Karyological and dental identification of Microtus limnophilus in a large focus of alveolar echinococcosis (Gansu, China)

0 Academic Animal

des sciences

biology

and

/ Elsevier,

pathology

Paris

/ Sio/ogie

et pathologic

animales

Karyological and dental identification of Microfus hnophihs in a large focus of alveolar echinococcosis (Gansu, China) Ident$cation caryologique et dentaire de Microtus limnophilus foyer important de 1Zcbinococcose alvkolaire (Gansu, Chine)

dans un

Fr6deric Couranta*, Patrick Brunet-Lecomtea, Vitaly Volobouevb, Jean Chaline”, Jean-Pierre Q&t+“, Adam Nadachowskid, Sophie Montuire”, Genshu Baoe, Laurent Viriotf, Robert Rauschg, Margarita Erbajevaarh, Dazhong Shi’, Patrick Ciraudoux’ “UMR CNRS 5561 et Laboratoire de palkobiodiversite et prehistoire de I’EPHE, universitb de Bourgogne, Centre des sciences de la Terre, 6, boulevard Gabriel, 21000 Dijon, France bMammifPres et oiseaux, Museum national d’histoire naturelle, 55, rue Buffon, 75005 Paris, France ‘UMR CNRS 5554, Laboratoire d’&co&hologie des rongeurs, universite de Montpellier-II, 34095 Montpellier cedex 5, France dlnstitute of Systematics and Evolution of animals, Polish Academy of Sciences, Slawkowska 17, 31-016 Krakow, Poland ‘Department of Parasitology, Lanzhou Medical College, 85 Dun Gang Xi Road, 730000 Lanzhou, China ‘Laboratoire de ghobiologie, biochronologie et palkontologie humaine, faculte des sciences fondamentales et appliqukes, universite de Poitiers, 40, avenue du Recteur-Pineau, 86022 Poitiers, France gDepartment of Comparative Medicine, University of Washington, Seattle, WA 98195-7190, USA hGeological Institute, Siberian Department, Russian Academy of Sciences, 670042 Ulan-Ude, Russia ‘Who Collaborating Centre, Laboratoire de biologie et &ophysiologie, universitb de Franche-Comte, place Leclerc, 25030 Besancon, France (Received Note

18 November

communicated

1998;

by Jean

accepted

11 February

1999)

Dorst

Abstract A study of voles (Arvicolidae, Rodentia) from Gansu (China) designed to identify a potential host of Echinococcus mu/ti/ocu/aris, responsable for human alveolar echinococcosis, leads to a general analysis of Micro&s limnophilus population karyotypes, M, of M. oeconomus populations from all of Eurasia and of M. limnophilus of Mongolia. The Microtus of Cansu belonging to the nominal subspecies M. limnophilus limnophilus (2n = 38; NF = 58) differs markedly in size and shape of M, from the M. limnophilus of Mongolia, which must therefore be considered as a new subspecies M. limnophilus malygini nov. ssp. (2n = 38; NF = 60) and the M. oeconomus of Mongolia should be ranked as M. oeconomus kharanurensis nov. ssp. (2n = 30; NF = 60). 0 Acadkmie des Sciences / Elsevier, Paris Microbus

/ Arvicolidae

/ taxonomy

I human

alveolar

echinococcosis

/ Gansu

(China)

R&umk - Une 6tude des campagnols (Arvicolidae, Rodentia) du Gansu (Chine) permettant d’identifier un hate potentiel d’Echinococcus multilocularis, l’agent parasitaire de l’tkhinococcose alvkolaire humaine, a impliquk une analyse g&kale des caryotypes des populations de Microtus limnophilus, de la variablilitk des M, des populations de M. oeconomusd’Eurasie et de M. limnophilus de Mongolie. Le Microtus

* Correspondence

and reprints:

C. R. Acad. Sci. Paris, Sciences 1999,322,473-480

[email protected] de la vie

/ Life Sciences

F. Courant

et al.

du Gansu appartenant 2 la sous-espke nominale de M. limnophilus limnophilus (2n = 38; NF = 58) diffke fortement par la taille et la forme des M, des M. limnophilus de Mongolie qui doivent done etre consid&& comme une nouvelle sous-espkce M. limnophilus malygini nov. ssp. (2n = 38; NF = 60) et les Microtus oeconomus de Mongolie doivent Etre class& comme M. oeconomus kbaranurensis nov. ssp.(2n = 30; NF = GO). 0 Acadkmie des Sciences / Elsevier, Paris Microtus

