Two new species of the genus Deuteraphorura Absolon, 1901
(Hexapoda, Collembola, Onychiuridae) from Georgian caves with

remarks on the subterranean biodiversity of the Caucasus Mountains

Andrea PARIMUCHOVÁ 1,*, Shalva BARJADZE 2,
Eter MAGHRADZE 3 & Ľubomír KOVÁČ 4

1,4 Institute of Biology and Ecology, Faculty of Science, Pavol Jozef Šafárik University in Košice,
Šrobárova 2, 04154 Košice, Slovakia.

2,3 Institute of Zoology, Ilia State University, Giorgi Tsereteli 3, 0162, Tbilisi, Republic of Georgia.

* Corresponding author: andrea.parimuchova@upjs.sk
2 Email: shalva.barjadze@yahoo.com

3 Email: eter.magradze.1@iliauni.edu.ge
4 Email: lubomir.kovac@upjs.sk

1 urn:lsid:zoobank.org:author:6533715D-0789-401A-BFFD-91ABDB36428B
2 urn:lsid:zoobank.org:author:AB36BEF1-C006-41A3-861E-E0B79EE35FBF

3 urn:lsid:zoobank.org:author:A5CE76D6-214B-40D8-B06D-181C0AB4D7C9
4 urn:lsid:zoobank.org:author:64C6117E-DB5C-4569-BF89-5C812C118760

Abstract. Specimens of Deuteraphorura collected in 11 Georgian caves were analysed morphologically
and molecularly based on the COI gene barcode region. Two molecular delimitation methods revealed
four species (MOTUs); however, only two of them were distinguished morphologically and are described
in this paper as new to science. Both new species, D. colchisi sp. nov. and D. kozmani sp. nov., belong to
the group with a pseudocellus on the first thoracic tergum; the differential diagnosis table to this species
group is provided. The potential of the Caucasus as a hotspot region of subterranean biodiversity and
evolution centre of subterranean animals is discussed.

Keywords. Cave fauna, species delimitation, MOTUs, hotspot, evolution centre.

Parimuchová A., Barjadze S., Maghradze E. & Kováč Ľ. 2023. Two new species of the genus Deuteraphorura
Absolon, 1901 (Hexapoda, Collembola, Onychiuridae) from Georgian caves with remarks on the subterranean
biodiversity of the Caucasus Mountains. European Journal of Taxonomy 879: 64–82.
https://doi.org/10.5852/ejt.2023.879.2161

Introduction
The biodiversity of caves has been one of the main topics of subterranean biology for decades. Within
the Palearctic region, “hotspot areas” were defined in the mountains of southern Europe based on the
diversity of obligate cave-dwellers (troglobionts and stygobionts) (Culver et al. 2004, 2006; Reboleira
et al. 2011). Europe is traditionally a centre of speleobiological research, resulting in rich data on karst

European Journal of Taxonomy 879: 64–82 ISSN 2118-9773
https://doi.org/10.5852/ejt.2023.879.2161 www.europeanjournaloftaxonomy.eu
2023 · Parimuchová A. et al.

This work is licensed under a Creative Commons Attribution License (CC BY 4.0).

R e s e a r c h a r t i c l e

urn:lsid:zoobank.org:pub:B8BE4779-EF8D-4B63-8643-E1468F306C5B

https://orcid.org/0000-0002-8977-9991
https://orcid.org/0000-0001-8992-4987
https://orcid.org/0000-0002-4796-9439
https://orcid.org/0000-0001-8194-2128
mailto:andrea.parimuchova%40upjs.sk?subject=
mailto:shalva.barjadze@yahoo.com
mailto:eter.magradze.1%40iliauni.edu.ge?subject=
mailto:lubomir.kovac%40upjs.sk?subject=
https://zoobank.org/urn:lsid:zoobank.org:author:6533715D-0789-401A-BFFD-91ABDB36428B
https://zoobank.org/urn:lsid:zoobank.org:author:AB36BEF1-C006-41A3-861E-E0B79EE35FBF
https://zoobank.org/urn:lsid:zoobank.org:author:A5CE76D6-214B-40D8-B06D-181C0AB4D7C9
https://zoobank.org/urn:lsid:zoobank.org:author:64C6117E-DB5C-4569-BF89-5C812C118760
https://doi.org/10.5852/ejt.2023.879.2161
https://creativecommons.org/licenses/by/4.0/
https://zoobank.org/urn:lsid:zoobank.org:pub:B8BE4779-EF8D-4B63-8643-E1468F306C5B

regions and their subterranean fauna. It has been observed that areas of maximum diversity occur in the
range of ca 42°–46° in Europe and 34° in the North America, but potential hotspots were also expected
in the karst regions of the western Caucasus in Georgia (Culver et al. 2006) and in South-East Asia, with
a high level of endemism in troglobiotic taxa (Deharveng & Bedos 2000). The last mentioned has been
confirmed, and the region of southern China is recently the richest in troglobiotic Trechini beetles (e.g.,
Tian et al. 2016, 2017). Biodiversity-rich areas were also identified in Brazil (Souza-Silva & Ferreira
2016).

