Procladius (Skuse, 1889)

Delimitation of Procladius View in CoL from other Tanypodinae

The family Chironomidae is usually divided into eleven subfamilies: Aphroteniinae , Buchonomyiinae , Chilenomyinae, Chironominae , Diamesinae , Orthocladiinae, Podonominae , Prodiamesinae , Tanypodinae , Telmatogetoninae , and Usambaromyiinae (Saether 2000; Bánki et al. 2024). Procladius is placed among the Tanypodinae .

The phylogenetic relationship of Tanypodinae worldwide outlined by Silva & Ekrem (2016) recognized the tribe Procladiini containing the genera Procladius , Djalmabatista , Laurotanypus , Lepidopelopia and Saetheromyia . The study was based on DNA barcodes and morphology of adults, pupae and larvae. Only Procladius is present in Europe.

Djalmabatista View in CoL is the closest relative to Procladius View in CoL according to Silva & Ekrem (2016). Krosch et al. (2022) arrived at the same conclusion but suggested that Djalmabatista View in CoL , based on barcodes mainly from Australia, might be a subgenus of Procladius View in CoL . This has been suggested before e.g. by Saether &Andersen (2000) who made morphological studies of adult males, adult females, pupae and larvae of a species in Africa assigned to Djalmabatista View in CoL . This species should be named Procladius reidi ( Freeman, 1955) View in CoL based on the description in Saether & Andersen (2000) and barcoding results of P. reidi View in CoL from China and Pakistan gained from BOLD ( Ratnasingham et al. 2024). Barcode sequences place P. reidi View in CoL closer to several specimens of Procladius View in CoL than those of Djalmabatista View in CoL . In the key to adult males below, Djalmabatista View in CoL is kept as a genus as more studies are needed to reveal its status as a genus or a subgenus of Procladius View in CoL . Djalmabatista View in CoL includes about 15 species found in Africa, Asia, Australia, North America and South America according to BOLD and Systema Dipterorum ( Evenhuis & Pape 2024). Findings closest to Europe are from Saudi Arabia.

Laurotanypus View in CoL , containing one species from Brazil, is regarded as a synonym of Procladius View in CoL . Dantas & Hamada (2018) placed L. travassosi (Oliveira, Messias & Silva-Vasconcelos, 1992) View in CoL in Procladius View in CoL . Silva (2019) regarded Procladius travassosi to be a junior synonym of Procladius stroudi Roback, 1982 View in CoL . Laurotanypus View in CoL is considered to be a subgenus of Procladius View in CoL in the key to adult males below.

Lepidopelopia View in CoL includes only one species which has been recorded from several countries in tropical Africa ( Harrison 1970; Saether & Andersen 2000). Saetheromyia View in CoL includes one species in Japan ( Niitsuma 2007) and one undescribed species in China with barcoding results in BOLD.

The following key can be used to distinguish males of the tribe Procladiini View in CoL from other tribes and subfamilies of Chironomidae View in CoL , and further to separate the genus Procladius View in CoL from other Procladiini View in CoL .

1. Wing crossvein MCu absent. Subfamilies Aphroteniinae , Chironominae , Orthocladiinae, Telmatogetoninae and Usambaromyiinae - Wing crossvein MCu present ( Fig. 5 View FIGURE 5 )..................................................................... 2. 2. Wing vein R2+3 absent. Subfamilies Buchonomyiinae , Chilenomyinae, Podonominae , some Tanypodinae - Wing vein R2+3 present ( Fig. 5 View FIGURE 5 ).......................................................................... 3. 3. Antenna with 5‒13 flagellomeres, last flagellomere longer than penultimate flagellomere. Subfamilies Diamesinae and

Prodiamesinae - Antenna with 14 flagellomeres, last flagellomere more than three times shorter than penultimate flagellomere ( Fig. 1 View FIGURE 1 ). Subfamily

Tanypodinae ......................................................................................... 4. 4. Leg fourth tarsomere broadened from base to end and shorter than fifth tarsomere. Hind leg tibial comb double. Coelopynia ,