/ Arvicolidae

Version

/ taxinomie

/ txhinococcose

ahkolaire

abr6gE

Dans plusieurs regions du monde, Microtus oeconomus est un des h8tes principaux d’Echinococcus multilocularis, cestode responsable de 1’6chinococcose alveolaire humaine. Depuis 1994, des etudes eco-6pid&miologiques ont et6 entreprises dans le sud du comt6 de Zhang (Gansu, Chine), zone oti la prevalence de cette maladie chez I’homme est de tri’s loin la plus 6levee enregistree au monde 1 ce jour. Les peuplements de rongeurs du Gansu sont extremement diversifi&, particulierement dans le sud du pays et il existe peu d’informations r&entes sur leur systematique. La decouverte de campagnols prksentant des afinites avec Microtus oeconomus et Microtus limnophilus impliquait de resoudre leur position taxinomique 5 partir des analyses dentaire et chromosomique de diverses populations mondiales. Le materiel analysC provient de trois populations de Microtus limnophilus de Chine (Gansu) et de Mongolie et de onze populations de Micro&s oeconomus d’Eurasie (Mongolie, Buriatie, Pays-Bas, Finlande, S&de, France) et d’Am&ique du Nord (ile Saint-Lawrence, Alaska). Les m&hodes utilisees concernent la mise en tvidence des caryotypes et I’analyse morphologique dentaire. Les analyses morphomCtriques de la M, montrent d’une part la dtiirenciation des deux populations de M. limnophilus du Gansu et d’autre part la position marginale de M. limnophilus et de M. oeconomus

1. Introduction Microtus

oeconomus

is one of principal flatworm that

hosts causes

for fchialveolar

biology

The discovery of voles mus and M. limnophilus

474

(Chine)

de Mongolie par rapport aux autres populations. Les analyses dentaires ont r&U l’homog6neit6 morphologique globale des populations de Microtus oeconomus d’Europe et notamment leur proximitk avec les populations de Buriatie, Les specimens de la population de Micro&s collect& dans le Gansu se referent par leur morphologie dentaire 2 M. oeconomus possCdant quelques caracteres de M. limnophilus de Mongolie. On pouvait penser qu’il s’agissait de la sousespece M. oeconomus Jlaviventris, originellement d&rite dans le sud du Gansu qui peut etre, dans 1’Ctat actuel des connaissances, mise en synonymie avec la population type de M. limnophilus de Qaidam (Qinghai, Chine). L’analyse caryologique montre qu’il s’agit bien de l’espgce Micro&s limnophilus. En conclusion, la synth?se de ces analyses montre que les deux populations de Microtus limnophilus du Gansu sont assez 6loign6es de la population de Microtus limnophilus de Mongolie, tant par la taille que par la forme de la M,. En con&quence d’aprPs ces donnkes, on peut conclure 5 une differentiation infraspecfique des Micro&s limnophilus du Gansu par rapport 2 ceux de Mongolie, Nous proposons done de distinguer la forme mongole sous le nom de M. limnophilus malygini nov. ssp. De m?me, le M. oeconomus de Mongolie presente de telles diff6rences avec ceux des autres populations d’Eurasie qu’on peut le distinguer sous le nom M. oeconomus kbaranurensis nou. ssp.

answer dental

in humans [I]. Since 1994, ecostudies about Echinococcus multiloculaconducted in the south of Zhang County where the prevalence of this disease in

humans is by far the highest recorded in the world to date [2]. These studies have entailed a survey of rodents as potential hosts of Echinococcus multilocularis in different biotopes. The rodent populations of Gansu are highly diverse, especially in the south of the region which lies at the crossroads of the Palearctic and Indo-Malayan zones [3,4]. However, there is little recent information about the systematics, cies [4, 51.

