as seen in the literature in

other species for the same structural levels of heterozygosity.We proceeded by matching the above identified cytotypes

with the morphological differences discussed elsewhere (Yal-

den et al. 1976, 1996; Bekele et al. 1993; Fadda and Corti2001; W. Verheyen and J. Hulselmans, personal communica-tion) and, limiting to Ethiopia, also with the morphology of

spermatozoa (Baskevich and Lavrenchenko 1995). Moreover,we took into account the percentage of cytochrome b sequencedivergence between pairs of species calculated by Ducroz et al.(1998). For the two new cytotypes here identified, we also

report the genetic divergence with their sister taxa as describedby Corti (unpublished data) (Table 3).

In evaluating genetic distance as a taxonomic tool we

followed some ideas by Bradley and Baker (2001) whoreviewed a set of genetic distances computed from the

Table 1. Species of Arvicanthis listed in Musser and Carleton (2006)

Taxon Type locality

A. abyssinicus (Ruppell, 1842) Ethiopia, Simien Prov.,Simien Mtns, Entschetqab

A. ansorgei Thomas, 1910 Bissau Guinea, GunnalA. blicki Frick, 1914 Ethiopia, South Chilalo Mtns,

Hora Mt base camp (2743 m)A. nairobae J. A. Allen, 1909 Kenya, NairobiA. neumanni (Matschie, 1894) Central Tanzania, Kondoa District,

BarungiA. niloticus (Desmarest, 1822) EgyptA. rufinus (Temminck, 1853) S Ghana, Elmina

224 Castiglia, Bekele, Makundi, Oguge and Corti

� 2006 The Authors JZS 44(3), 223–235Journal compilation � 2006 Blackwell Verlag, Berlin

cytochrome b sequences in four different rodent genera. Theyfound that the average percentage in sequence divergence

ranges from 0% to 6.3% at the intraspecific level, and from2.7% to 19.2% between sister species. These values also matchquite well with the values found within and among species of

Arvicanthis (Ducroz et al. 1998). However, because of theoverlapping divergence estimates between intraspecific andinterspecific comparison, one should be cautious when using

genetic distances for species identification. Nonetheless, valuesgreater than 10–11% have usually only been found betweendifferent species in mammals (Bradley and Baker 2001).

For the taxa identified in this manner we cannot categor-ically confirm names because the number of forms described inthe area is very high (Allen 1939), and it is impossible to givenew names to newly detected taxa before comparison with type

material, or extensive collections from the type localities.

Chromosomal diversity of Arvicanthis from EastAfrica

Data from the available literature and present investigation

provided a total of 116 specimens from 27 localities (Table 2,Fig. 1). Excluding intra-population polymorphism, a total of10 different cytotypes are represented. The comparisons of G-

banded karyotypes for seven cytotypes is shown in Fig. 2, anda selection of the C-banded karyotype and sex chromosomes isreported in Figs 3 and 4.

Arvicanthis niloticus (Desmarest, 1822)

2n ¼ 62 and NFa ¼ 60–62. This is the most widespread

karyotype as it has been found in Uganda, Kenya andEthiopia in 10 of the 27 localities reviewed (Fig. 1, Table 2).

The autosomal set is composed of all-telocentric chromosomeswith the exception of a small pair of metacentrics (no. 25)

(Fig. 2). This karyotype corresponds to the ones described forboth A. niloticus (previously ANI-1; Volobouev et al. 1987)and A. dembeensis (Corti et al. 1996). The karyotype shows a

consistent block of heterochromatin at the centromeres of allautosomes (Volobouev et al. 2002b).The X chromosome has a short arm that is entirely

heterochromatic, and the Y chromosome is entirely hetero-chromatic (Fig. 4). The sex chromosomes show variation inmorphology: a submetacentric X chromosome has been found

in several localities of Ethiopia and Kenya while Bulatovaet al. (2002) found a subtelocentric configuration in a localityof the Omo Valley. Variation in the autosomal set was alsofound by Bulatova et al. (2002) in the Gambela area (Ethi-

opia), where the karyotype lacks the small metacentric pairwhich characterizes all the species of the genus (2n ¼ 62;NFa ¼ 60) (Table 2).

Arvicanthis cf. somalicus Thomas, 1903

2n ¼ 62 and NFa ¼ 62–63. Baskevich and Lavrenchenko(2000) found a karyotype very close to the one described forA. niloticus in specimens from the Awash National Park,

Ethiopia, they identified as A. somalicus. They have also shownthe presence of a small addition of heterochromatin in twopairs of autosomes. Notwithstanding the similarity of thiskaryotype with that of A. niloticus, an incorrect assignment of

this specimens is unlikely because A. somalicus is a smallspecies and cannot be easily mistaken for larger A. niloticus inthe area. Moreover, the spermatozoon of these specimens is

the most distinctive among Arvicanthis for the presence of avery short sperm head (Table 3), and this was considered

Fig. 1. Map showing the locationsof the samples for which chromo-somal data are available in EastAfrica. Species names or acronymsare indicated. For locality num-bers, see Table 1

Chromosomal diversity in Arvicanthis 225

� 2006 The Authors JZS 44(3), 223–235Journal compilation � 2006 Blackwell Verlag, Berlin

sufficient to be considered as a different species from the othersurrounding 2n ¼ 62 ones (Baskevich and Lavrenchenko1995).Arvicanthis. cf. somalicus has been synonymized with

A. neumanni by Musser and Carleton (2006). However, wefollow, with caution, the interpretation of Baskevich andLavrenchenko (2000) because the two differ in karyotype and

in the shape of the skull (see below). No G-banding ormolecular work has been performed on these specimens for adefinitive identification.

