Citation
========

Zawierucha K, Dziamięcki J, Jakubowska N, Michalczyk Ł, Kaczmarek Ł (2014) New tardigrade records for the Baltic states with a description of*Minibiotus formosus* sp. n. (Eutardigrada, Macrobiotidae). ZooKeys 408: 81--105. doi: [10.3897/zookeys.408.6612](http://dx.doi.org/10.3897/zookeys.408.6612)

Introduction
============

The Baltic States, i.e. Estonia, Latvia and Lithuania, are located on the eastern coast of the Baltic Sea, and fall within the Palearctic ecozone ([@B27]). The topography of the three countries is dominated by lowlands with the highest peaks at *ca.* 300 m asl. The temperate climate is intermediate between maritime and continental. Even though the phylum Tardigrada is cosmopolitan and currently comprises *ca.* 1,200 species ([@B16]), so far only six tardigrade taxa have been reported from the Baltic States. Specifically, two from Estonia: *Eremobiotus alicatai* (Binda, 1969) and *Isohypsibius* cf. *marcellinoi* (Binda & Pilato, 1971), two from Latvia: *Paramacrobiotus richtersi* group and *Macrobiotus hufelandi* group and two from Lithuania: *Macrobiotus* sp. and *Ramazzottius* sp. ([@B79], [@B84], [@B85]).

In this study we report seventeen tardigrade species, which are all new records for the Baltic States. Moreover, one of these species is also new to science. The new species belongs to the genus *Minibiotus* R.O. Schuster, 1980, that until 1988 contained only a single species, *Minibiotus intermedius* (Plate, 1888). In 1988 [@B63] described *Minibiotus maculartus*, and within the last decade fourteen new *Minibiotus* species have been described. Several species have also been transferred to *Minibiotus* from the genus *Macrobiotus* based on characters defined by [@B10] and later supplemented by [@B23] ([@B46], [@B68], [@B48], [@B26], [@B52], [@B65], [@B38], [@B21], [@B22], [@B78], [@B44], [@B43], [@B45] [@B42]). Currently, the total number of *Minibiotus* species amounts to as many as forty seven.

In addition to the description of the new species, we also provide morphometric data and photographs of *Hypsibius* cf. *scabropygus*, a rare species that belongs to a large group of hypsibiids with at least partially sculptured dorsal cuticle and pharynx with two macroplacoids and without the microplacoid.

Material and methods
====================

Sixteen moss, lichen and mixed (moss/lichen) samples from trees, soil and stones were collected from 15 localities in Estonia, Latvia and Lithuania between the 29 April and the 5 May 2012 by the third author (more details below). Samples were collected and examined for tardigrades using standard methods (see [@B11]). After extraction, animals were mounted on microscope slides in Hoyer's medium. All specimens were examined measured and photographed using Phase Contrast Microscopy (PCM) or Scanning Electron Microscopy (SEM). In total 358 specimens (including 47 simplexes), one exuvium with 6 eggs, and 8 free-laid eggs were examined.

All measurements are given in micrometers \[μm\]. Structures were measured only if their orientation was appropriate. Body length was measured from the anterior extremity to the end of the body, excluding the hind legs. Buccal tube length and the level of the stylet support insertion point were measured according to [@B58]. Buccal tube width was measured as the external diameter at the level of the stylet support insertion point. Lengths of the claw branches were measured from the base of the claw to the top of the branch including accessory points for *Minibiotus* and according to [@B3] for *Hypsibius*. The *pt* ratio is the ratio of the length of a given structure to the length of the buccal tube expressed as a percentage ([@B58]). Macroplacoid length sequence is given according to [@B35], i.e. macroplacoids are listed from the shortest to the longest and their relative sizes are denoted with appropriate inequality, approximation and/or equality signs (\<, ≤, ≈, =). Morphometric data were handled using the 'Macrobiotoidea' ver. 1.1 template available from the Tardigrada Register ([www.tardigrada.net/register](http://www.tardigrada.net/register), [@B51]).

For species identification and differentiation, keys in [@B10], [@B19], [@B36], [@B54], [@B55] and [@B77], and original descriptions and redescriptions ([@B18], [@B75], [@B76], [@B28], [@B4], [@B12], [@B13], [@B6], [@B48], [@B52], [@B56], [@B21], [@B22], [@B44], [@B45]) as well as for insertion of the stylet muscles [@B60] were used. Tardigrade taxonomy is presented according to [@B40]. Only specimens determined to species level are provided in the list of species (we omitted all specimens determined only to the species group level, e.g. the *hufelandi* group or the *oberhaeuseri* group). In the species list Roman numbers indicate sample codes (see sampling localities) and Arabic numbers indicate the number of specimens, exuvia/simplexes and eggs.

Raw data underlying the description of *Minibiotus formosus* sp. n. are deposited in the Tardigrada Register ([@B51]) under [www.tardigrada.net/register/0012.htm](http://www.tardigrada.net/register/0012.htm).

Sampling localities
-------------------

I.  56°03\'08\"N, 24°24\'10\"E, *ca.* 33 m asl: Lithuania, Panevėžys county, Pasvalys district municipality, along the road E67, 0.5 km before the turning to Pasvalys, moss from tree and soil (slide code: LT 2422), date: 29.04.2012.

II. 55°25\'59\"N, 24°13\'32\"E, *ca.* 59 m asl: Lithuania, Kaunas county, Kėdainiai district municipality, Truskava city, near a church, lichens from tree and moss from soil (slide code: LT 2423), date: 29.04.2012.

III. 55°17\'12\"N, 23°58\'57\"E, *ca.* 30 m asl: Lithuania, Kaunas county, Kėdainiai district municipality, Kėdainiai city, Kranto II street; moss from wall (slide code: LT 2424), date: 29.04.2012.

IV. 55°17\'13\"N, 23°58\'56\"E, *ca.* 30 m asl: Lithuania, Kaunas county, Kėdainiai district municipality, Kėdainiai city, Paeismilgio street; moss from stone (slide code: LT 2425), date: 29.04.2012.

V.  55°43\'35\"N, 24°21\'30\"E, *ca.* 62 m asl: Lithuania, Panevėžys county, Panevėžys district municipality, Panevėžys city, Garden Street near Holy Trinity Rector; moss from tree (slide code: LT 2440), date: 05.05.2012.

VI. 56°38\'53\"N, 23°43\'18\"E, *ca.* 7 m asl: Latvia, Zemgale region, Jelgava municipality, Jelgava city, City Park; moss from soil (slide code: ŁO 2426), date: 29.04.2012.

VII. 57°10\'33\"N, 24°50\'32\"E, *ca.* 45 m asl: Latvia, Vidzeme region, Sigulda municipality, Gutmana Cave in the Gauja National Park; moss from rocks (slide code: ŁO 2427), date: 30.04.2012.