/ Gansu

around villages where the disease is most prevalent makes it important to determine the taxonomic status of the Microtus population of Gansu [6]. This study attempts to

nococcus multilocularis, the echinococcosis epidemiological ris have been (Cansu, China)

humaine

or ecology having within

of the

region’s

spe-

affinities with M. oeconothe rodent communities

this question on the analyses of the different

basis of chromosomal populations worldwide.

and

2. Material

M.

The material limnophilus

analysed of China

comes (Gansu)

from three populations and Mongolia and

of from

11 populations of M. oeconomus of Eurasia (Mongolia, Buryatia, the Netherlands, Finland, Sweden, France) and North America (St. Lawrence Island, Alaska). In addition, the lectotype of M. limnophilus [7], the holotype of M. limnophilus flaviventris [8] and two specimens of M. limnophilus of Mongolia were used as supplements to the main analyses. C. R. Acad.

Sci. Paris, Sciences

de la vie / Life Sciences 1999,322,473-480

Microtus

3. Karyological

analysis

Two

were

male

specimens

used

for karyological

sis. Both were caught in 1997, one in Shang 104”26E) and the other in Huang He (34”33N, in the same colonies as the animals caught 1996, respectively. Chromosome preparations ned from fibroblast cell cultures established muscle biopsies. A portion of the cells and

analy-

He (34”27N, 104”16E), in 1994 and were obtaiafter tissue

pectoral explants

of studied specimens were routinely preserved in liquid nitrogen in the cell and tissue collection of the Mammal and Bird Zoology Laboratory, of the National Natural History Museum (MNHN, Paris). The mitotic chromosomes were studied with G (GTG) following the procedures of ner [I 1 I, respectively. At least lysed for each specimen.

4. Morphometric 4.1.

General

The

M,

methods

and C (CBG) Seabright [lo] 20 metaphases

banding [91 and Sumwere ana-

morphology are

those

in alveolar

echinococcosis

and nine pairs of acrocentrics, one largest chromosome pair. The X chromosome and similar in size to the 4th and 5th pairs

defined

by

Brunet-

Lecomte [12, 131 from 23 measurements on the occlusal surface of the first lower molar, M, (figure 1). Principal component analysis was first carried out in order to ascertain differences between individuals. Then, canonical discriminant analysis was performed to establish the maximum differences between populations. These methods take account of the comparison of the parameters of total length of M, (TL), relative length of the anterior part of M,

After

application

of the

C-banding

technique,

all chro-

mosomes including X display moderate blocks of centromerit heterochromatin slightly varying from pair to pair in size and staining intensity. Moreover, two pairs, the 7th and 1 Oth, possess telomere c-bands. The Y chromosome is entirely heterochromatic (figure 3). The

karyotype

of M.

limnophilus

presented

here

corres-

ponds to the first description made on specimens from Mongolia [I41 except for a minor difference in the morphology of chromosome pair 9 involving a small hetero-

variability of the heterochromatic part of animal genomes on the one hand, and the limited number of karyotyped animals in both Mongolian and Chinese populations on the other, it seems too early to confirm that the differences found are fixed and thus take them into consideration for specifying

the taxonomic

status

6. M, morphology 6.1.

Comparison

This

comparison

of M, (figure

of these

forms.

analysis parameters 4) shows

in particular

that:

the total length of M, (TL), the Alaska population the largest of the M. oeconomus populations and limnophilus of Mongolia is also larger than M. limnophilus

tres

of Gansu;

4.2. For 1994)

parameter

AP is dimensionless.

-for

- for the relative the M. oeconomus

Parameters parameter analysis, and 2 (Gansu, July

populations 1996) were

1 (Gansu, May grouped together in

the China set; populations 5 (Selenga River), 6 (Kotokel Lake) and 7 (Kurba River) were grouped together in the Buryatia set; populations 8 (Zeeland), 9 (Texel Island) and 10 (Friesland) were grouped together in the Netherlands set, and populations 11 (Kilpisjarvi) and 12 (Jamtland) were grouped together in the Finland-Sweden set. Statistical analysis was conducted with SAS software and factorial maps displayed by ADES software.

value than of Mongolia;

Both

studied

specimens

results and discussion of M.

limnophilus

lar diploid numbers (2n = 38) and fundamental or numbers of chromosome arms (NF = 58). type contains eight pairs of metacentrics, CR. Acad. Sci. Paris, Sciences 1%9,322,473-480

de la vie / Life Sciences

possess

simi-

length of the anterior part population from Mongolia

populations

of Gansu

of the anterior of Buryatia,

and

M.

