Zootaxa 3599 (6): 564576

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Copyright 2013 Magnolia Press

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Article

ZOOTAXA
ISSN 1175-5334 (online edition)

http://dx.doi.org/10.11646/zootaxa.3599.6.5
http://zoobank.org/urn:lsid:zoobank.org:pub:67AAA0AF-0626-4694-A1F4-1F0BD753D018

Chironomus polonicus sp. n. (Diptera: Chironomidae) from southern Poland

PARASKEVA MICHAILOVA1,4, ANDRZEJ KOWNACKI2 & PETER H. LANGTON3
1

Institute of Biodiversity and Ecosystem Research, Bulgarian Academy of Sciences, 1 Tzar Osvoboditel boulv., 1000 Sofia, Bulgaria
Institute of Nature Conservation, Polish Academy of Sciences, al. A. Mickiewicza 33, 31120 Krakow, Poland
3
University Museum of Zoology, Downing Street, Cambridge, UK (Address for correspondence: 16 Irish Society Court, Coleraine, N.
Ireland BT52 IGX)
4
Corresponding author. E-mail: michailova@zoology.bas.bg
2

Abstract
The paper describes larval, pupal and adult morphology as well as the karyotype of Chironomus polonicus sp. n. from
southern Poland. The material has been obtained from reared egg masses collected in Bolesaw pool, near Krakw. The
species belongs to the pseudothummi cytocomplex with 2n = 8 and chromosome arm combinations AE, BF, CD, G.
Several homozygous inversions distinguish arm A of the new species from that of C. pseudothummi Strenzke. Arm F is
similar to that of C. aprilinus Meigen and differs from it by few steps of homozygous inversions. Few morphological
differences in the pupa and adult are also presented.
Key words: Diptera, Chironomus polonicus, new species, polytene chromosomes, morphology, southern Poland

Introduction
Chironomidae are a widely distributed and abundant group of insects in freshwater ecosystems. They play an
important role in these ecosystems, accounting for 25100 % of the macroinvertebrate species (Kownacki 2011).
Currently 1259 species and subspecies of Chironomidae are known in Europe (Sther & Spies 2012). One of the
most abundant is the genus Chironomus Meigen-85 species.
Chironomidae species possess easily identifiable polytene chromosomes, with species-specific banding
patterns and these can be used as cytogenetic markers to provide additional signs for studying taxonomy, evolution
and phylogeny (Michailova 1989, Kiknadze et al. 1991, Wuelker et al. 2011). Most species of the genus
Chironomus have been described on the basis of the species-specific cytogenetic markers.
In the present paper a new Chironomus species is described from southern Poland on the basis of all stages of
its metamorphosis (except female) together with its karyotype.

Material and methods

The material was obtained from egg masses collected from a small pond (max. depth 1 m) situated on a recent mine
spoil in Bolesaw in the Olkusz region with Zn-Pb ore deposits (southern Poland) in June 2001 and reared in
laboratory conditions in Sofia, following Michailovas (1985) method.
The 4th larval instar was fixed in ethanol-glacial acetic acid (3:1). The chromosome preparations were made
from salivary glands following Michailova (1989). The identification of chromosome banding patterns follows
Keyl (1962) for arms A, E, and F. In order to identify the position of the centromere regions the chromosome C
banding method has been applied (Michailova 1987).
For each specimen the chromosome preparation, larval head capsule and body were slide mounted in Euparal.
The description of larval morphology follows Sther (1980).
The preparations of salivary gland chromosomes and larval morphology are stored in the Bulgarian Academy
of Sciences, Institute of Biodiversity and Ecosystem Research, Sofia.

564 Accepted by W.Gika: 26 Dec. 2012; published: 10 Jan. 2013

Results
Chironomus polonicus sp. n.
Type material. Holotype: larva, chromosome squash with larval head capsule and hind part of abdomen, labeled:
6 polytene chromosomes, larva - external morphology, 06.2002; Bolesaw (50o1726.61E, 19o2636.93 N)
(Olkusz Region with Zn-Pb ore deposits), near Krakw, Poland. Paratypes: same sampling data as holotype, males
(2) and females (3) in Krakw, Institute of Natural Conservation, PAS; pupal exuviae (3) slides in UK, coll. PHL.
Material examined. Ten chromosome preparations and external larval morphology; 3 adult males and 4
pupae.
Source of material. Egg masses on water plants in the Bolesaw pool, reared under laboratory conditions.
Derivation of the name. From the old name of Poland.