Caves in the Caspian territory of the Palearctic have been almost neglected in terms of biospeleology.
Recent studies have made a significant contribution to revealing the subterranean diversity and
documenting the great potential of this area for the discovery of the new taxa. Caucasian subterranean
fauna has been intensively studied mostly in Georgia, resulting in regularly annotated lists of cave
fauna (Barjadze & Djanashvili 2008; Barjadze et al. 2012, 2015) with a total of 86 troglobiotic and
stygobiotic species as of 2019 (Barjadze et al. 2019). In the last decade, new species of Collembola
have been described in the genera Deuteraphorura Absolon, 1901 (Jordana et al. 2012), Arrhopalites
Börner, 1906 and Pygmarrhopalites Vargovitsh, 2009 (Vargovitsh 2012, 2013, 2017, 2022), and
Plutomurus Yosii, 1956 (Barjadze et al. 2022) and the new genus Troglaphorura, with highly
troglomorphic species (Vargovitsh 2019). Unexpected diversity was documented in Diplopoda
(Antić & Makarov 2016; Antić & Reip 2020), and new troglomorphic taxa were described in
Isopoda (Gongalski & Taiti 2014) and Opilionida, with the highly specialised species in the genus
Nemaspela (Martens et al. 2021) and Pseudoscorpionida of the genus Globochtonius (Zaragoza
et al. 2021) .

In the present contribution, we describe two new collembolan species of the genus Deuteraphorura
discovered during recent biological investigations in the caves of western Georgia, and discuss the
diversity indicated by molecular analyses. We emphasise the significance of caves in the Caucasian
Mountains and consider this important mountain range to be a hotspot area of subterranean biodiversity.

Material and methods
Study area
In Georgia, the total area of the karstic rock outcrop occupies 4475 km2 with more than 1500 caves
(Asanidze et al. 2019).

Specimens of Deuteraphorura from 11 relatively warm horizontal caves of low altitudes in Georgia
were morphologically studied and molecularly analysed (Table 1). We called the cave close to
Prometheus and Datvi Cave as Sakadzhia Cave; however, there are doubts about the real location of this
cave (Barbakadze pers. com.). In this study, populations were defined as individuals collected in caves
located in different karst areas of Georgia with a minimum distance of ca 2.5 km between the closest
caves (Motena–Inchkhuri) and maximum distance of ca 85 km (Motena–Kozmani) (Fig. 1).

Morphological examination

For morphological study, specimens were separately mounted on permanent slides in Swann medium
(Liquido de Swann) modified after Rusek (1975) and studied in phase-contrast Carl Zeiss Axio 5
microscope and Leica DM 2500 microscope equipped with DIL optics (differential interference
contrast), a measuring eyepiece (micrometric ocular) and a drawing arm. The images were taken with
an Axiocam 208 color (Carl Zeiss) camera with ZEN imaging software. Drawings were edited using
Adobe Photoshop CS6. Chaetotaxy of the tibiotarsus is presented after Deharveng (1983), and of the
labium after Fjellberg (1999).

PARIMUCHOVÁ A. et al., Two new cave Deuteraphorura species from Georgia

Molecular data analysis and species delimitation methods
One to three specimens from each population were analysed in the molecular laboratory of the Department
of Zoology, IBE FS UPJS, Košice, Slovakia.

To prevent contamination, all DNA laboratory work was conducted under sterile conditions with the use
of barrier tips. Total DNA was extracted with the Machrey-Nagel NucleoSpin Tissue Kit according to
the modified manufacturer´s protocol with 50 μL of elution buffer twice. A polymerase chain reaction
(PCR) (Saiki et al. 1988) was carried out using a 12.5 μL reaction volume consisting of 1 μL of template
DNA (not quantified), 10× PCR Buffer (TopBio), 12.5 mM of dNTP mix, 5 μM of each primer and 0.125
units of Taq polymerase (TopBio) on a GenePro (Bioer Co. Ltd, China) thermal cycler. A fragment of the
COI gene (588 bp) was amplified using the primers LCO1490_JJ (5’ cha cwa ayc ata aag ata tyg g-3’)
and HCO2198_JJ (5’- awa ctt cvg grt gvc caa ara atc a -3’; Astrin & Stüben 2008). Thermal cycling
conditions were as follows: 94°C for 3 min followed by 5 cycles of 94°C for 30 sec., 45°C for 1 min
30 sec. and 72°C for 1 min, followed by 35 cycles of 94°C for 30 sec., 51°C for 1 min 30 sec. and 72°C
for 1 min followed by 1 min in 72°C. After verification on agarose electrophoresis, reaction products
were purified using Exo I/FastAP (Thermo Fisher Scientific). The sequencing of the purified products
was performed using LCO1490_JJ by the Sanger method (Eurofins Genomics, Ebersberg, Germany).
In cases when the primer failed to produce high quality chromatogram, reverse primer sequencing was
employed. Sequences were edited and trimmed of unreadable short stretches (ca 30 bp at the 5’ and 3’
ends) with Geneious Prime ver. 2022.1.1 (Copyright © 2005–2022 Biomatters Ltd).