A. abyssinicus (Ruppel, 1842)

2n ¼ 62, NFa ¼ 64. The karyotype of this species is knownfrom three localities of Ethiopia (Orlov et al. 1992; Corti et al.1996). The karyotype differs from that of A. niloticus due to apericentric inversion in chromosome pair number 4 (Corti

et al. 1996; Fig. 2). C-banding shows conspicuous blocks ofheterochromatin at the centromeres of all the chromosomes, atthe short arm of the X chromosome and in all the Y

chromosome (shown in Corti et al. 1996). A different Xchromosome has been found in heterozygous condition

showing a smaller short arm (Corti et al. 1996, Fig. 4). Thespecies is endemic to the highlands of Ethiopia where it occursat altitudes above 1600 m (Yalden et al. 1976).

A. blicki Frick, 1914

2n ¼ 48, NFa ¼ 64. The karyotype of this species is known

from two specimens only (Corti et al. 1996; Lavrenchenkoet al. 1997). The karyotype is characterized by a reduction indiploid number following seven Robertsonian (Rb) fusions.

Given that these fusions are described with a differentnumeration (Corti et al. 1996; Volobouev et al. 2002b), wepresent here the G-banded chromosomes where the numer-

ation follows the standard of A. niloticus, following Volobouevet al. (2002b): Rb(9.15), Rb(10.19), Rb(11.16), Rb(12.17),Rb(13.22), Rb(21.26) and Rb(24.27) (Fig. 2). The C-bandingshows faint spots of heterochromatin at the centromeres of all

autosomes and heterochromatin is also present on the shortarm of X chromosome (Corti et al. 1996).

This species is endemic to Ethiopia and confined to the

moorlands of the Eastern plateau (Bale mts) above 3000 m(Yalden et al. 1976).

Table 2. Species, geographical origin of the samples (site number as indicated in Fig. 1, locality, country, latitude and longitude), number of maleand females studied, diploid number, autosomal fundamental number (NFa), morphology of the X and Y chromosomes (SM ¼ submetacentric;M ¼ metacentric; A ¼ acrocentric; ST ¼ subtelocentric; ? ¼ unknown) and the reference of the original description

Species Site Locality, country Latitude–longitude ## $$ 2n NFa X Y Reference

A. niloticus 1 Kyambara, Uganda Equator - 29�59¢E 1 – 62 62 SM ? Ducroz et al. (1998)2 Masai Mara, Kenya 01�01¢S–35�19¢E 1 1 62 62 SM ? Ducroz et al. (1998)3 Gambella, Ethiopia 07�53¢N–34�22¢E 53 62 62 SM M Orlov et al. (1992)4 Awash Nat. Park,

Ethiopia09�28¢N–40�18¢E 23 62 62 SM – Orlov et al. (1992)

5 Gambela area, Ethiopia 07�55¢N–34�19¢E – 1 62 60 SM – Bulatova et al. (2002)5 Gambela area, Ethiopia 07�55¢N–34�19¢E 123 62 62 SM M Bulatova et al. (2002)6 Omo Valley, Ethiopia 05�00¢N–36�07¢E ? ? 62 62 ST ? Bulatova et al. (2002)7 Zway, Ethiopia 07�55¢N–38�43¢E 5 – 62 62 SM M Present study8 Kitale, Kenya 01�01¢N–35�00¢E 1 – 62 62 SM M Present study9 Koka, Ethiopia 08�24¢N–39�01¢E 2 1 62 62 SM M Corti et al. (1996)10 Koka, Ethiopia 08�13¢N–38�55¢E 203 62 62 SM M Baskevich and

Lavrenchenko (2000)A. somalicus1 11 Awash Nat. Park,

Ethiopia09�00¢N–40�10¢E 53 62 62–63 SM M Baskevich and

Lavrenchenko (2000)A. blicki 15 Sanetti Plateau,

Bale, Ethiopia06�52¢N–39�52¢E – 2 48 62 SM – Corti et al. (1996),

Lavrenchenko et al. (1997)A. abyssinicus 16 Sululta, Ethiopia 09�15¢N–38�43¢E 1 4 62 64 SM-ST M Corti et al. (1996)

17 Managhesha, Ethiopia 09�00¢N–38�35¢E 5 – 62 64 SM M Corti et al. (1996)18 Ambo, Ethiopia 08�56¢N–38�58¢E 113 62 64 Orlov et al. (1992)

A. neumanni 19 Matongolo, Tanzania 05�46¢S–36�28¢E 7 4 53–54 62 SM M Castiglia et al. (2003)20 Zoissa, Tanzania 05�40¢S–36�25¢E 1 2 53–54 62 SM M Castiglia et al. (2003)21 Singida, Tanzania 04�49¢S–34�45¢E 1 1 54 62 SM M Castiglia et al. (2003)22 Ndaleta, Tanzania 05�14¢S–36�29¢E 1 – 53 62 SM M Castiglia et al. (2003)23 Itigi, Tanzania 05�41¢S–34�28¢E 2 – 53–54 62 SM M Castiglia et al. (2003)24 Berega, Tanzania 06�14¢S–37�10¢E 1 – 54 62 SM M Ducroz et al. (1998)

A. nairobae 25 Luwami, Tanzania 03�41¢S–37�32¢E – 2 62 78 ST – Castiglia et al. (2003)26 Manolo, Tanzania 04�37S–38�13¢E 3 2 62 78 ST M Castiglia et al. (2003)27 Nairobi, Kenya 01�16¢S–36�49¢E – 1 62 78 ST – Present study

ANI-5 13 Rongai, Kenya 00�10¢S–35�51¢E 3 2 56 62 SM M Present studyANI-6 7 Zway, Ethiopia 07�55¢N–38�43¢E – 1 60 72 ST – Present studyANI-6a 12 Gamo Gofa,

Konso, Ethiopia05�25¢N–37�20¢E 123 60 76 Orlov et al. (1992)

ANI-7 5 Gambela area,Ethiopia

07�55¢N–34�19¢E 1 – 56 78 A A Bulatova et al. (2002)

ANI-8 14 Afgoi, Somalia 02�08¢N–45�07¢E – 1 44 722 ? ? Capanna and Civitelli (1988)

1These specimens were assigned to A. somalicus by Baskevich and Lavrenchenko (2000) but synonymized with A. neumanni by Musser andCarleton (2006).2Fundamental number (FN).3Sex not indicated.