Larval morphology
Dark red body, length up to 80 mm, with very short lateral tubules and 2 pairs of long ventral rounded at apex
tubules. Posterior part of gula and frontoclypeus light. The setae SI serrated on both side, apically with strong setae,
well seen on one side. SII simple.
Mandible (Fig. 1a, b). Three very dark teeth - 3, 4, 5. The 6th tooth the smallest and very pale (Fig. 1b). The
minute 1st tooth very rudimentary and observable only in the right orientation of the mandible. The 2nd tooth small
and pale.
Submentum (Fig. 1c). The teeth of submentum dark with median tooth trifid (4145 m wide); 6 pairs of
lateral teeth, the lateral teeth overall decreasing in size from 1st to 6th; 1st and 2nd very closely approximated; 4th and
5th teeth of much the same size (Fig. 1c). The inner surface of the ventromental plates smooth. (Fig. 1f).
Premandibles (Fig. 1d). Pale, with 2 teeth. The ventral tooth thinner and slightly shorter than the dorsal tooth.
Pecten epipharyngis with 1214 teeth.
Antenna (Fig. 1e). Antennal blade extending to the end of the 3rd segment. LO distinct. RO on the basal
segment at about 1/3 the length of the segment. Measurements of the antenna in Table 1.

TABLE 1. Chironomus polonicus sp. n., measurements on larval antennae. L1 and L2length of antennal segments 1 and 2 in
m; W1width of antennal segment 1 through ring organ.
Number of specimens

L1

L2

W1

L1/L2

L1/W1

131.2

38.7

32

3.39

4.1

127.1

32

41

3.96

3.09

123

32

41

3.84

127.1

38.7

41

3.28

3.1

127.1

38.7

41

3.28

3.1

123

32

38.7

3.84

3.18

127.1

32

41

3.96

3.09

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FIGURE 1. Larva morphology of Chironomus polonicus sp. n. a: mandible; b: mandible with distinct teeth 3, 4, 5, 6; c: submentum; d: premandible; e: antenna; f: ventromental plate of submentum. Bar = 100 m

Karyotype
The species belongs to the pseudothummi cytocomplex with 2n = 8 and chromosome arm combinations: AE BF
CD G (Fig. 2c; Fig. 3a; Fig. 4a, b). Chromosome morphology was defined by localization of the constitutive
heterochromatin (Figs. 5ad). Chromosome AE - submetacentric, easily visible by C band staining (Fig. 5a).
Chromosomes BF and CD - metacentric (Fig. 5b, c), chromosome G - acrocentric (Fig. 5d), with one Balbiani ring
(BR) and one nucleolar organizer (NOR) located at the telomere (Fig. 4b, Fig. 5d).
Arm A (Figs. 2 ac): different from C. piger Strenzke and C. pseudothummi Strenzke by several steps of
homozygous inversions.
C. piger (Fig. 2a): 1 - 2 - 3 - 4 - 5 - 6 - 7 - 8 - 9 - 10 - 11 - 12 - 13 - 14 - 15 - 16 - 17 - 18 - 19.

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C. pseudothummi (Fig. 2b): 1 - 2 - 3 - 12 - 11 - 10 - 9 - 8 - 7 - 6 - 5 - 4 - 13 - 14 - 15 - 16 - 17- 18 - 19.

Chironomus sp.: 1 - 2 - 3 - 4 - 5 - 6 - 7 - 8 - 9 - 10 - 11 - 12 - 13 - 14 - 15 - 16 - 17 - 18 - 19.
C. polonicus (Fig. 2c): 1 - 2 - 3 - 10 - 9 - 8 - 7- 6 - 5 - 4 - 11 - 12 - 13 - 14 - 15 - 16 - 17 - 18 - 19.