Clinotanypus , Coelotanypus and Naelotanypus - Leg fourth tarsomere cylindrical and longer to slightly shorter than fifth tarsomere. Hind leg tibial comb simple ( Fig. 10 View FIGURES 9‒10 ) or absent............................................................................................... 5. 5. Wing vein Cu fork FCu (division into M3+4 and CuA) proximal to opposite MCu. - Wing vein Cu fork FCu distal MCu ( Figs. 5 View FIGURE 5 and 6 View FIGURE 6 )........................................................... 6. 6. Distance between MCu and FCu much less than half as long as M3+4. - Distance between MCu and FCu at least half as long as M3+4 ( Figs. 5 View FIGURE 5 and 6 View FIGURE 6 ). Procladiini ........................... 7. 7. Wing with more the 20 macrotrichia. Gonostylus with or without inner lobe....................................... 8. - Wing without or less than 5 macrotrichia. Gonostylus with distinct convex inner lobe with strong setae. Procladius subgenus

Psilotanypus 8. Gonostylus with protruding convex inner lobe with strong setae................................................. 9. - Gonostylus without distinct inner lobe, inner margin straight or concave (rarely slightly convex) without strong setae..... 10. 9. Scutal tubercle absent. Abdomen tergites light and mostly with darker median longitudinal band. Djalmabatista - Scutal tubercle present. Abdomen tergites not with darker median longitudinal band. Procladius subgenus Laurotanypus 10. Scale-like setae on thorax, legs and wings. Wing vein R2+3 not divided. Gonostylus without indication of outer process.

Lepidopelopia - No scale-like setae. Wing vein R2+3 divided into R2 and R3 ( Fig. 5 View FIGURE 5 ). Gonostylus with or without outer process......... 11. 11. Gonostylus without indication of outer process. Scutal tubercle present. Saetheromyia

Gonostylus with indicated to very long outer process ( Figs. 17 View FIGURES 16‒17 and 18 View FIGURES 18‒19 ). Scutal tubercle absent. Procladius subgenus

Holotanypus and Procladius

Species delimitation and identification of European Procladius

The iterative process comparing morphological data and barcoded data resulted in a list of 27 species of Procladius present in Europe ( Table 6) of which four are new species to science, namely P. exilis Brodin , P. gemma Brodin , P. saeticubitus Brodin and P. tenebricosus Brodin & Hellberg.

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Barcodes are available for 25 of the species. Table 6 shows that BOLD contained barcodes and BINs for 23 species. P. clavus Roback, 1971 and P. tatrensis have a few good barcodes of more than 550 base pars not yet incorporated into BOLD or GenBank. P. fimbriatus Wülker, 1959 and P. gemma lack known barcodes, but they are readily identified species based on several unique morphological characters.

Each of the 42 BINs in Table 6 contains only one species of Procladius , but several species have more than one BIN. Several other studies of Chironomidae ( Song et al. 2016; Chimeno et al. 2023) and other insects have shown that it is common that species have several BINs. The BINs for Procladius are constructed by barcodes of 6 414 specimens, ranging from 1 for P. clavus to 2 336 for P. dentus . The BINs contain specimens from 35 countries or autonomous regions. About 75% of the barcoded specimens are from Canada. P. culiciformis and P. lugens Kieffer, 1915 have four BINs each, while the others have tree to one BIN each.

Intraspecific distance ranges from 0 to 3.3% ( Table 6). Only P. ferrugineus (Kieffer, 1918) has an intraspecific distance exceeding that of the interspecific distance. Interspecific distance, expressed as distance to nearest neighbour in BOLD, ranges from 2.0 to 8.8%. Six species have an interspecific distance below 3.3%. Notably, five of these are among those most easily identified morphologically. The BIN distance between the sixth species P. tenebricosus and its nearest neighbour P. ferrugineus is only 2.0%. The species are readily distinguished from each other by the form of the phallapodeme. Some species have BINs with the closest neighbour BIN not of morphologically very similar species but of species with very different morphology. The most striking example is P. appropinquatus ( Lundström, 1916) . with almost no gonostylus process and P. dentus with a very long gonostylus process.