VIII. 56°23\'55\"N, 24°07\'33\"E, *ca.* 25 m asl: Latvia, Zemgale region, Bauska municipality, along Road No P103, 0.5 km from Saulaine; lichens from tree (slide code: ŁO 2428) date: 29.04.2012.

IX. 57°09\'55\"N, 24°51\'03\"E, *ca.* 73 m asl: Latvia, Vidzeme region, Sigulda municipality, Turaida city, Turaida Castle; moss from stone (slide code: ŁO 2430), date: 30.04.2012.

X.  56°54\'32\"N, 24°08\'45\"E, *ca.* 10 m asl: Latvia, Riga Region, boundary of Ķekava municipality, along road no A2; moss from tree (slide code: ŁO 2431), date: 30.04.2012.

XI. 57°09\'59\"N, 24°50\'59\"E, *ca.* 91 m asl: Latvia, Vidzeme region, Sigulda municipality, Sigulda city, Sigulda Castle; moss from stone (slide code: ŁO 2432), date: 30.04.2012.

XII. 56°41\'22\"N, 23°47\'43\"E, *ca.* 4 m asl: Latvia, Zemgale region, Ozolnieki municipality, Ozolnieki city, about 100 m from the Ozolnieki Lake; moss from soil (slide code: ŁO 2433), date: 29.04.2012.

XIII. 57°45\'43\"N, 24°20\'59\"E, *ca.* 3 m asl: Latvia, Vidzeme region, Salacgriva municipality, Salacgriva city; moss from soil, near the beach (slide code: ŁO 2434), date: 01.05.2012.

XIV. 59°10\'44\"N, 24°30\'06\"E, *ca.* 59 m asl: Republic of Estonia, Harju county, Kernu Parish municipality, Road No 4, moss from tree (slide code: ES 2420), date: 04.05.2012.

XV. 59°10\'44\"N, 24°30\'06\"E, *ca.* 59 m asl: Republic of Estonia, Harju county, Kernu Parish municipality, Road No 4, moss from tree (slide code: ES 2421), date: 04.05.2012.

XVI. 58°48\'47\"N, 24°24\'46\"E, *ca.* 32 m asl: Republic of Estonia, Rapla County, Märjamaa municipality, forest near Konuvere village, moss from tree (slide codes: ES 2487), date: 29.04.2012.

Results
=======

Taxonomic accounts of species found in the study
------------------------------------------------

### Phylum: Tardigrada (Spallanzani, 1777)

#### Class: Eutardigrada Richters, 1926

##### Order: Apochela Schuster, Nelson, Grigarick and Christenberry, 1980

###### Family: Milnesiidae Ramazzotti, 1962

####### Genus: *Milnesium* Doyère, 1840

######## Milnesium asiaticum

Tumanov, 2006

######### Localities and specimen numbers.

XV: 1 specimen.

######### Remarks.

Our specimen corresponds perfectly to the original description. *Milnesium asiaticum* was originally described from Kirghizstan and subsequently found in the Svalbard archipelago ([@B83], [@B37]).

######## Milnesium tardigradum tardigradum

Doyère, 1840

######### Localities and specimen numbers.

VIII: 31 specimens (including 6 simplexes) + 1 exuvium with 6 eggs.

######### Remarks.

Specimens correspond perfectly with the redescription by [@B54], [@B55]). This species was reported from many localities throughout the World, however records prior to [@B54], [@B55]) need to be verified. So far, all confirmed localities are exclusively European ([@B54], [@B55]).

##### Order: Parachela Schuster, Nelson, Grigarick & Christensen, 1980

###### Superfamily: Hypsibioidea Pilato, 1969 (in [@B40])

####### Family: Hypsibiidae Pilato, 1969

######## Subfamily: Diphasconinae Dastych, 1992

######### Genus: *Diphascon* Plate, 1888

########## Subgenus: *Diphascon (Diphascon)* Pilato, 1987

########### Diphascon (Diphascon) bullatum

Murray, 1905

############ Localities and specimen numbers.

XIII: 1 specimen.

############ Remarks.

Specimens correspond well with the limited original description ([@B57]) and also with later descriptions ([@B1] and [@B11], [@B12]). This species is very similar to *Diphascon (Diphascon) patanei* (Binda & Pilato, 1971) and, as suggested by [@B12], these two species could be synonymous. Thus, to clarify the taxonomic status of *Diphascon (Diphascon) patanei*, a re-description of *Diphascon (Diphascon) bullatum* is necessary based on material from *locus typicus* in Scotland (the type material probably does not exist) ([@B12]).

########### Diphascon (Diphascon) pingue pingue

(Marcus, 1936)

############ Localities and specimen numbers.

XV: 11 specimens.

############ Remarks.

Although we have found only 11 specimens, we were confident in identifying them to *Diphascon (Diphascon) pingue* because they corresponded perfectly to the partial redescriptions by [@B61], [@B62]) and we also identified them with the key by [@B19]. The species belongs to the *pingue* group and has been previously recorded from numerous localities throughout the World, however the majority of records should be verified based on the modern taxonomy ([@B61], [@B62]). Currently, exclusively verified localities of this species are only from Europe and North America ([@B61]).

########### Diphascon (Diphascon) recamieri

Richters, 1911

############ Localities and specimen numbers.

XV: 1 specimen.

############ Remarks.

The species has previously been found in many localities, mostly in the Holarctic ([@B41]).

########### Diphascon (Diphascon) rugosum

(Bartoš, 1935)

############ Localities and specimen numbers.

II: 3 specimens.

############ Remarks.

The species has previously been found in many localities in the Holarctic ([@B41]).

######### Genus: *Diphascon* Pilato, 1987

########## Subgenus. Diphascon (Adropion) prorsirostre

Thulin, 1928

########### Localities and specimen numbers.

XIV: 2 specimens, XV: 2 specimens.

########### Remarks.

The species has been previously found in many localities, mostly in the Holarctic ([@B41]).

######## Subfamily: Hypsibiinae Pilato, 1969

######### Genus: *Hypsibius* Ehrenberg, 1848

########## Hypsibius convergens

(Urbanowicz, 1925)

########### Localities and specimen numbers.

IX: 6 specimens (including 2 simplexes).

########### Remarks.

Belonging to the cosmopolitan *convergens*-*dujardini* complex of species ([@B41], [@B56], [@B30], [@B34]), *Hypsibius convergens* used to be considered cosmopolitan, but it is most likely a complex of very similar (possibly also cryptic) species found throughout the world. The original *Hypsibius convergens* description no longer conforms to modern standards and therefore a redescription is required. Nevertheless, the examined specimens correspond perfectly with the original description and the *Hypsibius convergens* characteristics reviewed by [@B56].

########## Hypsibius dujardini

(Doyère, 1840)

########### Localities and specimen numbers.

XIV: 2 specimens (including 1 simplex), XV: 5 specimens.

########### Remarks.