FIGURE 2. Salivary gland chromosome arms A and E. a: Chromosome arm A of Chironomus piger Strenzke (according to
Keyl, 1962); b: Chromosome arm A of Chironomus pseudothummi Strenzke (according to Keyl, 1962); c: Chromosome AE of
Chironomus polonicus sp. n. [band patterns of arm A can compare with the same arm of C. piger (a) and C. pseudothummi (b);
band patterns of arm E can compare with the same arm of C. pseudothummi (d)]; d: Chromosome arm of Chironomus
pseudothummi Strenzke [according to Keyl (1962)]. The arrow indicates the localization of the centromere. Bar = 100 m.

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FIGURE 3. Salivary gland chromosome arms B and F. a: chromosome BF of Chironomus polonicus sp. n. [band patterns of
arm F can compare with the same arm of C. aprilinus (b)]; b: chromosome arm F of Chironoimus aprilinus Kieffer [according
to Keyl (1962)]. The arrow indicates the localization of the centromere. Bar = 100 m.

FIGURE 4. Salivary gland chromosome arms C, D and G. a: chromosome CD of C. polonicus sp. n; b: chromosome G of C.
polonicus sp. n. BRBalbiani Ring, NORNucleolar organizer. The arrow indicates the position of the centromere. Bar = 100
m.

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FIGURE 5. Localization of the centromere region detected by C banding method. a: chromosome AE; b: chromosome BF;
c: chromosome CD; d: chromosome G. BRBalbiani Ring, NORNucleolar Organizer. The arrow indicates the position of
the centromere. Bar =100 m.

Arm E (Fig. 2c, d): similar to band patters of C. pseudothummi.

C. pseudothummi (Fig. 2d): 1 - 2 - 3 - 4 - 5 - 6 - 7 - 8 - 9 - 10 - 11 - 12 - 13.
C. polonicus (Fig. 2c): 1 - 2 - 3 - 4 - 5 - 6 - 7 - 8 - 9 - 10 - 11 - 12 - 13.
Arm B (Fig. 3a, b): near to the telomere a dark band with many light bands after it.
Arm F (Fig. 3a, b): Similar to that of C. aprilinus Kieffer but distinguished by several homozygous inversions
steps:
C. aprilinus (Fig. 3b): 1 - 2 - 3 - 4 - 5 - 6 - 7 - 8 - 9 - 10 - 15 - 14 - 13 - 12 - 11 - 16 - 17 - 18 - 19-20.
Chironomus sp.: 1 - 2 - 3 - 4 - 5 - 6 - 10 - 9 - 8 - 7 - 15 - 14 - 13 - 12 - 11 - 16 - 17 - 18 - 19 - 20.
C. polonicus (Fig. 3a): 1 - 2 - 3 - 4 - 5 - 6 - 10 - 9 - 8 - 7 - 14 - 15 - 13 - 12 - 11 - 16 - 17 - 18 - 19 - 20.
Arm C (Fig. 4a): the two groups of dark bands which are separated by a distinct single band can be used as a
marker. The typical constriction of arm C is located near to the centromere region (indicated by an arrow).
Arm D (Fig. 4a): the band sequences near to the telomere are the markers for the arm. The thick dark band located
in the middle of the arm is also a good marker.
Arm G (Fig. 4b): paired, the NOR at one end, a BR in the middle and after it a constriction.
The species is rich in somatic inversions (Michailova et al. 2012) and in 9 individuals there are heterozygous
translocations between chromosomes BF and CD.
Diagnosis. The karyotype of the species is easily distinguished from C. pseudothummi (cytocomplex
pseudothummi) by homozygous inversions in arm A. Band patterns of chromosome E is similar to that of C.
pseudothummi. Arm F has a similar banding pattern to that of C. aprilinus (Keyl 1962) and distinguished from it by

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two steps of homozygous inversions. Chromosome G of C. polonicus is small, always with paired homologues, one
NOR and one BR.