In addition to the 42 BINs thoroughly studied, there are 6 more BINs of Procladius in BOLD which contain at least one specimen from Europe. These BINs (BOLD:ABX4068, BOLD:ACQ5869, BOLD: ADA 6883, BOLD: ADA 8287, BOLD:AEE2525, BOLD:AEE7861) were not included in the analyses of the present study as they contain morphologically deformed males, contain no adult males, or lack adult males available for studies. Whether or not these BINs represent valid species of Procladius is an open question.

Measurement data of the analyses of one hundred morphological characters (Supplement 1: https://doi.org/10.6084/ m9.figshare.28189610) showed that nine of the Procadius species can be identified by at least one non-overlapping character. Seven species can be identified by one or more non-overlapping genitalia characters, viz. the unique form of the gonocoxite ( P. dentus ), gonostylus ( P. clavus and P. crassinervis ), phallapodeme ( P. ferrugineus ) or medioapodeme ( P. gemma , P. signatus and P. dentus ). P. flavifrons Edwards, 1929 can be distinguished by the numerous setae on the katepisternum. P. fimbriatus is effortlessly identified by its femalized head and an antenna with a much lower AR than that of all other species. Reliable identification of each of the other eighteen species mostly requires measurements of several morphological characters.

The most useful character for species identification is GspR, a ratio constructed as the gonostylus outer length divided by the gonostylus process length ( Table 2 item G22, Fig. 17 View FIGURES 16‒17 ). GspR is always one of the characters when more than one character is needed for species identification. An advantage is that GspR is usually easily measured for specimens prepared on slides or kept in alcohol.

Other frequently useful morphological characters for separation of morphologically similar species are antenna AR ( Table 2, item H2, Fig. 1 View FIGURE 1 ), macrotrichia on the median anepisternum ( Table 2, item T10, Fig. 4 View FIGURE 4 ), front leg beard ratio ( Table 2, item L10, Fig. 9 View FIGURES 9‒10 ), wing length ( Table 2, item W1, Fig. 5 View FIGURE 5 ), and thickness of the gonostylus measured as GsmR ( Table 2, item G23, Fig. 18 View FIGURES 18‒19 ).

Species key to adult males of European Procladius

Each step (couplet) in a key for morphological species identification should preferably be dichotomous and give two options for one morphological character. This approach is often not sufficient for distinguishing species of Procladius because of frequent and sometimes considerable overlaps in male characters.

In the key below one to three characters are provided for major divisions, while three characters are always available for species separation. The characters of each division are ranked according to importance (I‒III) for species identification. Taking all three characters and the drawings of male genitalia ( Figs. 20‒127 View FIGURES 20‒23 View FIGURES 24‒27 View FIGURES 28‒31 View FIGURES 32‒35 View FIGURES 36‒39 View FIGURES 40‒43 View FIGURES 44‒46 View FIGURES 47‒49 View FIGURES 50‒52 View FIGURES 53‒55 View FIGURES 56‒58 View FIGURES 59‒61 View FIGURES 62‒64 View FIGURES 65‒67 View FIGURES 68‒70 View FIGURES 71‒73 View FIGURES 74‒76 View FIGURES 77‒79 View FIGURES 80‒82 View FIGURES 83‒85 View FIGURES 86‒88 View FIGURES 89‒91 View FIGURES 92‒94 View FIGURES 95‒97 View FIGURES 98‒100 View FIGURES 101‒103 View FIGURES 104‒106 View FIGURES 107‒109 View FIGURES 110‒112 View FIGURES 113‒115 View FIGURES 116‒118 View FIGURES 119‒121 View FIGURES 122‒124 View FIGURES 125‒127 ) into account is often needed to achieve reliable species identification. Most characters have a code within parenthesis, e.g. (A1), that is explained in Table 2 and illustrated in Figs. 1‒19 View FIGURE 1 View FIGURES 2‒3 View FIGURE 4 View FIGURE 5 View FIGURE 6 View FIGURES 7‒8 View FIGURES 9‒10 View FIGURES 11‒12 View FIGURE 13 View FIGURES 14‒15 View FIGURES 16‒17 View FIGURES 18‒19 .