*Hypsibius dujardini* belongs to the cosmopolitan *convergens*-*dujardini* complex of species ([@B41], [@B56], [@B30], [@B34]), and used to be considered cosmopolitan, but it is most likely a complex of similar (possibly cryptic) species found throughout the world. Due to the limited original description, *Hypsibius dujardini* needs a modern redescription. Nevertheless, the examined specimens correspond perfectly with the original description and the *Hypsibius dujardini* characteristics reviewed by [@B56].

########## Hypsibius cf. scabropygus

Cuénot, 1929

[Table 1](#T1){ref-type="table"} [, Figs 1](#F1){ref-type="fig"} [--7](#F2){ref-type="fig"}

########### Localities and specimen numbers.

XI: 1 simplex, XII: 1 specimen, XIV: 34 specimens (including 4 simplexes), XV: 24 specimens (including 6 simplexes).

########### Description

**(measurements in [Table 1](#T1){ref-type="table"}). Adults.** Body transparent/white (after preparation), eyes present in 6 of 15 (40%) specimens mounted in Hoyer's medium ([Fig. 1](#F1){ref-type="fig"}). Dorsal cuticle sculptured: from head to legs II without tubercles but clearly thickened, from legs II to the caudal end of the body (including legs IV) with irregular tubercles and platelets. Tubercles increasing in size from the anterior to the posterior part of the body, reaching maximum dimensions between legs III and IV, where tubercles sometimes merge and form irregular platelets ([Figs 2--5](#F1){ref-type="fig"}). Irregular tubercles 1.0--6.0 μm in diameter. Ventral cuticle smooth (i.e. without sculpturing). Gibbosities and cuticular pores absent.

![*Hypsibius* cf. *scabropygus* Cuénot, 1929: **1** habitus (dorso-lateral view) **2--4** caudo-dorsal cuticle with distinct sculpturing -- tubercles and tubercles merged into platelets **5** a single caudo-dorsal platelet. (**1--3:** PCM, **4--5:** SEM).](zookeys-408-081-g001){#F1}

###### 

Measurements and *pt* values of selected morphological structures of *Hypsibius* cf. *scabropygus* Cuénot, 1929 mounted in Hoyer's medium (N -- number of specimens/structures measured, RANGE refers to the smallest and the largest structure among all measured specimens; SD -- standard deviation).

  -------------------------------- ------ ------- ------ ------ -------- ---- -------- ------ -------- ----- --------
  CHARACTER                        N      RANGE   MEAN   SD                                                  
  µm                               *pt*   µm      *pt*   µm     *pt*                                         
  Body length                      14     183     --     293    *808*    --   *1132*   235    *949*    33    *97*
  Buccopharyngeal tube                                                                                       
  Buccal tube length               15     22.0    --     28.2            --            24.6   --       2.0   --
  Stylet support insertion point   15     12.0    --     15.9   *51.3*   --   *57.1*   13.5   *54.9*   1.1   *1.6*
  Buccal tube external width       15     1.5     --     2.0    *6.5*    --   *8.0*    1.8    *7.2*    0.2   *0.5*
  Buccal tube internal width       15     0.6     --     0.9    *2.1*    --   *3.4*    0.7    *2.7*    0.1   *0.4*
  Placoid lengths                                                                                            
  Macroplacoid 1                   15     1.8     --     3.2    *7.7*    --   *12.3*   2.4    *9.6*    0.4   *1.3*
  Macroplacoid 2                   15     1.7     --     2.7    *7.2*    --   *11.3*   2.2    *8.9*    0.3   *1.1*
  Macroplacoid row                 15     4.7     --     6.3    *19.1*   --   *26.5*   5.3    *21.7*   0.5   *1.8*
  Claw 1 lengths                                                                                             
  External base                    14     2.2     --     4.5    *9.1*    --   *17.2*   3.5    *14.1*   0.7   *2.1*
  External primary branch          14     4.0     --     8.8    *18.2*   --   *36.1*   6.9    *28.1*   1.5   *5.0*
  External secondary branch        14     2.3     --     5.9    *10.5*   --   *24.4*   4.4    *17.9*   1.0   *3.9*
  Internal base                    12     2.1     --     4.1    *9.3*    --   *15.5*   3.3    *13.4*   0.6   *1.8*
  Internal primary branch          12     3.8     --     5.9    *16.0*   --   *23.5*   4.8    *19.3*   0.6   *2.0*
  Internal secondary branch        12     2.4     --     4.3    *9.9*    --   *16.5*   3.3    *13.3*   0.6   *2.0*
  Claw 2 lengths                                                                                             
  External base                    11     3.0     --     5.2    *12.4*   --   *19.9*   4.1    *16.7*   0.7   *2.2*
  External primary branch          13     6.7     --     10.4   *29.7*   --   *43.7*   8.5    *34.7*   1.3   *4.4*
  External secondary branch        13     4.3     --     6.7    *19.0*   --   *27.2*   5.4    *21.9*   0.7   *2.6*
  Internal base                    10     2.4     --     4.5    *10.9*   --   *18.9*   3.6    *14.6*   0.7   *2.2*
  Internal primary branch          12     4.0     --     6.7    *17.7*   --   *27.2*   5.4    *22.0*   0.9   *2.9*
  Internal secondary branch        12     2.6     --     5.4    *11.8*   --   *22.0*   4.1    *16.7*   0.9   *3.0*
  Claw 3 lengths                                                                                             
  External base                    9      2.7     --     6.2    *11.9*   --   *23.8*   4.3    *17.3*   1.0   *3.5*
  External primary branch          9      7.2     --     10.4   *29.3*   --   *43.7*   8.8    *35.7*   1.1   *4.4*
  External secondary branch        9      3.6     --     6.5    *12.8*   --   *27.3*   5.2    *21.0*   1.0   *4.3*
  Internal base                    11     2.3     --     4.1    *10.5*   --   *17.2*   3.4    *13.9*   0.6   *1.9*
  Internal primary branch          13     3.8     --     6.5    *17.3*   --   *27.3*   5.4    *21.8*   0.9   *3.1*
  Internal secondary branch        12     2.7     --     6.1    *12.2*   --   *24.8*   3.9    *16.0*   0.9   *3.5*
  Claw 4 lengths                                                                                             
  Anterior base                    13     3.3     --     5.6    *12.8*   --   *20.1*   4.1    *16.8*   0.6   *2.2*
  Anterior primary branch          13     4.4     --     7.5    *19.5*   --   *31.1*   5.9    *24.2*   1.1   *3.9*
  Anterior secondary branch        11     3.1     --     13.2   *13.0*   --   *47.3*   4.8    *18.9*   2.9   *9.7*
  Posterior base                   12     2.7     --     5.4    *12.3*   --   *21.5*   4.5    *18.2*   0.9   *3.1*
  Posterior primary branch         12     4.9     --     14.9   *22.0*   --   *60.6*   10.3   *41.9*   2.9   *11.2*
  Posterior secondary branch       12     4.0     --     6.5    *15.4*   --   *25.6*   5.2    *21.2*   0.9   *3.4*
  -------------------------------- ------ ------- ------ ------ -------- ---- -------- ------ -------- ----- --------

Bucco-pharyngeal apparatus of the *Hypsibius* type, without the ventral lamina, and with forked apophyses for stylet muscles ([Fig. 6](#F2){ref-type="fig"}). Peribuccal lamellae absent. Teeth in the oral cavity armature absent or not visible under PCM. Pharyngeal bulb with apophyses and with two granular macroplacoids (both, without constrictions). Macroplacoid length sequence 2\<1. Microplacoid and septulum absent.