Since none of the sequences contained stop codons or indels in ORF, all were considered to be true
mitochondrial and not nuclear copies. All the sequences were verified as consistent with Onychiuridae
congeners using the GenBank BLASTn search (the Mega Blast algorithm with the default setting).
Sequences were aligned with the Geneious Prime ver. 2022.1.1 (Copyright © 2005–2022 Biomatters
Ltd) software by Muscle (Codons) algorithm using the Invertebrate Mitochondrial GeneCode and

Fig. 1. Locations of studied caves in Georgia. For cave abbreviations see Table 1.

European Journal of Taxonomy 879: 64–82 (2023)

default parameters. Standard DNA barcoding distance analysis was conducted in MEGA X (Kumar
et al. 2018) F using the Tamura-3 parameter method (Tamura 1992). A neighbour-joining tree (Saitou &
Nei 1987) with Tamura-3 parameter method (Tamura 1992) was constructed and the robustness of the
tree nodes was assessed by bootstrap analysis with 1000 replications, values under 60 are not shown.

Both barcoding gap- and evolutionary models were applied for COI marker. Assemble Species by
Automatic Partitioning (ASAP) method (Puillandre et al. 2021) used genetic distances to propose species
hypotheses. The Kimura (K2P) model with default parameters was used to merge sequences into groups.

The Poisson tree processes (PTP) model, used for species delimitation based on the number of
substitutions, was performed using on-line software (Zhang et al. 2013). A maximum likelihood (ML)
tree was inferred using Auto substitution model and 1000 Ultrafast bootstrap analysis (Hoang et al.
2018) in IQ-TREE software (Nguyen et al. 2015).

Correlation between geographical and genetic distances (Tamura-3 parameter model, pairwise deletion
option) of populations was evaluated by Mantel test (999 permutations) using the GenAlEx 6.5 program.

All new sequences are available in GenBank (accession numbers: OQ271838–OQ271861).

Abbreviations
Ant. = antennal segment
Abd. = abdominal tergum
AOIII = antennal organ of the third antennal segment

Table 1. List of caves with their characteristics, and administrative and geographic location in Georgia.
Cave characteristics according to Tatashidze et al. (2009) and Tsikarishvili & Bolashvili (2013). Names
of tourist caves are bolded. Abbreviations: AK = Askhi karst massif; Ch = Chiatura; EE = entrance
elevation; et = eutrophic; IM = Imereti; Kg = Kharagauli; Kh = Khoni; Ma = Martvili; mt = mesotrophic;
OP = Odishi Plateau; ot = oligotrophic; SAM = Samegrelo; ST = Sataplia-Tskaltubo; Ts = Tskaltubo;
X = site of air temperature measurements in caves is unclear; Y = elevation between the entrance and the
deepest site of the cave; ZI = Zemo-Imereti Plateau; * trophic level estimation is based on presence of
organic material in internal parts of the cave; – = not measured.

Abb. Cave name EE
(m a.s.l.)

Cave
length (m)

Cave
temp. X
(°C)

Cave
depth Y
(m)

Trophic
level*

Region District Karst
area

Xom Khomuli 95 70 13.5–14 2 mt IM Ts ST
Mel Melouri 424 5300 12–13 15 ot IM Ts ST
Sak Sakadzhia 141 – – – mt IM Ts ST
Ssb Satsurblia 305 125 11.7 20 mt IM Ts ST
Pro Prometheus 147 2900 13.5–14.5 – et IM Ts ST
Dat Datvi 140 56 – – mt IM Ts ST
Sat Satevzia 215 250 – 0 mt IM Kh ST
Koz Kozmani 652 200 – – mt IM Kg ZI
Shv Shvilobisa 730 1000 12.3 10 ot IM Ch ZI
Mot Motena 570 95 13–13.6 14 ot SAM Ma AK
Ink Inchkhuri 380 65 – – et SAM Ma OP

PARIMUCHOVÁ A. et al., Two new cave Deuteraphorura species from Georgia

IBE FS UPJS = Institute of Biology and Ecology, Faculty of Science, P.J. Šafárik University,
Košice, Slovakia
IZISU = Institute of Zoology, Ilia State University, Tbilisi, Georgia
ms = microsensillum
MVO = male ventral organ
PAO = postantennal organ
pso = pseudocellus
psx = parapseudocellus
Tita = tibiotarsus
Th. = thoracic tergum
VT = ventral tube

Results
Molecular species delimitation
We employed delimitation methods based on the COI mitochondrial gene to define the molecular
operational taxonomic units (MOTUs) and assess their congruence with the current species level based
on morphology and geographic distribution.

We obtained alignment of 24 COI sequences with a length of 605 bp.

Fig. 2. A neighbour-joining tree (NJ) with species delimitation of Georgian cave populations of
Deuteraphorura Absolon, 1901 based on COI molecular marker, morphology and geographic location
in karst areas. Numbers and coloured columns indicate groups (species) identified by particular methods
ASAP (Assemble Species by Automatic Partitioning) and bPTP (Bayesian Poisson tree processes).
The question mark (?) indicates ambiguous result in Shvilobisa Cave due to low number of studied
specimens. For abbreviations of caves in the NJ tree see Table 1.