The key contains 28 of the 100 characters chosen for the present study. Before using the key, measurement of GspR, gonostylus process length divided by gonostylus outer length (G22, Fig. 17 View FIGURES 16‒17 ) is recommended as this character forms the base of species identification.

If the key does not result in satisfactory species identification, two additional instruments can be consulted. The helpdesk (heading below, Table 7) contains an overview of 21 morphological characters important for species separation. The subheadings “Diagnostical characters” and “Geographical distribution and ecology” for each species (below under heading “The species; systematics, distribution and ecology”) contains information particularly useful to separate morphologically similar species. The helpdesk and diagnostical characters are also useful for quality assurance of species determinations.

characters of a species are marked in bold. The characters are explained in Table 2 and illustrated in Fig. 1‒19 View FIGURE 1 View FIGURES 2‒3 View FIGURE 4 View FIGURE 5 View FIGURE 6 View FIGURES 7‒8 View FIGURES 9‒10 View FIGURES 11‒12 View FIGURE 13 View FIGURES 14‒15 View FIGURES 16‒17 View FIGURES 18‒19 .

......continued on the next page characters of a species are marked in bold. Characters are explained in Table 2 and illustrated in Fig. 1‒19 View FIGURE 1 View FIGURES 2‒3 View FIGURE 4 View FIGURE 5 View FIGURE 6 View FIGURES 7‒8 View FIGURES 9‒10 View FIGURES 11‒12 View FIGURE 13 View FIGURES 14‒15 View FIGURES 16‒17 View FIGURES 18‒19 .

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1. I) Wing; membrane with more than 20 macrotrichia (W3, Fig. 6 View FIGURE 6 ). II) Genitalia; gonostylus inner margin slightly concave to slightly convex without strong setae (G28, Fig. 17 View FIGURES 16‒17 )........................................................... 2.

- I) Wing; membrane without macrotrichia or rarely 1‒5 at wing apex (W3). II) Genitalia; gonostylus inner margin distinctly convex with strong setae (G28, Fig. 113 View FIGURES 113‒115 ).................................................................. 23.

2. I) Head; antenna AR 0.3‒0.6 (H2). II) Genitalia; gonostylus process long GspR 0.34‒0.40 (G22, Fig. 47 View FIGURES 47‒49 ). III) Genitalia; gonocoxite width 264‒311 µm (G11, Fig. 16 View FIGURES 16‒17 ). Figs. 20 View FIGURES 20‒23 , 47‒49. P View FIGURES 47‒49 . fimbriatus Wülker, 1959

- I) Head; antenna AR 1.3‒3.2 (H2, Fig. 1 View FIGURE 1 ). II. Genitalia; gonostylus process very short to very long GspR 0.02‒0.48 (G22, Figs. 56 View FIGURES 56‒58 and 65 View FIGURES 65‒67 ). III) Genitalia; gonocoxite width 185‒532 µm (G11, Fig. 16 View FIGURES 16‒17 )......................................... 3.

3. I) Genitalia; medioapodeme long and inner section strongly curved backwards 30‒105° (G3, Fig. 14 View FIGURES 14‒15 ).................. 4.

- I) Genitalia; medioapodeme short and inner section not or slightly curved backwards 0‒10° (G3, Fig. 15 View FIGURES 14‒15 )............... 7.

4. I) Genitalia; gonostylus process short GspR 0.10‒0.22 (G22, Fig. 18 View FIGURES 18‒19 )............................................ 5.

- I) Genitalia; gonostylus process long to very long GspR 0.36‒0.48 (G22, Fig. 17 View FIGURES 16‒17 ).................................. 6.