![*Hypsibius* cf. *scabropygus* Cuénot, 1929: **6** bucco-pharyngeal apparatus (dorso-ventral projection, ventral placoids in the insert) **7** claws IV (arrow indicates a small cuticular bar near the posterior claw). (Both PCM).](zookeys-408-081-g002){#F2}

Claws of the *Hypsibius* type, internal claws much smaller and of a different shape than the external claws ([Fig. 7](#F2){ref-type="fig"}). All main branches with large accessory points. Smooth, indistinct areoles under claws usually visible only on posterior claws IV. Cuticular bars under claws I-III absent but a small bar is present near the posterior claw IV ([Fig. 7](#F2){ref-type="fig"}, arrow).

**Eggs.** Unknown.

########### Remarks.

*Hypsibius scabropygus* has been recorded from many localities, mostly in the Holarctic ([@B41]). In general, our specimens correspond to the original description by [@B8], [@B9]) and later descriptions by [@B39] (=*Hypsibius callimerus* spec. nov.), and by [@B77]. However, importantly, none of the above mentioned descriptions reported a bar between anterior and posterior claws IV, which is present in all our specimens. Given the bar is small, it is possible that it was overlooked by Cuénot and later authors. If, however, *Hypsibius scabropygus* does not have the bar, then our specimens should probably be classified as a new species. Thus, until *Hypsibius scabropygus* is redescribed, our Latvian and Estonian records should be regarded as *Hypsibius* cf. *scabropygus*. As there is a possibility of our specimens belonging to a new species, we provide standard morphometrics ([Table 1](#T1){ref-type="table"}) and photographs ([Figs 1](#F1){ref-type="fig"}--[7](#F2){ref-type="fig"}).

######## Subfamily: Itaquasconinae Rudescu, 1964

######### Genus: *Astatumen* Pilato, 1997

########## Astatumen bartosi

(Węglarska, 1959)

########### Localities and specimen numbers.

XIV: 1 specimen.

########### Remarks.

Our specimen corresponds perfectly with characters of *Astatumen bartosi* proposed by [@B12] with the main difference between *Astatumen bartosi* and *Astatumen trinacriae* being the absence/presence of cuticular bars on legs II--III. Due to the notorious difficulties in differentiating the two species, the actual distribution of *Astatumen bartosi* cannot currently be described with confidence. [@B41] cited this species from several localities in Europe and from single African, Asian and South American sites.

###### Superfamily: Isohypsibioidea Marley, McInnes & Sands, 2011

####### Family: Isohypsibiidae Marley, McInnes & Sands, 2011

######## Genus: *Isohypsibius* Thulin, 1928

######### Isohypsibius ronsisvallei

Binda & Pilato, 1969

########## Localities and specimen numbers.

III: 1 specimen.

########## Remarks.

The species has previously been reported from several, mostly European, localities in the Holarctic ([@B41]).

######### Isohypsibius sattleri

(Richters, 1902)

########## Localities and specimen numbers.

IX: 1 specimen, XI: 8 specimens (including 5 simplexes), XIV: 3 specimens, XV: 5 specimens, XVI: 1 specimen.

########## Remarks.

The species has previously been reported from many localities throughout the World, thus it is considered cosmopolitan ([@B41], [@B34]).

###### Superfamily: Macrobiotoidea Thulin, 1928 in [@B40]

####### Family: Macrobiotidae Thulin, 1928

######## Genus: *Macrobiotus* C.A.S. Schultze, 1834

######### Macrobiotus harmsworthi harmsworthi

Murray, 1907

########## Localities and specimen numbers.

XI: 1 egg, XIV: 8 specimens, 1 egg.

########## Remarks.

The species belongs to the *harmsworthi* group which is widely distributed across a broad range of ecosystems throughout the world ([@B41], [@B34]). In the last decade many new species within this group were described from a variety of localities ([@B47], [@B69], [@B81], [@B64], [@B65], [@B70], [@B32], [@B31], [@B66], [@B78], see also [@B36] for the diagnostic key to the group). Due to many uncertain reports of *Macrobiotus harmsworthi harmsworthi*, especially in older literature, the distribution of the species is currently unknown. Specimens found in the present study correspond well to the characters presented in [@B72] and were successfully identified with the key by [@B36].

######### Macrobiotus hufelandi hufelandi

C.A.S. Schultze, 1833

########## Localities and specimen numbers.

XIV: 4 specimens, 1 egg.

########## Remarks.

The species belongs to the *hufelandi* group which is widely distributed across a broad range of ecosystems throughout the world ([@B41], [@B34]). In the last decade new species belonging to this group have been described from various localities ([@B68], [@B29], [@B14], [@B15], [@B20], [@B2], [@B31], [@B67], [@B5], [@B7], [@B74], [@B25], see also [@B4] for the diagnostic key to the group). Due to many uncertain reports of *Macrobiotus hufelandi hufelandi*, especially in older literature, the distribution of the species is currently unknown. Specimens found in the present study correspond well with the redescription by [@B4].

######## Genus: *Minibiotus* R.O. Schuster, 1980

######### Minibiotus formosus sp. n.

http://zoobank.org/BDBE49B7-84CF-4FE2-BE55-A399A537DE77

http://species-id.net/wiki/Minibiotus_formosus

http://www.tardigrada.net/register/0012.htm

[Tables 2](#T2){ref-type="table"} [--3](#T3){ref-type="table"} [, Figs 8](#F3){ref-type="fig"} [--15](#F4){ref-type="fig"}

########## Type material.

Holotype and 23 paratypes, 24 specimens (including 2 simplexes) and 3 unembryonated eggs).

########## Type locality.

57°10\'33\"N, 24°50\'32\"E, *ca.* 45 m asl: Latvia, Vidzeme region, Sigulda municipality, Gutmana Cave in the Gauja National Park; moss from rock (1 sample, slide codes: ŁO 2427/\*, where the asterisk can be substituted by any of the following numbers: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12).