226 Castiglia, Bekele, Makundi, Oguge and Corti

� 2006 The Authors JZS 44(3), 223–235Journal compilation � 2006 Blackwell Verlag, Berlin

Fig. 2. Comparison of autosomal G-banding patterns among different taxa of Arvicanthis from East and West Africa. a ¼ A. rufinus;b ¼ A. niloticus; c ¼ A. nairobae; d ¼ A. neumanni; e ¼ ANI-5; f ¼ A. blicki; g ¼ A. abyssinicus; h ¼ ANI-6. Lines include homologous por-tions. Arrows indicate the location of the centromere in the biarmed chromosomes. Only five chromosomes of ANI-6 are included in thecomparison, i.e. chromosomes 1, 3, 4, 5 and the Rb(2.6) showing homology in one arm with chromosome 2 and chromosome 6 for the other arm.Numeration follows the one proposed for A. niloticus by Volobouev et al. (2002a)

Chromosomal diversity in Arvicanthis 227

� 2006 The Authors JZS 44(3), 223–235Journal compilation � 2006 Blackwell Verlag, Berlin

Table

3.Summary

ofthetaxaidentified

bycytogeneticswiththedescriptionofmorphologicalcharacters(m

orphologyofthesperm

head,length

oftheupper

molarrow)andgeneticaldistinctivenessfrom

theirsister

taxa(percentageofdivergence

inCyt-bsequence)

Taxon

2n

nFA

Distribution

Minim

um-m

axim

um

upper

molarrow

length

(mm)

Sperm

head2

Genetic

differences

from

sister

taxa(%

)

A.abyssinicus

62

64

Ethiopia

6.2–7.4

1L

9.4–10.5;W

3.0–3.7;H

4.2–5.5

–A.cf.somalicus

62

62–63

Ethiopia,Kenya

5–5.9

1L

5.6–6.9;W3.5–4.2;H1.7–2.3

–A.blicki

48

64

Ethiopia

7.8–8.9

1–

–ANI-6

60

72–76

Ethiopia

(twolocalities)

–L

9.5–10.6;W

3.5–4.0;H

3.8–4.8

6–83

ANI-7

56

78

Ethiopia

(onelocality)

–L

9.6–10.2;W

2.9–3.9;H

2.2–3.6

–ANI-8

44

72

Somalia,(onelocality)

––

–A.nairobae

62

78

Ethiopia,Kenya,Tanzania

6.0–7.1

5(6.6)6

–9–104

A.niloticus

62

60–62

East

andWestAfrica

6.2–7.2

1(7.2)6

L9.4–11;W

3.0–4.0;H

4.0–5.1

8–114

A.neumanni

53–54

62

Tanzania

5.6–6.5

5–

10–124

ANI-5

56

62

Kenya(onelocality)

(6.6–6.9)6

–23

1Yalden

etal.(1976).

2BaskevichandLavrenchenko(1995):L¼

totallength;W

¼width;H

¼length

ofthehook(lm).

3Corti(unpublished

data).

4Ducrozet

al.(1998).

5Verheyen

andHulselmans,

personalcommunication:A.nairobae,

70specim

ensfrom

Machakos(01.31S-37.16E,Kenya);A.neumanni,37specim

ensfrom

Kujungu(05.25S-37.10E,Tanganica)and

Dodoma(06.11S-35.45E,Tanzania).

6Specim

ensanalysedcytogeneticallyin

thepresentstudy.

(a)

(b)

(c)

Fig. 3. C banded karyotypes of Arvicanthis: a, A. nairobae; b, ANI-5;c, A. neumanni

228 Castiglia, Bekele, Makundi, Oguge and Corti

� 2006 The Authors JZS 44(3), 223–235Journal compilation � 2006 Blackwell Verlag, Berlin

A. nairobae J. A. Allen 1909

2n ¼ 62, NFa ¼ 78. The karyotype of this species has beendescribed from individuals caught in two localities in Tanzania(Castiglia et al. 2003). The karyotype for the type locality of

the species (Nairobi, Kenya) is reported here. The autosomalset is composed of nine pairs of biarmed autosomes and 21pairs of telocentric chromosomes which decrease in size

(Fig. 2). The C-banding pattern (Fig. 3) shows an absence ofheterochromatin in the two largest pairs of biarmed chromo-somes (nos 1 and 3) and in five pairs of large acrocentrics. All

the other chromosomes have centromeric heterochromatin. Inthree pairs of medium-sized submetacentrics (nos 16, 17, 22),the heterochromatic blocks entirely involve the short arms,

while in the other pair (no. 18), heterochromatin is apparentlylimited to the centromeric area. The X chromosome is a largesubtelocentric and the Y chromosome is a small subtelocentric(Fig. 4). The short arm and the proximal portion of the long

arm of the X chromosome are entirely heterochromatic. The Ychromosome is entirely heterochromatic.

Comparison of the G banding pattern led to the recognition

of rearrangements relative to the karyotype of A. niloticus, i.e.pericentric inversions in pairs 1, 3, 18, 20 and 29; heterochro-matin additions on pairs 16 and 17 and 22 (Fig. 2) and a

reciprocal translocation between chromosomes 7 and 4.The southern and northern limits of A. nairobae are still

unclear (Musser and Carleton, 2006). We found one limit inthe South of Tanzania (08�15¢S–34�E) at the border between

the Somali-Maasai and Zambezian domains. Bulatova et al.(2002) reported it around the lower Omo Valley in Ethiopia,but with a 2n ¼ 62, NFa ¼ 62 that does not correspond to

what has been found by us for the species. However, theauthors consider the specimens analysed as A. nairobae onlyon the basis of the geographical position. And no morpholo-

gical or other genetical analysis support it belongs to A. nai-robae. Therefore, here we discuss their data in a A. niloticusframework (Table 2).