loop (AL), the the Netherlands

of Gansu, - for

Buryatia

variable Netherlands and the

Gansu

Principal

and

is M.

of M, (AP), has a lower limnophilus M. oeconoand Finlan-

d-Sweden have a less open anterior loop than those Alaska and of M. limnophilus of Mongolia; - for variable v7, the M. limnophilus population Mongolia has a greater value than those of the populations

6.2.

numbers Their karyoone pair of

the

-for closure mus populations

than

5. Karyological

of

autosomes, whereas the Y chromosome is acrocentric and similar in size to the smallest pair of acrocentric autosomes (figure 2).

(AP), anterior loop closure (AL), to which are added morphological variables v7 and vl5 by one-factor analysis of variance supplemented by the Scheffe contrast test 1131. Parameters TL, AL, v7 and vl5 are expressed in millimewhile

of

chromatic segment. After our analyses, this chromosome looks clearly acrocentric, whereas the latter authors identified it as submetacentric, resulting in different NF 58 versus 60. However, taking into account the extreme

methods

utilized

subtelocentrics which is the is metacentric

implied

of of

Europe;

~15, the populations Finland-Sweden have

of Alaska, a greater

the value

population. component

analysis

This analysis shows that axis 1 bears 48.6% of total variance (figure 5A). This is the result of a size-effect, common in such analyses. The gradient is pulled by the large form of M. oeconomus from St. Lawrence Island.

F. Courant

et al.

6 I 5

ZIN 2335

II

I ’ 2-J 7

I

ZJN 2334

D *... .:. ..:.:. :: :: ..‘a. . :. B JC02

JCOl

::.,:. .: .‘:. .I...... ii! 1299

1255

JD04

808

146

lmm

Figure 1. M, morphology of different morphotypes of the main populations of M. limnophilus and M. oeconomus. A: Morphometry of Microtus M, using 23 measurements; 6: lectotype M. limnophilus Btichner, 1889, n’ZlN 2335, Qaidam (Qinghai, China); C: holotype M. /imnophilus flaviventris Satunin, 1903, n”ZIN 2334, Chertynton (Cansu); D: Morphotypes. M. limnophilus of Gansu 05/94: JCO2, JCOl, JD04 (Shang He); M. limnophilus of Mongolia: 808, 811 [holotype mabygiri nov ssp.], 809 (Dzhungar Gobi), 1299 (Dzabkhan oeconomus of Mongolia: 1255 [holotype (Kharanurensis nov. ssp.)], 1305 (Amar-Khiyda); M. oeconomus of Buryatia: 146 (Kotokel (Kurba River); M. oeconomusof Netherlands: 586B (Zeeland); M. oeconomusof Sweden: 001 (Jamtland, Valadalen); M. oeconomusof (S’. Thibault de Couz); M. oeconomus of Alaska: 7, 41, 10 (St. Lawrence Island, Savoonga). (scale given for B-D). C. R. Acad.

Sci.

Paris,

Sciences

de

Gol); M. Lake), 530 France: 08

la vie / Life Sciences 1999,322,473-480

Microbus

14 APwhcdurity

I .

l-2 3 4 5-6-l 8-9-10 11.12 l3 14

10

implied

I

in alveolar

3.llz

N 33 11

I

30 w 10 10

1%

1 1

N 38

1 1

O.llO

I

Mean OAR3 0.4% OAsl odb7

echinococcosis

SdDev onl3 o.ols onw ON4 ml9 oJn4 0~316 0.m

1 A tklldre’s A A B

mr C

ABC B

C

ABC ABC A

AUmmm)

11

12

16

13

17

14

I-2 3 4 S-6-7 S-9-10

15

18

2. Karyotype of M. limnophilus after Chromosomes are arranged in correspondence presented in Malygin et al., 1990.

G-banding with

the

shown

from

Finland.

here,

lead

lian Microtus from land). As a whole, clearly the two

Axes

to similar

M. the

stdlh WJ3l

arm OJE 0.M o.c65 on67 om6 0.062

(CTG). karyotype

a058 0.062 OBl 0.042

Axes 2 and 3 (figure 5B) eliminate the size-effect and represent 18.4% of variance. They separate at one end the two Micro&s from Mongolia and at the other end M. not

1 1

1 Sdm=ifdsW 1 BCD A B AB C C

D D D

C

D D

BCD A

XY

Figure

oeconomus

h&m OA9?