European Journal of Taxonomy 879: 64–82 (2023)

The ASAP method delimited four species, and the best partition had an ASAP score of 2.0 (p <0.05).
The bPTP method estimated four groups (species) with support from 0.519 to 1.0, thus corresponding
to the ASAP delimitation (Fig. 2).

The distribution of K2P distances revealed a clear barcode gap. As determined with ASAP, specimens
diverging at a K2P distance above 3% belong to different species (Fig. 3).

Species 1 comprised specimens from most of the Sataplia-Tskaltubo Karst caves: Khomuli, Satsurblia,
Melouri, Prometheus, Datvi and Sakadzhia. Species 2 contained specimens from caves of three karst
areas represented by Satevzia Cave, Motena Cave and Inchkhuri Cave. Species 3 comprised only a
single specimen from Shvilobisa Cave, and species 4 consisted exclusively of specimens from Kozmani
Cave (Fig. 2).

Morphological character analysis was able to confirm the species status of only two of the four MOTUs
revealed by molecular delimitation methods; these two species have been given scientific names and are
described taxonomically below.

Fig. 3. Histogram of COI K2P distances between specimens of Deuteraphorura Absolon, 1901. The
red line indicates the threshold distance above which specimens are considered to belong to different
species, according to ASAP method.

PARIMUCHOVÁ A. et al., Two new cave Deuteraphorura species from Georgia

Taxonomy
Phylum Arthropoda von Siebold, 1848
Subphylum Hexapoda Blainville, 1816

Class Collembola Lubbock, 1870
Order Poduromorpha Börner, 1913

Family Onychiuridae Lubbock (in Börner, 1913)
Subfamily Onychiurinae Börner, 1901
Genus Deuteraphorura Absolon, 1901

Deuteraphorura colchisi Parimuchová, Barjadze & Kováč sp. nov.
urn:lsid:zoobank.org:act:AAC4848A-79D3-4A5C-8458-A8CB7BB2EA8F

Fig. 4, Table 2

Deuteraphorura sp. – Zaragoza et al. 2021.

Etymology
The name is derived from ‘Colchis’ – the historical geographical, ethnical and political entity of Georgia
which today is located in the west of the country.

Type material
Holotype

GEORGIA • ♀; Imereti, Tskaltubo, Satsurblia Cave; 42.38805000° N, 42.60626700° E; 12 Mar. 2020;
Eter Maghradze leg.; hand collecting on wood; IBE FS UPJS.

Paratypes
GEORGIA – Imereti, Tskaltubo • 3 ♀♀; Khomuli Cave; 42.31562° N, 42.63613° E; 11 Apr. 2020; Eter
Maghradze leg.; hand collecting on wood, guano, water surface • 1 ♀, 3 ♂♂; Melouri Cave; 42.38752° N,
42.62819° E; 28 May 2019; Eter Maghradze leg.; pitfall traps with pork liver, hand collecting on guano
and speleothems; IBE FS UPJS • 3 ♀♀; Prometheus Cave; 42.37716° N, 42.60086° E; 13 Feb. 2018; Eter
Maghradze leg.; pitfall traps with pork liver, hand collecting on wood, guano and speleothems; IBE FS
UPJS • 1 ♀; Datvi Cave; 42.37444° N, 42.59583° E; 27 Dec. 2019; Eter Maghradze leg.; pitfall traps with
pork liver; IZISU • 1 ♀; same collection data as preceding; IBE FS UPJS • 2 ♀♀; Satsurblia Cave; same
collection data as for holotype; IBE FS UPJS • 1 ♂; same collection data as preceding; IZISU • 1 ♀, 1 ♂;
Sakadzhia Cave; 42.36756387° N, 42.59123348° E; 28 Dec. 2020; Eter Maghradze leg.; hand collecting
on guano and detritus • 3 ♀♀, 3 ♂♂; Imereti, Khoni, Satevzia Cave; 42.43153377° N, 42.56590444° E;
10 Feb. 2020; Eter Maghradze leg.; hand collecting on guano and water surface; • 10 ♀♀, 2 ♂♂; same
collection data as preceding; IZISU IBE FS UPJS • 1 ♀, 1 ♂; Imereti, Chiatura, Shvilobisa Cave; 42.3254°
N, 43.26786° E; 8 Oct. 2021; Eter Magradze, Shalva Barjadze, Lado Shavadze, Mariam Gogshelidze leg.;
hand collecting on guano, wood and detritus; IBE FS UPJS • 3 ♀♀; Samegrelo, Martvili, Inchkhuri Cave;
42.45678637°N, 42.40425674°E; 18 Jul. 2020; 10 Jul. 2021; Eter Maghradze, Shalva Barjadze, Lado
Shavadze, Mariam Gogshelidze leg.; hand collecting guano, wood, water surface and walls; IZISU • 5 ♀♀;
same collection data as preceding; IBE FS UPJS • 1 ♀; Motena Cave; 42.47657295° N, 42.39126228° E;
10 Jul. 2021; Shalva Barjadze, Lado Shavadze, Mariam Gogshelidze leg.; hand collecting on walls; IBE
FS UPJS .