A. neumanni (Maschie, 1894)

2n ¼ 53–54, NFa ¼ 62. The karyotype of this species shows a

widespread Rb polymorphism (Castiglia et al. 2003). The

chromosomal set of the 2n ¼ 54 specimens is composed of fivepairs of biarmed chromosomes and 21 pairs of telocentricchromosomes. The reduction in diploid number of the 2n ¼ 53

specimens is due to additional Robertsonian fusions betweentwo telocentric chromosomes of the 2n ¼ 54 karyotype.C-positive heterochromatic blocks occur at a paracentromeric

position on all the autosomes, either biarmed or uniarmed(Fig. 3). The X chromosome is a large submetacentric and theY is a short metacentric, which however, is larger than the

smallest metacentric autosomal pair. The short arm of the Xchromosome is entirely heterochromatic together with theproximal portion of the long arm. The Y chromosome is

entirely heterochromatic (Fig. 4).The arms forming the large metacentrics of A. neumanni

correspond to 12 pairs of telocentrics of the A. niloticuskaryotype after Robertsonian fusion: Rb(3.15), Rb(4.21),

Rb(5.12), Rb(6.14), Rb(8.10) and Rb(11.20) (Fig. 2). Thepolymorphic Rb chromosomes are Rb(3.15) (found in four ofthe five localities examined) and Rb(11.20) found in a single

specimen. The range of this species should be confined toTanzania but its limits are still uncertain (see after).

ANI-5 (present study)

2n ¼ 56, NFa ¼ 62. This karyotype, described for the firsttime by standard coloration only in Corti et al. (2005) is

found in the locality of Rongai, Kenya. Here we report theG and C banded karyotype (Figs 1 and 2; Table 1). Thekaryotype is composed of 23 pairs of telocentric and four

pairs of meta-submetacentric chromosomes, one of which isparticularly small. C-bands show the presence of a largeamount of heterochromatin in the paracentromeric areas of

all the autosomes (Fig. 3). The X chromosome is a largesubmetacentric and the Y is a medium-sized metacentric.The short arm of the X chromosomes is entirely

heterochromatic, even if its telomeric portion is lesspronounced. The Y chromosome is entirely heterochromatic(Fig. 4).The different chromosomal arms can be correctly identi-

fied by G-banding (Fig. 2) following Volobouev et al.(2002b). The three largest pairs of metacentrics are due to

Fig. 4. Selected sex chromosomes(G bands on the left and C bandson the right) from different taxa ofArvicanthis

Chromosomal diversity in Arvicanthis 229

� 2006 The Authors JZS 44(3), 223–235Journal compilation � 2006 Blackwell Verlag, Berlin

Rb fusion of telocentric chromosomes identified in A. nil-oticus. These are as follows: Rb(3.10), Rb(11.15) andRb(17.23).

ANI-6 (present study), ANI-6a (Orlov et al. 1992)

2n ¼ 60 NFa ¼ 72/76. The ANI-6 karyotype (2n ¼ 60;NFa ¼ 72) has been described for the first time in Corti et al.(2005), and has been found in only one female from Zway,Ethiopia (Fig. 1, Table 1). It is composed of one pair of large

metacentrics, three pairs of medium/large submeta-metacen-trics and of three pairs of small metacentrics. The X chromo-some is a large subtelocentric with a heterochromatic short

arm (Fig. 4). C-banding revealed little heterochromatin in theautosomes as only three pairs of small metacentrics hadcentromeric heterochromatic blocks. No heterochromatin

occurred in the three pairs of large biarmed chromosomes(not shown). The G-banding pattern of this karyotype werenot as clear as those of other karyograms presented here. In

view of this, and the possibility that this specimen may be theonly representative of a so far undescribed species, only thebanded chromosomes labelled as �h� in Fig. 2 are presented.Five chromosomal rearrangements relative to A. niloticus can

be recognized. These are: Rb(2.6), an inversion in pairs 1 and 3and a reciprocal translocation between pairs 7 and 4. Anotherfour pairs of biarmed chromosomes occurred in this karyotype

indicating the presence of other unidentified rearrangementscompared with A. niloticus. A similar karyotype (named hereas ANI-6a; 2n ¼ 60 NFa ¼ 76) was found by Orlov et al.

(1992) who described it by standard staining. Differencesbetween ANI-6 and ANI-6a probably consist in an addition ofheterochromatin or inversion in two autosomal pairs (Orlov

et al. 1992).

ANI-7 (Bulatova et al. 2002)

2n ¼ 56, NFa ¼ 78. This karyotype has been reported byBulatova et al. (2002) in the Gambela area (Ethiopia). It ischaracterized by two pairs of large metacentrics and 10 pairs of

sub- and small metacentrics. The remaining pairs of chromo-somes are telocentrics. Some of these biarmed chromosomesshow heterochromatic addition to the small arms. The X and

Y chromosomes are telocentrics. Due to the lack of G-banding, it was impossible to ascertain chromosomal homol-ogies between this karyotype and the others known for thegenus.

Bulatova et al. (2002) did not attribute these samples to anyknown taxon.

ANI-8 (Capanna and Civitelli 1988)

2n ¼ 44, Fn ¼ 72. This karyotype was described on the basis

of a single female from Afgoi, Somalia (Capanna and Civitelli1988) (Fig. 1, Table 1). This specimen on the basis of linearmeasurement could be recognized as A. niloticus (see later).

However, it is characterized by a strong reduction in diploidnumber that allows the allocation to a different taxon. The Xchromosome could not be positively identified. Should the Xchromosome be biarmed the NFa would be 68. This karyotype

suggests the occurrence of at least eight Rb fusions, and fourrearrangements which can be inversions or heterochromatinadditions/deletions.