########## Description

**(measurements in [Table 2](#T2){ref-type="table"}).** Body white/colourless ([Fig. 8](#F3){ref-type="fig"}). Eyes present in 18 of 24 (75%) specimens mounted in Hoyer's medium. Entire cuticle covered with small (0.4--1.1 μm) and large (1.9--2.5 μm) round or oval pores ([Fig. 9](#F3){ref-type="fig"}). Pores arranged in 9--10 poorly defined transverse bands. Pores on the dorsal cuticle arranged more densely than on the ventral cuticle. A single large pore (diameter: 2.1--2.9 μm) present on external side of legs I--III ([Fig. 12](#F4){ref-type="fig"}, arrow). A ring of pores around the mouth opening absent. Cuticle without granulation, except for legs which are all covered with fine and regular granulation (better developed on legs IV) visible only in larger specimens ([Fig. 12](#F4){ref-type="fig"}, arrowhead).

![*Minibiotus formosus* sp. n.: **8** habitus (holotype, ventral view) **9** dorsal cuticle with pores (holotype) **10--11** bucco-pharyngeal apparatus (**10** dorso-ventral projection, paratype **11** lateral view, paratype). All PCM.](zookeys-408-081-g003){#F3}

![*Minibiotus formosus* sp. n.: **12** leg II with claws, granulation (arrowhead) and a single large pore (arrow) (holotype) **13** claws IV (paratype) **14** egg (mid-section) **15** egg surface with processes. All PCM.](zookeys-408-081-g004){#F4}

###### 

Measurements and *pt* values of selected morphological structures of *Minibiotus formosus* sp. n. mounted in Hoyer's medium (N -- number of specimens/structures measured, RANGE refers to the smallest and the largest structure among all measured specimens; SD -- standard deviation, ? -- trait oriented unsuitably for measurement).

  -------------------------------- ------ ------- ------ ------ ---------- ---- ------ ------ ------ ----- ----- ------ ------
  CHARACTER                        N      RANGE   MEAN   SD     Holotype                                                
  µm                               *pt*   µm      *pt*   µm     *pt*       µm   *pt*                                    
  Body length                      7      113     --     236    636        --   1034   184    901    39    139   194    848
  Buccal tube                                                                                                           
  Length                           9      17.7    --     22.9              --          20.3   --     1.7   --    22.9   --
  Stylet support insertion point   9      9.5     --     12.2   49.5       --   56.2   10.9   53.8   0.9   2.0   12.2   53.3
  External width                   9      1.3     --     2.0    7.0        --   9.9    1.6    7.8    0.2   0.8   1.7    7.4
  Internal width                   9      0.5     --     0.7    2.4        --   3.4    0.6    2.8    0.1   0.3   0.7    3.1
  Placoid lengths                                                                                                       
  Macroplacoid 1                   9      1.3     --     1.9    6.9        --   8.3    1.5    7.6    0.2   0.4   1.8    7.9
  Macroplacoid 2                   9      1.2     --     1.7    6.8        --   8.3    1.5    7.5    0.2   0.4   1.7    7.4
  Macroplacoid 3                   9      1.4     --     2.2    7.4        --   9.6    1.7    8.2    0.3   0.6   1.9    8.3
  Microplacoid                     8      0.5     --     0.9    2.4        --   4.7    0.7    3.3    0.1   0.8   0.8    3.5
  Macroplacoid row                 9      4.5     --     6.8    24.3       --   33.7   5.5    26.9   0.7   3.0   5.9    25.8
  Placoid row                      8      5.2     --     7.9    27.1       --   39.1   6.4    31.0   0.9   3.9   6.7    29.3
  Claw 1 lengths                                                                                                        
  External primary branch          4      4.7     --     6.5    22.8       --   31.6   5.8    27.7   0.8   3.7   6.2    27.1
  External secondary branch        3      3.6     --     4.3    17.5       --   20.9   3.9    19.4   0.4   1.7   ?      ?
  Internal primary branch          6      4.7     --     6.6    24.3       --   30.4   5.7    27.4   0.7   2.4   6.3    27.5
  Internal secondary branch        5      3.1     --     4.6    15.0       --   20.4   4.0    18.7   0.5   2.3   4.6    20.1
  Claw 2 lengths                                                                                                        
  External primary branch          5      5.2     --     6.9    25.2       --   34.0   5.9    28.6   0.6   3.5   5.9    25.8
  External secondary branch        4      3.3     --     4.7    16.0       --   23.2   3.9    19.4   0.6   2.9   ?      ?
  Internal primary branch          5      5.2     --     6.4    27.1       --   31.9   6.0    29.7   0.5   2.3   6.2    27.1
  Internal secondary branch        3      3.5     --     4.1    18.5       --   20.9   3.9    19.9   0.3   1.3   ?      ?
  Claw 3 lengths                                                                                                        
  External primary branch          5      5.1     --     6.9    27.0       --   33.2   6.3    30.6   0.7   2.4   6.9    30.1
  External secondary branch        6      3.6     --     4.9    19.0       --   24.1   4.6    22.1   0.5   1.8   4.9    21.4
  Internal primary branch          7      5.1     --     6.6    25.7       --   32.5   5.9    29.2   0.6   2.5   6.4    27.9
  Internal secondary branch        4      4.1     --     4.6    20.9       --   22.8   4.3    21.6   0.2   0.8   ?      ?
  Claw 4 lengths                                                                                                        
  Anterior primary branch          5      6.0     --     8.0    31.6       --   39.6   7.0    34.2   0.8   3.5   ?      ?
  Anterior secondary branch        4      4.0     --     6.0    21.2       --   29.7   5.0    24.1   0.9   3.9   ?      ?
  Posterior primary branch         6      6.0     --     7.8    30.7       --   38.4   7.0    33.9   0.7   3.1   ?      ?
  Posterior secondary branch       5      3.9     --     5.5    20.6       --   27.1   4.8    23.8   0.6   2.6   ?      ?
  -------------------------------- ------ ------- ------ ------ ---------- ---- ------ ------ ------ ----- ----- ------ ------

Mouth antero-ventral. Ten peribuccal papulae present. Bucco-pharyngeal apparatus of the *Minibiotus* type ([Figs 10--11](#F3){ref-type="fig"}). Oral cavity armature absent or not visible under PCM. Buccal tube with a poorly visible ventral lamina and with an anterior and a posterior bend (both visible in lateral view only, [Fig. 11](#F3){ref-type="fig"}). Buccal tube walls thickened just below the stylet supports insertion point. Pharyngeal apophyses triangular, very near to the first macroplacoid. Three granular macroplacoids and a minute microplacoid present in the pharyngeal bulb. All macroplacoids of similar but not identical sizes, the macroplacoid length sequence: 2\<1\<3. Septulum absent.

Claws of the *Macrobiotus* type ([Figs 12--13](#F4){ref-type="fig"}). Primary branches of claws with thin, but obvious accessory points detaching at the apogee of the primary branch curve. Smooth lunules present on all legs, distinctly larger under external and posterior claws. Bars and other cuticular thickenings on legs absent.