2 and results,

oeconomus principal

shows a) a morphological Mongolian species, b) the

4 (16.2%

variance)

separating

Mongo-

from Friesland (Holcomponent analysis convergence morphological

between conver-

S-6-7 s-9-10 11-u

13 14

25 30 W 10 10

-zF s0343 0367 om 0347 02u6

A

A A A A A

Figure 4. Ml parameters. TL: total length; AP: anterior part; AL: anterior ~15: variable 15. Analysis of variance: P< 0.001

B B

CD

B

B

loop; v7: variable for all five variables.

7;

For the Scheffb test: * means with same letter are not significantly different (P > 0.05). 1 : M. limnophilus - Gansu 05/94 (China); 2: M. limnophilusGansu 07/96 (China); 3: M. /imnophik~s -Mongolia; 4: M. oeconomus - Mongolia; 5: M. oeconomus - Selenga River (Bu-

2

3

‘-7

4

.**

8

12

5

9

13

10

14

15

ryatia); 6: M. oeconomusKotokel Lake - Kurba River (Buryatia); 8: M. oeconomus 9: M. oeconomus - Texel (Netherlands); land (Netherlands); oeconomus Thibault-de-Couz (Alaska).

Figure

17

3.

Karyotype

CR. Acad. Sci. Paris, 1999.322,473-480

of M. limnophilus Sciences

de

after la vie

C-banding

/ Life Sciences

(CBC).

-

12: M. SaintIsland

M. limnophilus from oeconomus and that to the conclusion that oeconomus is greater M. oeconomusand

limnophilus. 6.3. Canonical

18

11: M. oeconomusKilpisjgrvi (Finland); Jgmtland (Sweden); 13: M. oeconomus (France); 14: M. oeconomus - St. Lawrence

gence of the two populations of Gansu with the Euro-Siberian M. from St. Lawrence Island. This leads the within-species variability of M. than the between-speciesvariabilityof M.

16

(Buryatia); 7: M. oeconomus - Zeeland (Netherlands); 10: M. oeconomus - Fries-

discriminant

The distribution of the populations is presented

analysis centres of gravity of the different on axes l-2 of the canonical

477

F. Courant et al.

i

i

Figure 6. Canonical discriminant analysis: comparative M, morphology of the different populations on axes 1 x 2. Population labels as in figure 4.

correlated with canonical variables 1 (r = 0.41) and 2 (r = 0.39). These observations suggest finally that axes 1 and 2 reflect shape rather than size. In conclusion, morphometric analyses of M, show the differentiation of the two populations of M. limnophilusof Gansu and also the marginal position of M. limnophilus and M. oeconomus of Mongolia compared with the other populations.

7. Systematic Principal component analysis: comparative M, morphology of the different populations on axes 1 x 2 (A) and 2 x 3 (B). Population labels as in figure 4.

Figure

5.

discriminant analysis (figure 6). The canonical discriminant variables of axes 1 and 2 express 43 and 22% of variance, respectively. Axis 1 allows the two Mongolian populations (M. oeconomus and M. limnophilus) to be separated from all of the other populations. Axis 2 separates the two Gansu populations from the others. The projection of the centres of gravity of the various populations shows that the two most remote sets are the two populations from Mongolia and the two Gansu populations, and also that the Netherlands populations are close to those of Buryatia. The morphological variables most closely correlated with canonical variable 1 are vl6 (r = 0.63), v7 (r = 0.59), v20 (r = 0.55) and v23 (r = 0.51). The morphological variables most closely correlated with canonical variable 2 are vl5 (r = 0.69), v2 (r = 0.69), v21 (r = 0.68) and vl (r = 0.65). Variable v6 (total length of M,) is not closely