Pupa (n = 4, exuviae)
Medium sized pupae, 8.08.8mm long. Exuviae pale brown, cephalothorax blotched gold and brown, lateral
margins of abdominal tergites VVIII brown; tergites IIIV(V) with brown streak along inner margins of lateral
adhesion markings, truncated on segment V. Spur of segment VIII brown. Anal lobes are entirely brown (Fig. 7a).
Cephalothorax. Cephalic tubercles broadly conical, 88132m long, ending in frontal setae 3650m long (Fig.
6c). Thoracic horn richly branched, plumose. Basal ring of thoracic horn 160x72176x80 m, tracheal patch
120x40132x40 m about 1516 tracheoles across, tracheoles 2 m in diameter. Thorax small granulate anteriorly,
with no stronger granules by suture, by mid scutum reticulate (Fig. 6a), the granules larger and shallower and
squashed in on themselves in mounts.

FIGURE 6. Pupal exuviae structure of Chironomus polonicus sp. n. (a, c, e, g) and C. commutatus (b, d, f, h). a, b: scutal
granulation around dorsocentral setae (anterior to right); c, d: frontal apotome and cephalic tubercles; e, f: spur of segment VIII;
g, h: armament of tergite IV mid lateral. Bar = 0.1 mm.

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FIGURE 7. Pupal exuviae of Chironomus polonicus sp. n. a: general view of andomen; b: segment II dorsal; c: segment IV
dorsal (the same patch shape occurs on thergites III and V); d: segment VI dorsal; e: segment VII dorsal (setae D1 are broken,
its place attachment D1 without points is shown); f: segment VIII dorsal with a spur, and anal segment. Bar = 100 m.

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Abdomen. Tergite I unarmed, IIV with median undivided patches of dense short non-imbricating points (Fig.
6g), larger posteriorly. Armament on TII distinct, rectangular (Fig. 7b). Armament on TIIIV bell-shaped, similar
in size on each tergite (Fig. 7c). TVI with point patch narrowing posteriad, extending to a little past mid segment;
nearly circular patch of stronger points around each seta D5. Point patch of TVI may be divided into three parts
(Fig. 7d). TVII with transverse band of minute points anterior to setae D1 narrowed or completely divided (Fig.
7e). TVIII with 2 lateral patches of small points, not arranged in rows (Fig.7 f). Anal segment without shagreen.
Paratergite V with anterior longitudinal band of small points; V, VI and VII with small patch of points posteriorly.
Pleura of segment IV unarmed. Ventral shagreen: on segment II lateral longitudinal bands connect with median
patch posteriorly, on segment III lateral bands extend for about 2/3 segment length, on segment IV absent. Hook
row 0.50 segment breadth, with about 100 hooks. Vortex distinct on segment IV; conspicuous pedes spurii B
present on segment II. Segment VIII with dark brown, strong postero-lateral spurs each with 1(2) sturdy apical
tooth (Figs.6 e, f). For chaetotaxy of abdominal segments see Table 2.
TABLE 2. Chironomus polonicus sp. n., chaetotaxy of abdominal segments (one side).

Ds

II

III

IV

VI

VII

VIII

Dt
Ls

IX
1

Lt

(4)5

100

Comments. The pupal exuviae runs to Chironomus (s. str.) commutatus Keyl in Langton (1991), couplet 71
and Langton and Visser (2003), couplet 212. The main distinguishing character between C. commutatus and C.
polonicus lies in the granulation of the thorax: in C. commutatus it is composed of very small granules over the
whole scutum, whereas in C. polonicus it is small granulate only anteriorly, the greater part of the scutum covered
with a reticulate pattern formed of larger, shallower, weaker granules (Figs. 6a, b); the granules close to the suture
anteriorly are much enlarged in C. commutatus, but remain small in C. polonicus. There is generally more than one
tooth on the spur of segment VIII in C. commutatus (24), the teeth narrowed to apex, whereas in C. polonicus
there is generally one tooth that suddenly narrows to an apiculate tip (Figs. 6e, f). Both C. polonicus and C.
commutatus have non-imbricating points on tergite IV (Figs. 6g, h), a character not recorded for C. commutatus
previously.