**Eggs (measurements in [Table 3](#T3){ref-type="table"}).** White/transparent, laid freely ([Fig. 14](#F4){ref-type="fig"}). Spherical, without areolation. Processes in the shape of short, smooth, slightly flexible cones ([Fig. 15](#F4){ref-type="fig"}). Processes are distributed on the surface of the egg close one to another but never in contact. Surface between processes smooth under PCM ([Fig. 15](#F4){ref-type="fig"}).

###### 

Measurements of selected morphological structures of *Minibiotus formosus* **sp. n.** eggs mounted in Hoyer's medium.

  ---------------------------------------------- ---------- ---------- ----------
  CHARACTER                                      egg 1      egg 2      egg 3
  Diameter of egg without processes              45.7       44.1       ?
  Diameter of egg with processes                 55.6       55.1       ?
  Process height                                 4.5--5.2   4.8--5.2   4.6--5.3
  Process base width                             2.8--3.4   2.8--3.1   2.4--2.6
  Process base/height ratio                      57%--69%   54%--65%   47%--57%
  Distance between processes                     2.0--2.5   1.9--3.9   1.8--2.0
  Number of processes on the egg circumference   30         29         30
  ---------------------------------------------- ---------- ---------- ----------

########## Remarks.

Since ventral lamina is very poorly visible, the measurements of this structure are not included in [Table 2](#T2){ref-type="table"}. Three unembryonated eggs have been found alongside the described specimens. Given that *Minibiotus formosus* sp. n. was the only *Minibiotus* species in the sample and because no *Ramazzottius* Binda & Pilato, 1986 was found in the samples, we assumed that these eggs belong to the new species.

########## Etymology.

Given that we found the composition of small and large pores in the new species beautiful, we decided to name the animal after this impression (in Latin '*formosus*' means 'beautiful').

########## Type depositories.

Holotype 23 paratypes and 3 eggs are deposited in the Department of Animal Taxonomy and Ecology at the Adam Mickiewicz University (Poznań, Poland).

Differential diagnosis
----------------------

The new species is most similar to *Macrobiotus gumersindoi* Guil & Guidetti, 2005, but it differs from it by: the presence of two types of cuticular pores (small and large) in the new species *vs* pores of a uniform size in *Macrobiotus gumersindoi*, the absence of a triangular or a pentagonal arrangement of pores placed above a single large pore on legs, the presence of granulation on legs, a different macroplacoid length sequence (2\<1\<3 in the new species *vs* 1=2=3 in *Macrobiotus gumersindoi*), and by slightly larger macroplacoids (I: 1.3--1.9 μm; II: 1.2--1.7 μm III: 1.4--2.2 μm in the new species *vs* 1.0 μm in *Macrobiotus gumersindoi*).

Other species to which *Minibiotus formosus* sp. n. is similar by some characteristics of adult and/or egg morphology (e.g. pores in transverse bands, eggs with conical processes), include species listed below. The new species differs specifically from:

-   ***Macrobiotus bisoctus*** ([@B28]) by: the absence of trilobed and star-shaped pores (although their presence was not mentioned in the original description, they are clearly visible in Fig. 114 in [@B28], and by stylet supports inserted in a more anterior position (*pt=49.5--56.2* in the new species *vs pt≈60.3* in *Macrobiotus bisoctus* (according to [@B10])).

-   ***Macrobiotus eichhorni*** Michalczyk & Kaczmarek, 2004 by: a different arrangement of pores on the dorsal cuticle (9--10 transverse bands in the new species *vs* 8 bands in *Macrobiotus eichhorni*), the absence of star-shaped pores, the absence of four pores around the mouth opening, the presence of a single large pore on lateral sides of legs I-III, slightly shorter buccal tube (17.7--22.9 μm in the new species *vs* 24.7--34.2 μm in *Macrobiotus eichhorni*), stylet supports inserted in more anterior position (9.5--12.2 μm \[*pt=49.5--56.2*\] in the new species *vs* 16.2--23.8 μm \[*pt=65.4--70.6*\] in *Macrobiotus eichhorni*), a different macroplacoid sequence (2\<1\<3 in the new species *vs* 2\<3\<1 μm in *Macrobiotus eichhorni*), slightly shorter placoids, and by slightly smaller external claws I--IV (compare [Table 2](#T2){ref-type="table"} below and Table 1 in [@B48] for exact differences in dimensions of placoids and claws).

-   ***Macrobiotus furcatus*** (Ehrenberg, 1859) (according to [@B6]) by: the absence of tri- and quadrilobed cuticular pores, the presence of two types of cuticular pores (small and large in the new species *vs* uniformly small pores present in *Macrobiotus furcatus*), the presence of a single large pore on each of legs I--III, the presence of granulation on legs, the absence of the oral cavity armature, stylet supports inserted in a more anterior position (*pt=49.5*--*56.2* in the new species *vs pt≈68.4* in *Macrobiotus furcatus*), a different macroplacoid length sequence (2\<1\<3 in the new species *vs* 2\<3\<1 in *Macrobiotus furcatus*), and by egg processes without an obvious flexible portion (and never bifurcated).

-   ***Macrobiotus harrylewisi*** Meyer & Hinton, 2009 by: the absence of tri- and quadrilobed cuticular pores, the presence of two types of pores (small and large) over the entire cuticle in the new species *vs* small pores present only in the anterior part of the body and large pores present only in the posterior part of the body in *Macrobiotus harrylewisi*, the presence of a single large pore on each of legs I--III, stylet supports inserted in a more anterior position (*pt=49.5*--*56.2* in the new species *vs pt=61.4*--*67.6* in *Macrobiotus harrylewisi*), a different macroplacoid length sequence (2\<1\<3 in the new species *vs* 2≤3\<1 μm in *Macrobiotus harrylewisi*), a different shape of egg processes (short, single-tipped cones in the new species *vs* elongated, tapering cones with bulbous bases in *Macrobiotus harrylewisi*), a smaller diameter of eggs without and with processes (44.1--45.7 μm and 55.1--55.6 μm in the new species *vs* 66.1--80.0 μm and 78.2--101.9 μm in *Macrobiotus harrylewisi*), a slightly lower number of processes on egg circumference (29--30 in the new species *vs* 32--41 in *Macrobiotus harrylewisi*), and by smaller egg processes (4.5--5.3 μm in the new species *vs* 7.6--12.8 μm in *Macrobiotus harrylewisi*).