Description
Body length 1.3–2.3 mm in females, 1.85–2.1 in males (average 1.78 mm; n = 46), shape cylindrical
(Fig. 4a). Colour white to pale brownish in ethyl alcohol. Cuticular granulation fine and uniform, slightly
dense around pseudocelli. Antennae almost as long as head, area antennalis relatively well marked. PAO

European Journal of Taxonomy 879: 64–82 (2023)

https://zoobank.org/urn:lsid:zoobank.org:act:AAC4848A-79D3-4A5C-8458-A8CB7BB2EA8F

Fig. 4. Deuteraphorura colchisi Parimuchová, Barjadze & Kováč sp. nov. a. Dorsal chaetotaxy (the
same scale as in Fig. 4b). b. Ventral chaetotaxy of abdomen. c. PAO. d. MVO in adult specimen (other
than in Fig. 4b).

PARIMUCHOVÁ A. et al., Two new cave Deuteraphorura species from Georgia

with 10–14 compound vesicles (Fig. 4c). Ant. I with 8–9 chaetae in one row, Ant. II with 14–15 chaetae.
AOIII with 5 papillae, 5 guard chaetae, 2 sensory rods almost as long as papillae, 2 rough sensory
clubs and lateral ms (as in Fig. 5b). Lateral ms on Ant. IV placed basally at the level of second row of
chaetae. Apical organite simple in unprotected cavity. Maxillary outer lobe simple with 1 basal chaeta
and 2 sublobal hairs. Labium of AB-type, with 6 proximal chaetae. Basomedian field with 4 chaetae,
basolateral field with 5 chaetae. Head ventrally with 4 postlabial chaetae.

Pso formula dorsally as 33/133/3(4)3(4)4(3)5(6)3-4 (Fig. 4a) (2 pso on Th. I sometimes appear); ventrally
as 12/011/3212 (Fig. 4b for abdominal ventral pso formula); head ventrally with 1 anterior 1 postero-
medial and 1 postero-lateral pso. Psx weakly visible. Subcoxae 1 of I–III pairs leg with 2,2,2 pso.

Dorsal body chaetae only weakly differentiated into macro and mesochaetae. Th. I with 7 chaetae per
half. ThII–AbdIII with 3 + 3 medial chaetae respectively. VT with 5–7 chaetae per half, basal chaetae

Table 2. List of species with 3 pso on hind margin of the head and 1 pso on Th. I. Abbreviations: abs =
absent; f = forked; l = long; MVO = number of setae in male ventral organ; s = simple; t = thick.

Species Distribution Habitat Body length
(mm)

PAO Dorsal pso Ventral
pso

Subcoxae
1 pso

MVO

D. akelaris
Jordana &
Beruete, 1983

Spain
(Navarra)

cave 1.2 12–14 33/133/45454 3/011/
2211

? 2t/8t

D. arminiaria
(Gisin, 1961)

Austria cave 1.5–2.2 12 33/133/33354 3/011/
2112

2,2,2 ? 4t/6t

D. bizkaiensis
Beruete, Arbea &
Jordana in Beruete
et al. 2021

Spain
(Basque)

cave 0.8–1.06 12–13 33/133/33353 3/011/
3111

2,2,2 2t/8t

D. closanica
Gruia, 1965

Romania caves 1.25–1.8 12 33-
4/133/33353

3/011/41-
21-22

1,1,1 4s/35-40s

D. dashtenazensis
Arbea,
Yahyapour &
Shayanmehr in
Yahyapour et al.
2020

Iran soil,
litter

1.4–1.9 13-15 33/133/33353 3/000/
1221

2,2,2 -/8t

D. galani Beruete,
Arbea & Jordana,
2001

Spain
(Navarra)

cave 1.0–1.2 12–14 33/133/3-443-
454

3/011/
4212

? 2t/8t

D. harrobiensis
Beruete, Arbea &
Jordana, 2001

Spain
(Navarra)

cave 1.1.–1.3 12–15 33/133/3-
44464

3/011/
4111

? 2t/8t

D. jitkae
(Rusek, 1964)

Slovakia soil,
forest

2.1 21 33/133/33342 1/???/2-
321-22

1,1,1 2/
numerous
s,f

D. kosarovi
(Zonev, 1973)

? ? ? 15 33/133/33354 3/011/
4222

? ?

D. trisilvaria
(Gisin, 1962)

Austria cave 1.6–2.4 18 33/133/33354 3/011/
3211

2,2,2 abs

D. colchisi
sp. nov.

Georgia cave 1.1–2.3 10–14 33/133/3(4)
3(4)4(3)5(6)3-4

3/011/
3212

2,2,2 -/20-25
t, s

D. kozmani
sp. nov

Georgia cave 1.5–2.6 14–16 33/133/4(3)4
(5)3-45(6)3(4)

3/011/
3222

2,2,2 -/ 20-25
l, t, f

European Journal of Taxonomy 879: 64–82 (2023)

Fig. 5. Deuteraphorura kozmani Parimuchová, Barjadze & Kováč sp. nov. a. Dorsal chaetotaxy.
b. AOIII. c. MVO (enlargment of modified chaeta). d. Tita and claw of leg III (DIC contrast image;
chaeta in C-row not visible from this view).