5. I) Genitalia; medioapodeme anterior margin undulating ( Fig. 21 View FIGURES 20‒23 ). II) Abdomen; tergite IX hind mid margin with a ball-formed projection (A7, Fig. 21 View FIGURES 20‒23 ). III) Genitalia; gonostylus process length/width 0.8‒1.1 (G21, Fig. 50 View FIGURES 50‒52 ). Figs. 21 View FIGURES 20‒23 , 50‒52. P View FIGURES 50‒52 . gemma Brodin, new species

- I) Genitalia; medioapodeme anterior margin straight ( Fig. 22 View FIGURES 20‒23 ). II) Abdomen; tergite IX hind mid margin with a semicircular projection (A7, Fig. 22 View FIGURES 20‒23 ). III) Genitalia; gonostylus process length/width 0.5‒0.8 (G21, Fig. 53 View FIGURES 53‒55 ). Figs. 22 View FIGURES 20‒23 , 53‒55. P View FIGURES 53‒55 . clavus Roback, 1971

6. I) Genitalia; gonocoxite with a marked dorsal ridge ( Fig. 23 View FIGURES 20‒23 ). II) Genitalia; medioapodeme mid with long anteriorly directed process ( Fig. 23 View FIGURES 20‒23 ). III) Head; antenna AR 2.5‒3.0 (H2). Figs. 14 View FIGURES 14‒15 , 23 View FIGURES 20‒23 , 56‒58. P View FIGURES 56‒58 . dentus Roback, 1971

- I) Genitalia; gonocoxite without a marked dorsal ridge ( Fig. 24 View FIGURES 24‒27 ). II) Genitalia; medioapodeme mid without anteriorly directed process ( Fig. 24 View FIGURES 24‒27 ). III) Head; antenna AR 1.9‒2.5 (H2). Figs. 17 View FIGURES 16‒17 , 24 View FIGURES 24‒27 , 59‒61. P View FIGURES 59‒61 . signatus ( Zetterstedt, 1850)

7. I) Genitalia; gonostylus process long GspR 0.35‒0.43 (G22, Fig. 63 View FIGURES 62‒64 ). II) Wing; length 3.5‒4.5 mm (W1). III) Thorax; median anepisternum with 7‒21 setae on each side (T10). Figs. 11 View FIGURES 11‒12 , 25 View FIGURES 24‒27 , 62‒64. P View FIGURES 62‒64 . tatrensis Gowin, 1944

- I) Genitalia; gonostylus process very short to long GspR 0.02‒0.39 (G22), if>0.34 then II) Wing; length <3.5 mm (W1) and III) Thorax; median anepisternum with 0‒6 setae on each side (T10, Fig. 4 View FIGURE 4 )....................................... 8.

8. I) Genitalia; gonostylus process slightly indicated to short GspR 0.02‒0.10 (G22, Figs. 66 View FIGURES 65‒67 and 69 View FIGURES 68‒70 ). II) Genitalia; gonostylus process length/base width 0.1‒0.5 (G21, Fig. 66 View FIGURES 65‒67 ). III) Genitalia; gonocoxite width 282‒451 µm (G11, Fig. 16 View FIGURES 16‒17 )........... 9.

- I) Genitalia; gonostylus process short to very long GspR 0.10‒0.39 (G22). II) Genitalia; if gonostylus process length/base width <0.6 (G21, Fig. 17 View FIGURES 16‒17 ) then III) Genitalia; gonocoxite width <265 µm (G11, Fig. 16 View FIGURES 16‒17 )............................ 10.

9. I) Genitalia; gonocoxite width 377‒451 µm (G11). II) Leg; hind leg each tibial comb with 8‒10 spines (L18). III) Wing; length 3.7‒4.6 mm (W1). Figs. 9 View FIGURES 9‒10 , 26 View FIGURES 24‒27 , 65‒67. P View FIGURES 65‒67 . simplicistilus Freeman, 1948

- I) Genitalia; gonocoxite width 282‒350 µm (G11). II) Leg; hind leg each tibial comb with 11‒16 spines (L18, Fig. 10 View FIGURES 9‒10 ). III) Wing; length 2.5‒3.7 mm (W1). Figs. 27 View FIGURES 24‒27 , 68‒70. P View FIGURES 68‒70 . appropinquatus ( Lundström, 1916)

10. I) Leg; hind leg tibial comb with 6‒9 spines (L18). II) Leg; tarsi of front leg with very long setae BRI 5.5‒8 (L10, Fig. 9 View FIGURES 9‒10 ). III) Thorax; median anepisternum with 10‒18 setae on each side (T10)............................................. 11.