-   ***Macrobiotus jonesorum*** Meyer et al., 2011 by: the absence of trilobed and polygonal pores, the presence of two types of cuticular pores (small and large) in the new species *vs* small pores present only in the anterior part of the body, intermediate in size in the middle of the body and large pores in the posterior part of the body in *Macrobiotus jonesorum*), the presence of a single large pore on each of legs I-III, the presence of granulation on all legs, a slightly shorter buccal tube (17.7--22.9 μm in the new species *vs* 24.4--29.6 μm in *Macrobiotus jonesorum*), stylet supports inserted in a more anterior position (*pt=49.5*--*56.2* in the new species *vs pt=63.0*--*65.6* in *Macrobiotus jonesorum*), a slightly smaller external diameter of the buccal tube (1.3--2.0 μm \[*pt=7.0-9.9*\] in the new species *vs* 2.1--2.6 μm \[*pt=7.4*--*10.7*\] in *Macrobiotus jonesorum*), a different macroplacoid length sequence (2\<1\<3 in the new species *vs* 1\<2\<3 in *Macrobiotus jonesorum*), slightly shorter macroplacoids II and III (II: 1.2--1.7 μm \[*pt=6.8*--*8.6*\]; III: 1.4--2.2 μm \[*pt=7.4*--*9.6*\] in the new species *vs* II: 1.9--2.3 μm \[*pt=7.1*--*8.8*\]; III: 2.4--2.6 μm \[*pt=8.4*--*9.9*\] in *Macrobiotus jonesorum*), a slightly shorter macroplacoid row (4.5--6.8 μm \[*24.3*--*33.7*\] in the new species *vs* 7.0--8.4 μm \[*pt=27.0*--*34.4*\] in *Macrobiotus jonesorum*), the presence of a microplacoid, and by slightly shorter primary and secondary branches of external claws I--IV (compare [Table 2](#T2){ref-type="table"} below and [Table 2](#T2){ref-type="table"} in [@B45]).

-   ***Macrobiotus keppelensis*** Claxton, 1998 by: the lack of red pigment granules, the presence of two types of cuticular pores (small and large) in the new species *vs* pores uniform in size (*ca.* 1.0 μm) in *Macrobiotus keppelensis*), the presence of a single large pore on each of legs I-III, a slightly shorter buccal tube (17.7--22.9 μm in the new species *vs* 24.9--28.4 μm in *Macrobiotus keppelensis*), stylet supports inserted in a more anterior position (*pt=49.5*--*56.2* in the new species *vs pt≈60.6* in *Macrobiotus keppelensis*), a different macroplacoid length sequence (2\<1\<3 in the new species *vs* 2=3\<1 in *Macrobiotus keppelensis*), a slightly shorter macroplacoid row (4.5--6.8 μm in the new species *vs* 7.0--7.6 μm in *Macrobiotus keppelensis*), the lack of a membrane around egg processes, a smaller diameter of eggs with processes (55.1--55.6 μm in the new species *vs* 65.0--85.0 μm in *Macrobiotus keppelensis*), a larger number of processes on egg circumference (29--30 in the new species *vs ca.* 11 in *Macrobiotus keppelensis*), smaller egg processes (4.5--5.3 μm in the new species *vs* 11.0--16.0 μm in *Macrobiotus keppelensis*), narrower egg processes bases (2.4--3.4 μm in the new species *vs* 9.0--12.0 μm in *Macrobiotus keppelensis*), and by slightly smaller distances between egg processes (1.8--3.9 μm in the new species *vs* 4.0--6.0 μm in *Macrobiotus keppelensis*).

-   ***Macrobiotus orthofasciatus*** Fontoura et al., 2009 by: cuticular pores arranged in 9--10 transverse bands (11 transverse bands present in *Macrobiotus orthofasciatus*), the absence of tri- and quadrilobed cuticular pores, the presence of two types of pores (small and large) in the new species *vs* all pores of similar size in *Macrobiotus orthofasciatus*, the presence of a single large pore on each of legs I--III, the presence of granulation on all legs, stylet supports inserted in a more anterior position (*pt=49.5*--*56.2* in the new species *vs pt=66.5*--*67.8* in *Macrobiotus orthofasciatus*), a different shape of egg processes (short, single tip cones without a membrane in the new species *vs* screw-like processes with a membrane and six areoles in *Macrobiotus orthofasciatus*), a slightly larger number of processes on egg circumference (29--30 in the new species *vs ca.* 24 in *Macrobiotus orthofasciatus*), and by smaller distances between egg processes (1.8--3.9 μm in the new species *vs* 6.4--6.9 μm in *Macrobiotus orthofasciatus*).

-   ***Macrobiotus poricinctus*** Claxton, 1998 by: cuticular pores arranged in 9--10 transverse bands (8 transverse bands in *Macrobiotus poricinctus*), the presence of two types of pores (small and large) in the new species *vs* uniform pore size in *Macrobiotus poricinctus*), the presence of a single large pore on each of legs I--III, stylet supports inserted in a more anterior position (*pt=49.5*--*56.2* in the new species *vs pt≈59.5* in *Macrobiotus poricinctus*), a different macroplacoid length sequence (2\<1\<3 in the new species *vs* 2=3\<1 in *Macrobiotus poricinctus*), a different shape of egg processes (short, single-tipped cones without a membrane in the new species *vs.* screw-like processes within a membrane in *Macrobiotus poricinctus*), the absence of granulation on egg shell, a larger number of processes on egg circumference (29--30 in the new species *vs* 18--20 in *Macrobiotus poricinctus*), slightly smaller egg processes (4.5--5.3 μm in the new species *vs* 6.5--7.0 μm in *Macrobiotus poricinctus*), and by smaller distances between egg processes (1.8--3.9 μm in the new species *vs* 6.0--8.0 μm in *Macrobiotus poricinctus*).

-   ***Macrobiotus pustulatus*** (Ramazzotti, 1959) by: the absence of triangular and polygonal pores, the presence of two types of cuticular pores (small and large) in the new species *vs* small pores present only in the anterior part of the body, intermediate in size in the middle of the body and the large pores in the posterior part of the body in *Macrobiotus pustulatus*), the presence of a single large pore on each of legs I--III and, egg processes without a filiform bristle.

-   ***Macrobiotus ramazzottii*** Binda & Pilato, 1992 by: pores arranged in bands, the presence of two types of pores (small and large) in the new species *vs* universal pores size in *Macrobiotus ramazzottii*), the presence of a single large pore on each of legs I--III, the absence of the oral cavity armature, stylet supports inserted in a more anterior position (*pt=49.5*--*56.2* in the new species *vs pt=68.2*--*68.3* in *Macrobiotus ramazzottii*), a different macroplacoid length sequence (2\<1\<3 in the new species *vs* 3\<2\<1 in *Macrobiotus ramazzottii*), and by a lower number of processes on egg circumference (29--30 in the new species *vs ca.* 34--41 in *Macrobiotus ramazzottii*).

-   ***Macrobiotus subintermedius*** (Ramazzotti, 1962) by the presence of cuticular pores, the presence of granulation on all legs, and by fully developed lunules (only small open lunules present in *Macrobiotus subintermedius*).