PARIMUCHOVÁ A. et al., Two new cave Deuteraphorura species from Georgia

absent. Chaetae on Th. I–III sterna absent. Furca remnant with 2 + 2 thin chaetae in one row. MVO
present only in fully adult males in form of 10–20 thickened, short and bent spine-like chaetae only on
Abd. III sternum (Fig. 4d). Subcoxae 1 of legs I–III 4,4,4 chaetae, subcoxae 2 with 3, 13–14, 14–16
chaetae, trochanters with 8–9 chaetae each and femora with 15, 13–15, 13–15 chaetae, respectively.
Tita I–III with 18, 19, 17 chaetae including 9 chaetae in distal whorl. Tita I with 6 + M chaetae in B row
and 2 chaetae in C row, Tita II with 7 + M chaetae in B-row and 2 chaetae in C-row, Tita III with 6 +
M chaetae in B row and 1 chaeta in C row. Claw without teeth. Empodium with basal lamella, tip of
filament reaching two-thirds of the claw length (as in Fig. 5d).

Ecology and distribution

The species is known only from caves in western Georgia where it inhabits warm caves at low altitudes.
By its morphology, it does not display any obvious troglomorphic adaptations.

Remarks

See remarks for D. kozmani sp. nov.

Deuteraphorura kozmani Parimuchová, Barjadze & Kováč sp. nov.
urn:lsid:zoobank.org:act:BD48C182-8833-4537-9C44-024938DC47CD

Fig. 5, Table 2

Etymology

The species was named after the type locality, the Kozmani Cave in Georgia.

Type material

Holotype
GEORGIA • ♂; Imereti, Kharagauli, Kozmani Cave; 42.10092528° N, 43.28852625° E; 14 Sept. 2021;
Eter Maghradze leg.; hand collecting on detritus; IBE FS UPJS.

Paratypes
GEORGIA • 6 ♀♀, 2 ♂♂; same collection data as for holotype; IZISU • 8 ♀♀, 1 ♂; same collection
data as for holotype; IBE FS UPJS.

Description

Body length 1.8–2.6 mm in females, 1.5–2.0 in males (average 2.0 mm; n = 18), shape cylindrical
(Fig. 5a). Colour white to pale brownish in ethyl alcohol. Cuticular granulation fine and uniform, slightly
dense around pseudocelli. Antennae almost as long as head, area antennalis relatively well marked. PAO
with 14–16 compound vesicles. Ant. I with 8 chaetae in one row, Ant. II with 14–15 chaetae. AOIII with
5 papillae, 5 guard chaetae, 2 sensory rods almost as long as papillae, 2 rough sensory clubs and lateral
ms (Fig. 5b). Lateral ms on Ant. IV placed basally at the level of second row of chaetae. Apical organite
simple in unprotected cavity. Maxillary outer lobe simple with 1 basal chaeta and 2 sublobal hairs.
Labium of AB-type, with 6 proximal chaetae. Basomedian field with 4 chaetae, basolateral field with 5
chaetae. Head ventrally with 5 postlabial chaetae.

Pso formula dorsally as 33/133/4(3)4(5)3-45(6)3(4) (Fig. 5a); ventrally as 12/011/3222; head ventrally
with 1 anterior, 1 postero-medial and 1 postero-lateral pso. Psx weakly visible. Subcoxae 1 of I–III pairs
leg with 2,2,2 pso.

European Journal of Taxonomy 879: 64–82 (2023)

https://zoobank.org/urn:lsid:zoobank.org:act:BD48C182-8833-4537-9C44-024938DC47CD

Dorsal body chaetae only weakly differentiated into macro and mesochaetae. Th. I with 6–7 chaetae per
half. ThII–AbdIII with 3 + 3 medial chaetae respectively. VT with 5–6 chaetae per half, basal chaetae
mostly absent. Chaetae on Th. I–III sterna absent. Furca remnant with 2 + 2 thin chaetae in one row.

MVO present only in fully adult males in form of 20–25 thickened, long and forked chaetae only on
Abd. III sternum (Fig. 5c). Subcoxae 1 of legs I–III with 4, 4, 4 chaetae, subcoxae 2 with 3,14–17, 15–17
chaetae, trochanters with 8–10 chaetae each and femora with 14–15, 13–15, 13–15 chaetae, respectively.
Tita I–III with 18, 19, 17 chaetae including 9 chaetae in distal whorl. Tita I with 6 + M chaetae in B row
and 2 chaetae in C row, Tita II with 7 + M chaetae in B-row and 2 chaetae in C-row, Tita III with 6 +
M chaetae in B row and 1 chaeta in C row. Claw without teeth. Empodium with basal lamella, tip of
filament reaching two-thirds of the claw length (Fig. 5d).

Ecology and distribution
The species is known only from the type locality, occurring on guano and decaying organic material. It
does not display any obvious troglomorphic adaptations.