Phylogenetic relationships

Chromosomal rearrangements have been used as characters tohypothesize phylogenetic relationships among cytotypes. Inthis analysis we added the ANI-5 and ANI-6 and A. nairobae

karyotypes to the seven used by Volobouev et al. (2002a) toinfer phylogenetic relationships. As an outgroup, the karyo-type of Lemniscomys zebra was used as it shows a complete

banding homology with the chromosomes of Arvicanthis(Castiglia et al. 2003). Therefore, it is suitable to identify thedirection of chromosomal mutations. We identified 36 chro-mosomal rearrangements (Fig. 2) that were coded in this

analysis following Dobigny et al. (2004). The matrix ofchromosomal characters (Table 4) was analysed by maximumparsimony using the heuristic search option in PAUP 4.0

(Swofford 1998).Ten informative characters were found. The most parsimo-

nious tree found is shown in Fig. 5 (tree length ¼ 39;

consistency index ¼ 0.92, and excluding uninformative char-acters ¼ 0.77). Two major clades occur, corresponding toclades 1 and 2 as identified by Ducroz et al. (1998) throughmolecular phylogenetics.

Clade 1 includes A. ansorgei, A. rufinus, A. nairobae, ANI-2and ANI-6; A. nairobae clusters with A. ansorgei, while ANI-2with A. rufinus. ANI-6 has a basal position relative to

A. ansorgei and A. nairobae. Clade 2 comprises A. niloticus,A. neumanni, A. abyssinicus, A. blicki, and ANI-5. In thisclade only A. blicki and A. abyssinicus cluster together, the

other cytotypes having a bush-like unresolved relationshipaccording to chromosomal characters.

Three homoplasies occur in clade 1 and none in clade 2

(Fig. 5): the first is due to Rb(2.6), occurring independently inANI-2 and ANI-6; the second is the independent occurrence ofa small heterochromatic arm in chromosome 22 of A. nairobaeand the common ancestor of A. rufinus and ANI-2; the third is

the independent occurrence of inversion on small autosomenumber 29 in A. nairobae and A. rufinus.

Given the generally recognized polymorphism of hetero-

chromatic regions, especially short arms, and the repetitivenature of heterochromatin, such a character is likely to be asource of homoplasy. Moreover, we cannot ignore the

possibility of independent Rb fusion and pericentric inversion,in such a highly chromosomally diversified genus.

The chromosomal phylogenetic hypothesis illustrated inFig. 5 shows how the two clades are characterized by different

types of chromosomal rearrangements. In clade 2, Rb fusionsappear repeatedly in different lines and predominate over anyother kind of chromosomal rearrangement. There are no

shared Rb fusions among lines of this clade, and several casesof monobrachial homology occur (Table 4). This stronglysuggests a cytotype radiation from an �all telocentric� condi-tion. On the other hand, inversions, additions of heterochro-matic small arms and Rb fusions characterize chromosomalevolution within clade 1.

The differences in the type of chromosomal mutationbetween the two clades could depend on the differentmolecular architecture of centromeric heterochromatin in-volved in various types of chromosomal mutations (Garagna

et al. 1995; Redi et al. 2001). All the karyotypes of clade 2,with the exception of A. blicki, show conspicuous blocks ofheterochromatin at the centromere (Corti et al. 1996; this

study), but this is more variable and does not constitute acommon pattern for those of clade 1. It is impossible, at

230 Castiglia, Bekele, Makundi, Oguge and Corti

� 2006 The Authors JZS 44(3), 223–235Journal compilation � 2006 Blackwell Verlag, Berlin

present, to determine whether this reflects differences at themolecular level.Redi et al. (2001) reviewed the role of heterochromatin as a

molecular drive for structural rearrangements of the karyotypesuch as during the Robertsonian formation of metacentrics(Garagna et al. 1995, 2001). Heterochromatin transfer among

homologous and non-homologous chromosomes with repetit-ive DNA intragenomic movements suggests that repetitiveDNA families may have been involved at least in Robertsonianprocesses (see Garagna et al. 1995 and Hamilton et al. 1990

for rodents; Wichman et al. 1991 for equids) and might berelated to speciation.Intraspecific sex chromosome polymorphism due to hetero-

chromatic addition or deletion has been found in many speciesof the genus: A. niloticus (Bulatova et al. 2002; Volobouevet al. 2002b), A. abyssinicus (Corti et al. 1996) and A. rufinus

(Civitelli et al. 1995); thus, this variation in sex chromosomescannot be easily used as a taxonomic marker. Nonetheless, wecannot completely exclude a functional consequence of these

chromosomal rearrangements. As a matter of fact, they mayhave some effect in the pairing between X and Y chromosomes(Stitou et al. 2000). However, other functional roles may bepossible. For example, in populations of A. rufinus from

Benin, the occurrence of different Xs significantly affects thegenome size in the specimens, thus may affect the metabolicproperties at the cellular level (Garagna et al. 1999).

Implications of chromosomal variation for thetaxonomy of East African Arvicanthis

Five of the identified cytotypes correspond to the speciesreported by Musser and Carleton (2006), i.e. A. nairobae,

A. neumanni, A. blicki, A. abyssinicus and A. niloticus. Allthese cytotypes are distinguishable by their own chromosomal(Corti et al. 1996; Castiglia et al. 2003), genetic (Capula et al.1997; Ducroz et al. 1998) and morphological (Yalden et al.

1976, 1996; Baskevich and Lavrenchenko 1995; Corti andFadda 1996; Fadda and Corti 1998) differences (Table 3). Thetaxonomic status of A. cf. somalicus and of the cytotypes

named ANI-5, ANI-6/6a, ANI-7 and ANI-8 merits a morethorough discussion.Musser and Carleton (2006) synonymized A. somalicus with

A. neumanni from Tanzania. We prefer here to retain the taxonname somalicus because it presents some important differences.First, the two taxa show a different karyotype, where A. neu-manni has four to six additional Rb fusions in Tanzania.