478

discussion

The first reports of M. oeconomus (M. ratticeps was a former name for M. oeconomus Pallas, 1776) and M. limnophilus in central China are cited in Allen [51. Biichner [7] described M. limnophilus on the basis of five specimens from Tsaidam (Tibet-Qinghai plateaus). This area is located 800 km northwest of our study area. These specimens were collected by Przewalski in the course of his 1879 and 1884 expeditions. Satunin [8] described on the basis of a single specimen M. limnophilus flaviventris from the ‘Tschortentan’ temple in Gansu, 2 400 m elevation [5]. He compared this specimen with M. limnophi/us [7]: it shows ‘much agreement in tooth pattern and general proportions’. Allen gives M. ratticeps flaviventris as synonymous for this species and M. malcolmi [15]. He reports M. ratticeps flaviventrisas being not uncommon in the western part of Gansu, and provides details of the location of Thomas’ specimens. They were found 200-250 km east from our study area, less than 100 km from the border with Shanxi province. Since that time, M. oeconomus and M. limnophilus have obviously been confused, specimens having never C. R. Acad.

Sci. Paris, Sciences

de la vie / Life Sciences 1999,322,4?3-480

Microbus been identified instance, Huang

on the basis of karyotypes et al. [I 61 cite M. limnophilus

so far. For as a subs-

pecies of M. oeconomus. The subspecies M. oeconomus flaviventris 14, 17, 181, M. oeconomus limnophilus [19] and the species M. oeconomus [20] were still recently cited in studies of central China. Those records were located in the Hexi corridor, the Qilian Mountains, over the south Gansu plateaus chuan and in Qinghai.

and

grasslands,

in western

Si-

Study of the lectotype of M. limnophilus [71 from Qaidam (Qinghai, China) from the Przewalski collection (figure I/ shows an M, with a typically ‘ratticepoid’ structure with a simple outline of T6. The holotype of M. limnophi/us flaviventris [8] has exactly the same tooth morphology. Dental characteristics cannot therefore be used to distinguish between the two forms. In conclusion, the synthesis of these analyses shows that the two populations of M. limnophilus from Gansu are fairly remote from the M. limnophilus population of Mongolia both in size and shape of M,. Because of the recent genetic isolation, the two species of Mongolia are morphologically very similar on the structure of M, and it is thus difficult to separate the two Mongolian species. The occurrence of the subspecies M. limnophilus flaviventris [8] which has already been attributed to Gansu forms is confirmed and may be, in the present state of knowledge, synonymous with the type population of M. limnophilus of Qaidam (Qinghai, China). Actually,

karyological

analyses

confirm

the presence

of

M. limnophilus 171 in the Gansu province, but these studies are not available to attest the presence of M. oeconomus in the same area. In Mongolia [14], M. limnophilus was recorded from the south of the great lakes district, in the Dzhungar oasis, and the large valleys (Tatsin-Col, Tuin-Gol, etc.). Thus, the question arises of the actual distribution of M. oeconomus and M. limnophilus in China. Karyological evidence for M. oeconomus and M. limnophilus was provided in northern and southern golia, respectively, as well as in southern Gansu latter. In China, the biotopes where M. limnophilus captured were river banks (grassland with patches sp., Hippophae sp.), slope grassland, scrubland bland (Befula albosinensis, Rosa rufolantatus, mongolicum, Sorbaria kirilowii, Sambucus There, its optimal habitat seemed to be any

Monfor the was of %/ix

and shruViburnum sp., etc.). area with

reasonably dense grass cover and the presence of small bushes under which it could find shelter. In Mongolia, it thrived in wet grasslands, bushy areas, from salty morasses (Elkhon oasis in the vicinity of Altai’) and sand areas, to wetlands (reeds), river banks and lake borders. M. oeconomus was parapatric and could occupy the same kind of habitats, forests and mountain grasslands (up to 3 000 m elevation) in the northern part of the country. If favourable habitats do exist, the distribution range of M. limnophilus should western Gansu,

be expected to becontinuous, part of the Inner Mongolia eastern Xinjiang, northern

CR. Acad. Sci. Paris, Sciences 1999,322,473-480

covering province, Tibet-Qinghai

de la vie / Life Sciences

also the northern plateau

and their spurs. China is puzzling

implied

in alveolar

The presence of M. and calls for further

echinococcosis

oeconomus karyological

in central studies.