-   ***Macrobiotus vinciguerrae*** Binda & Pilato, 1992 by: pores arranged in bands, the absence of tri- and quadrilobed pores, the presence of two types of pores (small and large) in the new species *vs* uniform pore size in *Macrobiotus vinciguerrae*), the presence of a single large pore on each of legs I--III, the absence of the oral cavity armature, a larger mean body size (184 μm in the new species *vs* 380 μm in *Macrobiotus vinciguerrae*), stylet supports inserted in a more anterior position (*pt=49.5*--*56.2* in the new species *vs pt=66.1*--*68.7* in *Macrobiotus vinciguerrae*), a different macroplacoid length sequence (2\<1\<3 in the new species *vs* 2\<3\<1 μm in *Macrobiotus vinciguerrae*), a smaller diameter of eggs without and with processes (44.1--45.7 μm and 55.1--55.6 μm in the new species *vs ca.* 76.4 μm and *ca.* 88.0 μm in *Macrobiotus vinciguerrae*), a slightly larger number of processes on egg circumference (29--30 in the new species *vs ca.* 26 in *Macrobiotus vinciguerrae*), egg processes without flexible filaments, smaller egg processes (4.5--5.3 μm in the new species *vs ca.* 8.2 μm in *Macrobiotus vinciguerrae*), and by narrower bases of egg processes (2.4--3.4 μm in the new species *vs ca.* 5.0 μm in *Macrobiotus vinciguerrae*).

-   ***Macrobiotus weglarskae*** Michalczyk et al., 2005 by: the absence of bi-, trilobed and star-shaped pores, the presence of two types of pores (small and large) in the new species *vs* uniform pore size in *Macrobiotus weglarskae*), the absence of 3--5 large triangular or irregularly shaped pores on the caudo-dorsal cuticle above hind legs, the presence of a single large pore on each of legs I-III, a different shape of egg processes (short, single tip cones without a membrane in the new species *vs.* screw-like processes within a membrane in *Macrobiotus weglarskae*), a slightly larger number of processes on egg circumference (29--30 in the new species *vs ca.* 24 in *Macrobiotus weglarskae*), and by slightly wider bases of egg processes (2.4--3.4 μm in the new species *vs* 1.6--2.0 μm in *Macrobiotus weglarskae*).

-   ***Macrobiotus xavieri*** Fontoura et al., 2009 by: the absence of trilobed pores, the presence of two types of pores (small and large) in the new species *vs* all pores of similar size in *Macrobiotus xavieri*), the presence of a single large pore on each of legs I--III, the presence of granulation on all legs, a smaller body size (113--236 μm in the new species *vs* 275--410 μm in *Macrobiotus xavieri*), stylet supports inserted in a more anterior position (*pt=49.5*--*56.2* in the new species *vs pt=66.1*--*67.9* in *Macrobiotus xavieri*), a different macroplacoid length sequence (2\<1\<3 in the new species *vs* 2\<3\<1 in *Macrobiotus xavieri*). shorter macroplacoids (I: 1.3--1.9 μm \[*pt=6.9*--*8.3*\]; II: 1.2--1.7 μm \[*pt=6.8*--*8.6*\] III: 1.4--2.2 μm \[*pt=7.4*--*9.6*\] in the new species *vs* I: 3.6--4.5 μm \[*12.7*--*13.8*\]; II: 2.9--3.6 μm \[*10.3*--*11.1*\] III: 3.0--3.9 μm \[*pt=10.9*--*11.9*\] in *Macrobiotus xavieri*), a shorter microplacoid (0.5--0.9 μm \[*pt=2.4*--*4.7*\] in the new species *vs* 1.5--2.0 \[*5.0*--*6.2*\] in *Macrobiotus xavieri*), a shorter macroplacoid row (4.5--6.8 μm \[*pt=24.3*--*33.7*\] in the new species *vs* 9.8--12.6 μm \[*pt=35.6*--*38.5*\] in *Macrobiotus xavieri*), a shorter placoid row (5.2--7.9 μm \[*pt=27.1*--*39.1*\] in the new species *vs* 10.9--13.9 μm \[*39.6*--*43.3*\] in *Macrobiotus xavieri*), a different shape of egg processes (short, single-tipped cones in the new species *vs* long cones with bi- or multi-tipped tips in *Macrobiotus xavieri*), egg shell and processes without granulation, a smaller diameter of eggs without and with processes (44.1--45.7 μm and 55.1--55.6 μm in the new species *vs* 56.0--79.0 μm and 80.0--99.2 μm in *Macrobiotus xavieri*), a larger number of processes on egg circumference (29--30 in the new species *vs* 20--23 in *Macrobiotus xavieri*), smaller egg processes (4.5--5.3 μm in the new species *vs* 10.6--19.0 μm in *Macrobiotus xavieri*), and by slightly narrower bases of egg processes (2.4--3.4 μm in the new species *vs* 3.7--6.6 μm in *Macrobiotus xavieri*).

### Genus: *Paramacrobiotus* Guidetti, Schill, Bertolani, Dandekar & Wolf, 2009

#### Paramacrobiotus richtersi

(Murray, 1911)

##### Localities and specimen numbers.

XI: 2 specimens (including 1 simplex) and 1 egg.

##### Remarks.

*Paramacrobiotus* species (until recently a collection of species within *Macrobiotus*) can be divided into three groups: *areolatus*, *huziori* and *richtersi*, with respect to the combination of two traits: the presence/absence of the microplacoid in the pharynx and the type of egg areolation. *Paramacrobiotus richtersi*, considered cosmopolitan, is recognised as the nominal species for a group of very similar taxa that require careful taxonomic examination of adults and egg morphology for correct identification. In the last decade many new species of this group have been described from various localities (e.g. [@B69], [@B33], [@B82], [@B49], [@B50], [@B53], [@B70], [@B71], [@B17], [@B2], [@B74]), with more recent additions including molecular data ([@B24], [@B80]).

Supplementary Material
======================

###### XML Treatment for Milnesium asiaticum

###### XML Treatment for Milnesium tardigradum tardigradum

###### XML Treatment for Diphascon (Diphascon) bullatum

###### XML Treatment for Diphascon (Diphascon) pingue pingue

###### XML Treatment for Diphascon (Diphascon) recamieri

###### XML Treatment for Diphascon (Diphascon) rugosum

###### XML Treatment for Diphascon (Adropion) prorsirostre

###### XML Treatment for Hypsibius convergens

###### XML Treatment for Hypsibius dujardini

###### XML Treatment for Hypsibius cf. scabropygus

###### XML Treatment for Astatumen bartosi

###### XML Treatment for Isohypsibius ronsisvallei

###### XML Treatment for Isohypsibius sattleri

###### XML Treatment for Macrobiotus harmsworthi harmsworthi

###### XML Treatment for Macrobiotus hufelandi hufelandi

###### XML Treatment for Minibiotus formosus

###### XML Treatment for Paramacrobiotus richtersi

KZ would like to express sincere thanks to Małgorzata Kuźnik for her help in preparing figures.

[^1]: Academic editor: L. Penev