Putative F1 hybrids between these two taxa might carry fromfour to six trivalents at meiotic diakinesis. This meioticconfiguration usually implies high levels of ipofertility in

house mouse hybrids (Garagna et al. 1990; Wallace et al.2002). Moreover, Fadda and Corti (2001) found a differentskull shape in the Tanzanian sample compared with the

Kenyan, Somalian and Ethiopian samples. Only molecularanalysis or breeding data may help to understand if A. soma-licus from Ethiopia is a chromosomal form of A. neumanni or

that it represents a distinct species. If the Tanzanian popula-tion is a different species, A. neumanni would be the correctname for it, and A. somalicus would extend its range fromKenya to Somalia, south Ethiopia and south-east Sudan

(Musser and Carleton, 2006).The cytotype ANI-6 from Ethiopia (2n ¼ 60 NFa ¼ 72,

Fig. 1, Table 1) is very similar to the one described by Orlov

et al. (1992) (standard techniques only; ANI-6a, 2n ¼ 60,Table

4.Matrix

ofchromosomalcharacters(1–36)identified

inArvicanthisandusedforthephylogenetic

reconstruction.ThekaryotypeofLem

niscomyszebra

hasbeenusedasoutgroup

12

34

56

78

910

11

12

13

14

15

16

17

18

19

20

21

22

23

24

25

26

27

28

29

30

31

32

33

34

35

36

Ref.

A.niloticus(A

NI-1)

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

eA.niloticus(A

NI-1b)

10

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

eANI-2

01

11

11

11

10

00

00

00

00

00

00

00

00

00

00

00

00

00

eA.ansorgei

00

01

11

01

11

11

00

00

00

00

00

00

00

00

00

00

00

00

eA.rufinus

00

01

11

11

10

00

11

00

00

00

00

00

00

00

00

00

00

00

e,c

ANI-6

00

11

1?

0?

?1

00

00

00

00

00

00

00

00

00

00

00

00

00

fA.neumanni

00

00

00

00

00

00

00

11

11

11

00

00

00

00

00

00

00

00

bA.abyssinicus

00

00

00

00

00

00

00

00

00

00

10

00

00

00

00

00

00

00

dA.nairobae

00

01

11

10

11

01

00

00

00

00

01

10

00

00

00

00

00

00

bA.blicki

00

00

00

00

00

00

00

00

00

00

10

01

11

10

00

00

00

11

dANI-5

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

01

11

00

fLem

niscomyszebra

00

00

00

00

00

00

00

00

00

00

00

00

00

01

11

10

00

00

a

Thechromosomaltransform

ationsare

coded

asfollows:1,inv(30);2,Rb[1;trcp(4;7)];3,Rb(2;6);4,inv(3);5,trcp(4.7);6,inv(20);7,hadd/del(22);8,inv(28);9,inv(29);10,inv(1);11,in(2);12hadd/del

(17);13hadd/del(1);

14hadd/del(2);15,Rb(3;15);16,Rb(4;21);17,Rb(5;12);18,Rb(6;14);19,Rb(8;10);20,Rb(11;20);21,inv(4);22,hadd/del(16);23,inv(18);24,Rb(9;15);25,Rb(10.19);26,

Rb(11.16);27,Rb(12;17);28,tanfus/fis(1.20);29,tanfus/fis(6.24);30tanfus/fis(7.11);31,Rb(30.29);32,Rb(3.10);33,Rb(11.15);34,Rb(17.23);35,Rb(13;22).36,Rb(24;27).Inv¼

pericentricinversion;

Rb¼

Robertsonianfusion;trcp¼

reciprocaltranslocation;hadd/del

¼heterochromaticaddition/deletion;tanfus/fis¼

tandem

fusion-fission.References:

a,Castigliaet

al.2002;b,Castigliaet

al.

2003;c,

Civitelliet

al.1995;d,Cortiet

al.1996;e,

Volobouev

etal.2002a;f,presentstudy.

Chromosomal diversity in Arvicanthis 231

� 2006 The Authors JZS 44(3), 223–235Journal compilation � 2006 Blackwell Verlag, Berlin

NFa ¼ 76) in the Gambela region, which is approximately300 km from Zway. The authors, when considering theexternal morphology, decided these specimens are similar toA. niloticus, although they differ slightly in the morphology of

the spermatozoa (Baskevich and Lavrenchenko 1995) (Ta-ble 3). The G- banding pattern here presented clearly placesANI-6 within clade 1 and not in clade 2 as A. niloticus, as there

is the same reciprocal translocation between chromosomes 4and 7 characterizing this clade (see introduction). For thisreason here we confirmed the high chromosomal divergence

between ANI-6 and A. niloticus. Moreover, the analysis of thesequences of the entire cytochrome b of these specimens showsthat the closer representative of this clade is A. ansorgei (from

Senegal and Burkina Faso), which diverged by 6–8% (Corti,unpublished data), a moderately high level of divergence in asequence of cytochrome b. Recognized differences with thekaryotype of A. ansorgei include one Rb fusion and one

inversion. However, ANI-6 shows another six unidentifiedpairs of biarmed chromosomes. Due to this uncertainness inthe interpretation of karyotypes, it is difficult to evaluate the

effect of the observed chromosomal differences on fertility ofputative hybrids. Therefore, additional data are required totaxonomically evaluate this taxon correctly.

Orlov et al. (1992) recognized the large metacentric of ANI-6a as Rb(1.7), but from their picture no categorical identifi-cation of telocentrics involved in the fusion is possible.Therefore, it cannot be excluded that the chromosome

recognized as Rb(1.7) is, in fact, Rb(2.6). Were this to be thecase, then differences between ANI-6 and ANI-6a would beminimal and probably represented by an inversion or addition

of heterochromatic material, showing evidence of chromoso-mal variation within this undescribed taxon. This same Rb(2.6)chromosome is also shared by ANI-2 from the Central African

Republic (Table 2). However, in our phylogenetic tree (Fig. 5)it has been hypothesized that its occurrence in these twocytotypes represents an independent event, although a less

parsimonious solution could also depict the �true� phylogeny.