Remarks
Both species belong to the species group of Deuteraphorura with 3 pso on hind margin of the head
and possessing the pso on the first thoracic tergum. The vast majority of these species occupy caves
in southern and central Europe. As morphological characters vary within both new species, reliable
distinguishing from each other is possible only by ventral pseudocellar formula and shape of MVO in
matured males. Deuteraphoruracolchisi sp. nov. has simple thickened chaetae in MVO, while modified
chaetae in D.kozmani sp. nov. are longer and weakly forked at the tip. Similar to the new species,
D. dashtenazensis Arbea, Yahyapour & Shayanmehr, 2020 has MVO only on Abd. III, but it differs in
number of chaetae on this organ. Diagnostic morphological characters of both new species and other
species of this group are listed in Table 2.

Discussion
Only a few species of Deuteraphorura have been registered in Georgia to date: D. variabilis (Stach,
1954), D. kruberaensis Jordana & Baquero, 2012 (Barjadze et al. 2012, 2015) and D. inermis (Tullberg,
1869); however, the occurrence of the last species in Georgian caves is doubtful (Barjadze et al. 2012).
Intensive cave sampling using an integrative approach reveals a greater diversity of cave Deuteraphorura
in Georgia than previously thought.

Delimitation methods indicated the presence of several distinct molecular lineages (MOTUs) within
the (morpho)species colchisi, while only one in kozmani. Three groups within the species of colchisi
point to recent speciation of Deuteraphorura in Georgian caves, as revealed in Deuteraphorura and
Protaphorura from the Western Carpathians (Parimuchová et al. 2017, 2020). Three approaches to species
delimitation (morphological, molecular and geographical) contradict one another in the population from
Shvilobisa Cave. Molecularly, specimens from this cave represent unique species; however, it is located
in the same karst area as Kozmani Cave–Zemo-Imereti Plateau. Geography was considered a reliable
delimitation tool in Onychiuridae (Sun et al. 2017), but the in the case of such a complex karst area as
Zemo-Imereti, particular structural plateaus have a different geological history (Lezhava et al. 2019;
Tielidze et al. 2019), which may have a decisive impact on the isolation of subterranean populations
within this area and the evolution of independent phyletic lineages of Deuteraphorura. Morphologically,
specimens from Shvilobisa Cave are similar to those from the geologically similar Sataplia-Tskaltubo
karst area. But due to the relatively small number of specimens for morphological and molecular study,
the population from Shvilobisa Cave needs further examination. The Satevzia, Motena and Inchkhuri
caves are located in different karst areas, but they are geographically relatively close; they share the

PARIMUCHOVÁ A. et al., Two new cave Deuteraphorura species from Georgia

same cryptic species, which is documented by a positive correlation between geographic and genetic
distance.

As the family Onychiuridae reflects a high level of morphological variability and the left-right
asymmetry in chaetotaxy and pseudocellar patterns (e.g., Jordana et al. 2012; Kaprus' et al. 2014; Sun &
Wu 2014; Parimuchová et al. 2017, 2020; Vargovitsh 2019), identification of species-specific characters,
corresponding to molecular delimitation, is very problematic. Inadequacies in the morphological
taxonomy of Onychiuridae are caused by a lack of sufficient morphological characters and thus a high
level of cryptic diversity in this family (Sun et al. 2017).

Subterranean biodiversity of the Caucasus Mts – a hotspot area
Areas of the highest subterranean biodiversity (hotspots) were defined based on the number of species
adapted to subterranean life per cave (Culver & Sket 2000; Reboleira et al. 2011). Regarding the number
of troglobiotic species, we have to return to the definition of troglobiont/troglobite as a species that
exclusively inhabits a subterranean environment with a preference for its deep parts, and eventually
also showing morphological adaptations to subterranean life (Sket 2008; Trajano & Carvalho 2017;
Howarth & Moldovan 2018). The degree of troglomorphism is not correlated with occupied cave depths,
as documented by non-adapted animals occurring in the deepest parts of caves (Sendra & Reboleira
2012). To distinguish troglobionts based only on the level of morphological adaptations to the cave
environment is very ambiguous, particularly in pre-adapted groups of invertebrates living in deeper
soil horizons. Moreover, recent speciation could be a reason for the low development of troglomorphic
characters in Onychiuridae (e.g., Fiera et al. 2021). Thus, the real species richness of a given area could
be underestimated in this family when considering only morphological traits.

It is known that troglomorphic adaptations are not universal in all cave-adapted species. The level of
troglomorphy could be correlated with evolutionary age, showing up well in ‛old’ troglobionts and
weakly to moderately in ‛young’ ones (Kováč et al. 2016). Guanobionts regularly contradict the generally
accepted morphological traits of obligate cave-dwellers (e.g., Culver & Pipan 2009, 2015). In troglobiotic
Onychiuridae, a rather edaphomorphic appearance without progressive troglomorphic adaptations has
been documented in a large number of cave-dwelling species from the Romanian Carpathians (Fiera et
al. 2021). On the other hand, the highly troglomorphic Troglaphorura gladiator Vargovitsh, 2019, from
Georgia in the Caucasus, and Deuteraphorura muranensis Parimuchová & Kováč, 2020, distributed
at the northernmost distribution limit of troglobionts in Europe (Parimuchová et al. 2020), show an
extremely high level of troglomorphy, as much as the species of the genus Ongulonychiurus from Spain
and Croatia, respectively (Thibaud & Massoud 1986; Sun et al. 2019), and Pilonychiurus from Algeria
(Pomorski 2007). In contrast, Absolonia gigantea (Absolon, 1901) from Dinarides, of an unusually
large size, lacks distinct troglomorphy similar to Protaphorura janosik Weiner, 1990, and P. cykini
Parimuchová & Kováč in Parimuchová et al., 2017a, from the Western Carpathians and Siberia,
respectively, and Megaphorura arctica (Tullberg, 1877), which is abundant in the substrate at the foot
of bird cliffs in the Arctic. These discrepancies suggest that the microhabitat (or the trophic niche that a
species occupies) determines the level of troglomorphy to a greater extent than geographic distribution
in a biodiversity hotspot or evolutionary origin in terms of young and old troglobionts. Based on the
category of troglomorphisms (length of antennae, claws and furca), four life forms were distinguished in
troglobiotic Arrhopalitidae, including neustonic, atmobiotic, intermediate and intrasubstrate troglobionts
(Vargovitsh 2022). A similar approach could be applied to representatives of the family of Onychiuridae,
whose species living exclusively on the water surface have a clearly elongated claw, while species living
in guano and sediments generally have a short claw.