- I) Leg; hind leg tibial comb with 9‒14 spines (L18, Fig. 10 View FIGURES 9‒10 ), if <10 spines then II) Leg; tarsi of front leg with BRI <5 (L10, Fig. 9 View FIGURES 9‒10 ). III) Thorax; median anepisternum with 0‒26 setae on each side (T10, Fig. 4 View FIGURE 4 ).................................. 12.

11. I) Genitalia; gonostylus process GspR 0.23‒0.31 (G22, Fig. 28 View FIGURES 28‒31 ). II) Wing; Cu stem vein before MCu without setae (W11, Fig. 5 View FIGURE 5 ). III) Head; palpomere five length/width 7.2‒8.5 (H9). Figs. 28 View FIGURES 28‒31 , 71‒73. P View FIGURES 71‒73 . lugubris ( Zetterstedt, 1850)

- I) Genitalia; gonostylus process GspR 0.14‒0.19 (G22, Fig. 29 View FIGURES 28‒31 ). II) Wing; Cu stem vein before MCu with 4‒11 setae (W11). III) Head; palpomere five length/width 10.7‒11.3 (H9, Fig. 2 View FIGURES 2‒3 ). Figs. 29 View FIGURES 28‒31 , 74‒76. P View FIGURES 74‒76 . exilis Brodin, new species

12. I) Genitalia; phallapodeme inner section about as dark and basally as broad as outer section (G6, G7, Fig. 30 View FIGURES 28‒31 ). II) Genitalia; phallapodeme mid not to moderately angled 0‒30° (G5, Fig. 30 View FIGURES 28‒31 ). III) Genitalia; gonostylus process moderately long GspR 0.27‒0.33 (G22, Fig. 77 View FIGURES 77‒79 ). Figs. 30 View FIGURES 28‒31 , 77‒79. P View FIGURES 77‒79 . ferrugineus (Kieffer, 1918)

- I) Genitalia; phallapodeme inner section lighter to much lighter and basally distinctly thinner than outer section (G6, G7, Fig. 15 View FIGURES 14‒15 ). II) Genitalia; phallapodeme mid moderately to strongly angled 25‒90° (G5, Fig. 15 View FIGURES 14‒15 ). III) Genitalia; gonostylus process short to long GspR 0.10‒0.39 (G22)...................................................................... 13.

13. I) Genitalia, gonostylus process strongly oriented upwards 30‒60° (G33, Fig. 19 View FIGURES 18‒19 ). II) Genitalia; gonostylus process strongly diverging 40‒75° (G31, Fig. 80 View FIGURES 80‒82 ). III) Genitalia; gonostylus process long GspR 0.32‒0.39 (G22, Fig. 80 View FIGURES 80‒82 ). Figs. 19 View FIGURES 18‒19 , 31 View FIGURES 28‒31 , 80‒82. P View FIGURES 80‒82 . crassinervis ( Zetterstedt, 1838)

- I) Genitalia; gonostylus process not to moderately oriented upwards 0‒25° (G33, Fig. 19 View FIGURES 18‒19 ). II) Genitalia; gonostylus process not to strongly diverging 0‒50° (G31, Fig. 83 View FIGURES 83‒85 ). III) Genitalia; gonostylus process short to long GspR 0.10‒0.37 (G22)....... 14.

14. I) Thorax; median anepisternum with 5‒26 setae (T10). II) Head; palpomere five length/width 8.0‒9.3 (H9). III) Genitalia; gonostylus process moderately long GspR 0.25‒0.33 (G22, Fig. 83 View FIGURES 83‒85 ). Figs. 32 View FIGURES 32‒35 , 83‒85. P View FIGURES 83‒85 . frigidus ( Holmgren, 1869)

- I) Thorax; median anepisternum with 0‒6 setae (T10, Fig. 4 View FIGURE 4 ). II) Head; palpomere five length/width 9.1‒12.3 (H9, Fig. 2 View FIGURES 2‒3 ), if <9,4 then median anepisternum with <4 setae (T10). III) Genitalia; gonostylus process short to long GspR 0.10‒0.37 (G22).

.................................................................................................. 15.