8. Conclusion This

paper

provides

the

first

record

of Micro&s

limno-

phi/us in an area of high endemicity for Human alveolar echinococcosis. Epidemiological studies have shown that in South Gansu the prevalence of Human alveolar echinococcosis was higher where shrublands favourable limnophilus had a larger extent [6, 211. Moreover, been shown experimentally that M. limnophilus

to M. it has was a

good physiological host for Echinococcus multilocularis (Zhou et al., unpublished data). It can therefore be suspected that M. limnophilus should play an important role in the life cycle of Echinococcus multilocularis in southern Gansu. Microtus oeconomus, morphologically similar, has been shown to be a key species for transmission in other countries such as Siberia and Alaska [22]. Moreover, the species was believed to be recorded in western China, including Gansu Consequently, the cies in China and

and some bordering respective distribution their subsequent role

provinces [16]. of the two spein transmission

need clarification since earlier identifications med on the only basis of morphology which be considered valid.

were perforcan no longer

M. limnophilus of Mongolia is morphologically similar to M. oeconomus. Dental analyses indicate the overall morphological homogeneity of the populations of M. oeconomusof Europe and especially their proximity with the populations of Buryatia. The specimens of the Micro&s population collected in Zhang County lar dental morphology to M. oeconomus of M. limnophilus of Mongolia.

(Gansu) with

have some

simitraits

The two populations of M. oeconomus and M. limnophi/us of Mongolia, which are different species by their karyotypes, are morphologically very close to one another, perhaps because of recent genetic isolation, which makes them a good example of sibling species. Consequently, it can be concluded from these data that there is a within-species difference between M. limnophilus of Gansu

compared

In view the Gansu of Mongolia,

with

those

of Mongolia.

of the differences in shape observed between populations and the M. limnophilus population we propose that the Mongolian form should

be distinguished malygini nov. - Microtus

ssp.

under (figure

the name 7).

limnophilus

Holotypus: skull Malygin collection

malygini

Microtus nov.

Moscow,

Derivatio conducted

nominis: in tribute to V.M. many studies of these voles. typicus:

Hypodigm:

ssp.

with mandibles, specimen 811, (Zoological Museum of Moscow

University,

Locus Mongolia.

limnophilus

V.M. State

Russia).

Stream 11 specimens

Gun

Tata including

Bulak,

Malygin Dzhungar

who

has Gobi,

holotype.

479

F. Courant

et al

Diagnosis: the distance between sides of middle part of anteroconid broad and somewhat asymmetrical 38; NF = 60. Comparison: Microtus differs from nominative

external and internal is wide; large, very M,; karyotype: 2n =

Locus

limnophilus malygini nav. ssp. Microtus limnophilus limnophilus

by larger size of its M, (morphological conid of M,). - M. oeconomus kharanurensis Similarly, the Microtus different from the other be distinguished rensis nov. ssp.

under (figure

Hoiotypus: skull Malygin collection University, Moscow,

Acknowledgments: 1. Markowski Programme

structure nav.

the 1).

name

ssp.

of Mongolia of Eurasia that

M.

oeconomus

is so it can

kharanu-

This paper

is a contribution

Malygin for avancees

loan of material; franco-chinois

V.M. State

5561

Life-cycle patterns and geographic distribution of Echiin: Thompson R.C.A., Lymbery A.J.S. (Eds.), EchinococDisease, CAB International, Wallingford, Oxon, 1995,

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Cytogenetic chromosomes, centromeric

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Nur

holotype.

symmetrical a globulous

is similar

Finally, the presence which was hitherto

to that

M,

with

an ante-

shape

with

no ~6 to

of

M.

of M. oeconomus generally accepted

by karyological

Biogeosciences-Dijon. support from the European

the loop Morpholo-

limnophilus

a of

in central China must now be

analyses.

The authors Commission

1121 Brunet-Lecomte idae, Rodentia) actuels de Bourgogne, Dijon,

[31 Bobrov V.V., Neronov and Indo-Malayan fauna1 distribution of rodents), Russian).

from

Khara

Mongolia.

narrow,

varies

of origin

forms where T6 may be very developed giving ‘nivaloid’ shape, karyotype: 2n = 30; NF = 60.

confirmed

of UMR CNRS and financial (PRA E95-1).

Nur,

specimens

fairly that

its region

Mongolia.

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