No G-banding, morphological and molecular data areavailable for the cytotypes named ANI-7 and this makes iteven more difficult to establish its taxonomic status. Thepresence of large metacentrics in ANI-7 could relate this

cytotype to ANI-6, but the sperm head of these specimens israther distinctive with a short hook and this may indicatespecific distinction (Baskevich and Lavrenchenko 1995, Ta-

ble 3). Another possible relative cytotype is ANI-2 from theCentral African Republic, also characterized by large meta-centrics. At the moment the relationship between this cytotype

and the eastern and central relatives is unclear. However, itseems reasonable to suppose that ANI-7 represents an addi-tional species for the East African Arvicanthis fauna.

ANI-5 from Rongai differs from all other known karyo-types. Chromosomes 4 and 7 are present in the ancestral state(i.e. they are not involved in reciprocal translocation) and thismakes it possible to place this species within the group of the

strictly East African species, together with A. niloticus,A. abyssinicus, A. blicki and A. neumanni (clade 2, followingDucroz et al. 1998). Its diploid number and NFa resemble the

karyotype of A. neumanni (2n ¼ 53–54; NFa ¼ 62; Castigliaet al. 2003). However, the G-banding revealed several differ-ences. As a matter of fact, there are no metacentrics shared

with other cytotypes and, among these, Rb(3.15) of A. neu-manni shows a monobrachial homology with metacentricsRb(3.10) and Rb(11.15) of ANI-5. The metacentrics of ANI-5are likely to have arisen directly from an ancestral �all-telocentrics� karyotype as for A. niloticus (see later). The lengthof the upper molar row, a good indicator of dimension in thesespecimens (6.6–6.9) falls within the range known for A. niloti-

cus (6.2–7.2) (Table 3).The analysis of the sequences of the entire cytochrome b of

these specimens showed a 2.5% divergence from the 2n ¼ 62

NFA ¼ 62 A. niloticus from Kitale (Corti, unpublished data).This value falls within the range of intraspecific variation. Thelow distance between ANI-5 and A. niloticus is considerable,

and suggest a fast chromosomal evolution in these species.

Fig. 5. Most parsimonious treebased on chromosomal rearrange-ments in Arvicanthis. The tree hasbeen outgroup rooted on Lem-niscomys zebra. Character codingand rearrangement numbering isrepresented in Table 2. The differ-ent kinds of chromosomal rear-rangements are represented in fullsymbols, homoplasies in opensymbols

232 Castiglia, Bekele, Makundi, Oguge and Corti

� 2006 The Authors JZS 44(3), 223–235Journal compilation � 2006 Blackwell Verlag, Berlin

In putative hybrids with A. niloticus the three Rb fusions ina heterozygous condition would not severely impair gameto-genesis and theoretically would not represent a strong post-

zygotic isolation barrier, as occurs in other rodent species (seefor example, Castiglia and Capanna 2000). However, onlythrough the analysis of structural heterozygotes from an

eventual hybrid zone between ANI-5 and A. niloticus (thedistance between Rongai and Kitale is about 160 km) mayassess the taxonomic status of this cytotype that easilyrepresent a chromosomal race within A. niloticus.

An extreme reduction in diploid number was found byCapanna and Civitelli (1988) in one female from Somalia(ANI-8; 2n ¼ 44). In Somalia two morphospecies of Arvican-

this occur, a large one identified as A. niloticus and a small oneidentified as A. somalicus. The specimen analysed by Capannaand Civitelli clearly falls in the first group based on the large

size of the body and the skull. Chromosomal differencesrelative to the 2n ¼ 62 A. niloticus include eight fusions andother unidentified rearrangements. Putative hybrids between

these specimens and A. niloticus should have a severe impair-ment at gametogenesis leading to complete sterility in somecases (Gropp et al. 1982; Hauffe and Searle 1998; Castiglia andCapanna 2000). For this reason we follow Capanna and

Civitelli (1988) suggesting that this specimen may represent adifferent unidentified species.

Times of chromosomal evolution

It is believed that the diversification of African mammal

species has been caused by cycles of disruption and coalescenceamong biomes from the Pliocene onwards (Grubb 1999).Chromosomal phylogeny in Arvicanthis, however, suggests an

intricate relationship pattern between geography and phylo-geny, with members of the different clades coexisting in thesame area, even in strict syntopy (for example see the localityof Zway and the Gambela region in Ethiopia; Fig. 1, Table 2).

Moreover, current knowledge regarding the real ranges ofthese chromosomal species is still too scarce so that limitedhypotheses on the biogeographical causes of variation are

possible.The richness of species and karyotypes in this part of Africa

is higher relative to West Africa, with members of both clades

occurring. This reinforces the view that savannas and high-lands surrounding the Rift Valley represent the main, if not theoriginal, centre of diversification of the genus, although, forthe moment, no older fossils than the Omo Valley specimen

(3 Myr; Wesselman 1984) have been found.Based upon complete mitochondrial DNA cytochrome b

sequences (Ducroz et al. 1998), the deepest divergence time

estimated within Arvicanthis is 5.3 Myr (end of the Miocene),during which, therefore, the split between clades 1 and 2occurred. The within-clade diversification is more recent and

the causes of chromosomal diversification and cladogenesis areplausibly related to the geological and climatic events thatcharacterized East Africa during the late Pliocene and early

Pleistocene (Ducroz et al. 1998). As a matter of fact, Arvican-this occurs in all open habitats, from the very arid conditionsof Somalia to the moister Guinean region, and from sea levelto the altitudes of the Bale massif, showing patterns of

adaptation to these very diverse conditions (Fadda and Corti2001).