The occurrence of highly troglomorphic species is thus possible everywhere in the subterranean
environment where the microhabitat character requires specific adaptations; however, there are

European Journal of Taxonomy 879: 64–82 (2023)

environmental indicators to a higher incidence. Aside from hypotheses considering habitat heterogeneity,
historical circumstances and habitat productivity, a high terrestrial species richness is also enhanced by
the west-east orientation of mountains, which historically reduced the migration potential of invertebrates
and increased their invasion rate via subterranean habitats (Culver et al. 2006; Deharveng et al. 2012).
Similar to the geographic characteristics seen in mountain ranges in southern Europe, the Caucasus is
predestined to be a hotspot of subterranean biodiversity and potential evolution centre. The relatively
large extent and connectivity of the karst, especially in Abkhazia, Georgia, may enable subterranean
species to disperse more widely and access various microhabitats inside the karst, the pattern revealed in
Dinarides (Bregović & Zagmajster 2016). The length of passages and surface productivity as a variable
of subterranean species richness (Culver et al. 2004, 2006) point in favour of the Caucasus as a hotspot,
as well. The long-term stable areas of high precipitation are important for subterranean terrestrial
diversity, while productive energy is important only on a global scale (Bregović & Zagmajster 2016).

The fauna of the Western Caucasian caves importantly contributes to global subterranean diversity, as
was documented across multiple arthropod taxa (e.g., Sendra & Reboleira 2012; Antić & Makarov 2016;
Barjadze et al. 2019; Antić & Reip 2020; Martens et al. 2021; Zaragoza et al. 2021). The Caucasus
as a significant hotspot of subterranean biodiversity is well documented in Diplopoda, with a high
level of endemism and a high proportion of troglobiotic species, especially in the orders Julida and
Chordeumatida (Antić & Makarov 2016; Antić & Reip 2020). Altogether, the 19 troglobiotic species
known from a single cave in Georgia (Fiera et al. 2021) are almost equal to the threshold of 20 troglobiotic
and stygobiotic species in the hotspot caves (Culver & Sket 2000). Thus, it is herein documented that the
Western Caucasus is a centre of speciation in the Collembola genera of Arrhopalites, Pygmarrhopalietes
(Arrhopalitidae) and Plutomurus (Tomoceridae) (Fiera et al. 2021), and based on the present results,
potentially also in the genus Deuteraphorura (Onychiuridae).

Acknowledgements
This article is published in the frame of the grant: “Complex morphological and molecular investigations
of cave dwelling collembolans (Hexapoda) in Georgia and Slovakia” supported by the National
Scholarship Programme of the Slovak Republic. The study was financially supported by grant APVV-
21-0379 (Slovak Research and Development Agency), grant VEGA 1/0438/22 (Slovak Scientific Grant
Agency) and grant “Conservation actions and invertebrates’ investigations in Sataplia-Tskaltubo karst
caves, Georgia” supported by the Conservation Leadership Programme (CLP-04125220). We would like
to thank speleologist Valeri Barbakadze (Imereti Cave Protected Areas, Georgia) for assistance during
the expeditions. We are thankful to David Lee McLean for linguistic correction of the manuscript.

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Manuscript received: 21 October 2022
Manuscript accepted: 13 February 2023
Published on: 10 July 2023
Topic editor: Tony Robillard
Section editor: Javier I. Arbea
Desk editor: Marianne Salaün

Printed versions of all papers are also deposited in the libraries of the institutes that are members of the
EJT consortium: Muséum national d’histoire naturelle, Paris, France; Meise Botanic Garden, Belgium;
Royal Museum for Central Africa, Tervuren, Belgium; Royal Belgian Institute of Natural Sciences,
Brussels, Belgium; Natural History Museum of Denmark, Copenhagen, Denmark; Naturalis Biodiversity
Center, Leiden, the Netherlands; Museo Nacional de Ciencias Naturales-CSIC, Madrid, Spain; Leibniz
Institute for the Analysis of Biodiversity Change, Bonn – Hamburg, Germany; National Museum of the
Czech Republic, Prague, Czech Republic.

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