Adult male
Medium size species with total length about 7.0 mm. Wing length 3.5 mm.
Color. Gold-brown with dark-brown transverse markings on tergites IIV (V) of the abdomen. Wing lightgreyish with brown veins, without markings. Legs uniformly brown.
Abdomen (Fig. 8a). Tergites IIV with distinct, dark-brown, transverse markings. Marking of tergite I the
widest, on tergite V difficult to see. All tergites covered with very fine microtrichia, with setae arising on pale areas
without microtrichia. The length of setae not exceeding 1/3 the length of segment. Sternites are uniformly coloured.
Antenna. With 11 flagellomeres, bearing whorls of long setae. Ultimate flagellomere with tip slightly extended,
covered with short hairs, apex sharp. Flagellomere lengths: 77: 32: 32: 32: 34: 35: 33: 30: 29: 24: 1065 m. AR
3. Large globular pedicel without setae (Fig. 8b).
Head. Eye bare, with strong, parallel-sided dorsomedial extension. Vertical setae placed in row above dorsal
eye elongation (Fig. 8c). Frontal tubercles well developed. Ocellus conical, covered with short hairs, about 50 m
(Fig. 8 d). Clypeus with 1821 long setae regularly distributed over entire surface. Palpomeres lengths: 31: 16: 64:
62: 102 m. First palpomere with row of bristles, palpomeres 25 with a few long setae. All palpomeres densely
covered with rows of microtrichia.

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FIGURE 8. Male of C. polonicus sp. n. a: abdomen with marks on tergites; b: antenna; c: head; d: ocellus. Bar = 100 m.

Thorax. Antepronotum narrow, with 24 short dorsal setae. Mesonotum brown with pale brown longitudinal
bands, covered with very fine microtrichia. Acrostrichals arising from small, pale sockets on anterior part of
mesontum; dorsocentrals uniserial, arising on pale areas; 5 prealar setae present; humeral, anepisternal and
preepisternals indistinct. Humeral pit distinct, kidney-shaped. Scutellum with unordered long setae.
Wing. Wings and veins typical for this genus.
Legs. Fore tibia without spur, mid and hind tibiae with closely approximated combs, each bearing spur (about
2025 m long) (Figs. 9a, b). Legs densely covered with long setae. Legs measurements in Table 3.
TABLE 3. Chironomus polonicus sp. n., length of legs segments, bristle - and leg ratios.
fe

ti

ta1

ta2

ta3

ta4

ta5

BR

LR

p1

1307

1260

1800

1099

785

680

220

2.2

1.4

p2

1407

1353

825

516

406

290

220

2.2

0.6

p3

1625

1700

1195

730

580

380

210

2.9

0.7

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FIGURE 9. Legs of C.polonicus sp. n. a: tibia of first leg without comb and spur; b: hind legtibial comb and spur. Bar = 50
m.

Hypopygium (Fig. 10). Anal tergite bands dark brown, V-type, separated medially; pale brown median area
with about 68 long setae. Posterior margin of anal tergite rounded without lobes, with group of long setae on both
sides of anal point base (about 8). Anal point slender, narrowing in middle part, apex rounded. Superior volsella
distally sickle-shaped, pointed, without bristles (E-type according to Strenzke 1959). Inferior volsella twice as long
as superior volsella, reaching half length of gonostylus, with curved long setae in distal part. Gonostylus narrowed
subapically, with group of 8 bristles at inner margin near apex.
Comments. The male is similar to those of the plumosus complex as indicated by the slender anal point,
slender superior volsella, well developed frontal tubercles, LR1 1.4. However, it differs from that group in a number

of characters: AR 3 (species of the plumosus group have usually AR > 4), BR < 3 (vs. BR > 4 in plumosus
group). Gonostylus is narrowed subapically as in species of the riparius group.

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FIGURE 10. Hypopygium of C. polonicus sp. n..

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Acknowledgements
This study was supported by the exchange Program between the Polish and the Bulgarian Academy of Sciences
(2012). The authors thank both referees for evaluating the manuscript and valuable suggestions. The assistance of
Dr. W. Gika is greatly acknowledged.

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