Molecular phylogenetics support an earlier divergence of the

lines of clade 1, as well as an overall lower rate of chromo-

somal evolution with respect to the lineages forming clade 2,where chromosomal radiation occurred in more recent times(Ducroz et al. 1998). The discovery of chromosomal differ-

ences between A. niloticus and ANI-5 (with three fixed Rbmetacentrics) coupled with low genetic distances between taxaindicates that the process of differentiation in Arvicanthis may

be much more recent.Furthermore, the widespread Rb polymorphism in A. neu-

manni from Tanzania (Castiglia et al. 2003) indicates thatfactors other than geographical isolation may be driving

current chromosomal evolution in the genus. It has beensuggested that a new chromosomal mutation could spread andbecome fixed without any form of geographical isolation; this

can be due to population structure and/or some kind ofsuperiority of the new chromosomal variant (Pardo-Manuel deVillena and Sapienza 2001; Ratkiewicz et al. 2002).

Concluding remarks

Ten different cytotypes have been found in East AfricanArvicanthis, against the five species listed by Musser andCarleton in the area. Concerning the remaining five cytotypes,four (A. cf. somalicus, ANI-6, ANI-7, ANI-8) show a level of

chromosomal, genetic and morphological diversification thatcould suggest a specific status; one of the cytotypes (ANI-5) islikely to be a chromosomal race because of the low divergence

in mtDNA sequences compared with A. niloticus. This diver-sity must still be regarded as an underestimate of the realspecies number as few samples have been studied compared

with the suitable areas for the genus in this part of Africa. Thistaxonomical complexity has, without doubt, an impact onapplied studies such as ecology or parasitology. The syntopy of

cryptic species occurring in two localities (Fig. 1, Table 1)shows the need to investigate undiscovered areas and theecology of different species. Nonetheless, karyotype analysis isessential to determine the entire diversity in the genus

Arvicanthis, given that populations with low molecular diver-gence show different karyotypes. Therefore, this must also beevaluated to establish a correct taxonomy.

Acknowledgements

This paper is dedicated to Walter Verheyen, recently deceased, whodedicated his scientific life to the taxonomy of African rodents. Weexpress our gratitude to those who helped us in this research inEthiopia (Goitom Redda), Tanzania (A.W. Massawe, L. Mulungu, G.Mgode K. Kessy, T. Protas), Kenya (D. Ojwang, B. Agwanda) andItaly (E. Capanna, C. Fadda). Thanks are extended to W. Verheyenand J. Hulselmans for providing original information on the length ofupper molar row of Tanzanian Arvicanthis and to G. Contrafatto andan anonymous referee for helpful comments and suggestions. Thiswork was supported by grants �Programmi d’Ateneo 60%�, �MURST40%�, �STAPLERAT: Protecting staple crops in eastern Africa:integrated approaches for ecologically based field rodent pest man-agement� (European Union) (M.C.).

Riassunto

Diversita cromosomica nel genere Arvicanthis (Rodentia, Muridae)dell’Africa dell’est. Una valutazione tassonomica e sistematica.

In questo lavoro viene discusso il contributo che la citogenetica hafornito alla sistematica del genere Arvicanthis in Africa Orientale.Viene presentata una rassegna di tutti i dati cromosomici fino ad oraconosciuti per il genere nell’area e nuovi dati citogenetici da tre localita

Chromosomal diversity in Arvicanthis 233

� 2006 The Authors JZS 44(3), 223–235Journal compilation � 2006 Blackwell Verlag, Berlin

del Kenya e una localita dell’Etiopia. Inoltre, viene fornito il cariotipobandeggiato G- e C- per due cariotipi descritti recentemente e chiamatiprovvisoriamente ANI-5 (2n ¼ 56, NFa ¼ 62) e ANI-6 (2n ¼ 60,NFa ¼ 72). Il numero di citotipi totali rinvenuti nell’area e pari a 10.Di questi, cinque corrispondono alle specie presenti nell’ultimarevisione, vale a dire A. nairobae (2n ¼ 62, NFa ¼ 78), A. neumanni(2n ¼ 53–53, NFa ¼ 62), A. blicki (2n ¼ 48, NFa ¼ 62), A. abyssin-icus (2n ¼ 62, NFa ¼ 64) and A. niloticus (2n ¼ 62, NFa ¼ 60–62).Viene discusso lo status tassonomico dei rimanenti 5 citotipi (A. cf.somalicus, 2n ¼ 62 NFa ¼ 62–63, ANI-5, 2n ¼ 56, NFa ¼ 62; ANI-6/6a 2n ¼ 60, NFa ¼ 72–76; ANI-7, 2n ¼ 56, NFa ¼ 78 and ANI-8,2n ¼ 44, NF ¼ 72). Inoltre, sono state individuate lievi differenzecromosomiche tra A. niloticus e A. somalicus (2n ¼ 62, NFa ¼ 62–63)(precedentemente messo in sinonimia con A. neumanni). L’alberofilogenetico costruito con tutti i cariotipi del genere in Africa mostra lapresenza di due cladi principali, ciascuno caratterizzato da differentitipi di mutazioni cromosomiche. I tempi degli eventi cladogenetici,stimati tramite orologio molecolare, indicano che l’evoluzione crom-osomica ha accompagnato quasi tutti gli eventi speciativi dalla fine delMiocene ad oggi. La scoperta di un elevato differenziamento cromo-somico tra popolazioni che mostrano una bassa distanza genetica e lapresenza di polimorfismo intrapopolazione indicano che il processo didifferenziamento cromosomico e ancora in atto e che puo essereresponsabile degli eventi di speciazione.

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Author’s address: Riccardo Castiglia (for correspondence), Diparti-mento di Biologia Animale e dell’Uomo, Universita di Roma�La Sapienza�, Via A. Borelli 50, 00161 Rome, Italy. E-mail: castiglia@uniroma1.it

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