15. I) Genitalia; gonostylus process moderately to strongly diverging 30‒50° (G31, Fig. 86 View FIGURES 86‒88 ). II) Body; length 3.6‒4.4 mm (B1). III). Genitalia; gonostylus process rather long to long GspR 0.28‒0.37 (G22, Fig. 86 View FIGURES 86‒88 ). Figs. 33 View FIGURES 32‒35 , 86‒88. P View FIGURES 86‒88 . floralis Kieffer, 1915

- I) Genitalia; gonostylus process not to moderately diverging 0‒30° (G31, Fig. 89 View FIGURES 89‒91 ), if>25° then II) Body; length> 4.4 mm (B1). III) Genitalia; gonostylus process short to long GspR 0.10‒0.34 (G22).......................................... 16.

16. I) Genitalia; gonostylus process medium long to long GspR 0.23‒0.34 (G22, Fig. 89 View FIGURES 89‒91 ). II) Genitalia; gonostylus outer length/ mid width GsmR 5.2‒6.9 (G23, Fig. 89 View FIGURES 89‒91 ). III) Genitalia; gonostylus process length/width 1.2‒1.9 (G21, Figs. 17 View FIGURES 16‒17 and 89 View FIGURES 89‒91 )... 17.

- I) Genitalia; gonostylus process short to medium long GspR 0.10‒0.25 (G22, Fig. 18 View FIGURES 18‒19 ), if>0.23 then II) Genitalia; gonostylus outer length/mid width GsmR <5.2 (G23, Fig. 107 View FIGURES 107‒109 ) and III) Genitalia; gonostylus process length/width <1.4 (G21, Figs. 17 View FIGURES 16‒17 and 18 View FIGURES 18‒19 )................................................................................................ 19.

17. I) Wing; with distinct to very distinct dark patch in anal cell (W7, Fig. 6 View FIGURE 6 ). II) Leg; hind leg tibia length 0.84‒1.17 mm (L16). III) Genitalia; gonostylus process medium to rather long GspR 0.23‒0.29 (G22, Fig. 89 View FIGURES 89‒91 ). Figs. 6 View FIGURE 6 , 12 View FIGURES 11‒12 , 34 View FIGURES 32‒35 , 89‒91. P View FIGURES 89‒91 . tenebricosus Brodin & Hellberg, new species

- I) Wing; with no or faint dark patch in anal cell (W7, Fig. 5 View FIGURE 5 ). II) Leg; hind leg tibia length 1.21‒1.71 mm (L16). III) Genitalia; gonostylus process medium to long GspR 0.23‒0.34 (G22)................................................... 18.

18. I) Genitalia; outer section length/gonocoxite base width 0.34‒0.46 (G10, Fig. 13 View FIGURE 13 ). II) Genitalia; gonostylus process medium to long GspR 0.25‒0.34 (G22, Fig. 92 View FIGURES 92‒94 ). III) Genitalia; gonostylus outer length/mid width GsmR 5.5‒6.9 (G23, Fig. 92 View FIGURES 92‒94 ). Figs. 19 View FIGURES 18‒19 , 35 View FIGURES 32‒35 , 92‒94. P View FIGURES 92‒94 . longistilus (Kieffer, 1916)

- I) Genitalia; phallapodeme outer section length/gonocoxite base width 0.27‒0.36 (G10, Fig. 13 View FIGURE 13 ). II) Genitalia; gonostylus process medium long to rather long GspR 0.23‒0.30 (G22, Fig. 95 View FIGURES 95‒97 ). III) Genitalia; gonostylus outer length/mid width GsmR 5.2‒6.3 (G23, Fig. 95 View FIGURES 95‒97 ). Figs. 5 View FIGURE 5 , 8 View FIGURES 7‒8 , 16 View FIGURES 16‒17 , 36 View FIGURES 36‒39 , 95‒97. P View FIGURES 95‒97 . pruinosus ( Kieffer, 1924)

19. I) Genitalia; gonostylus outer length/mid width GsmR 5.9‒6.9 (G23, Figs. 18 View FIGURES 18‒19 and 98 View FIGURES 98‒100 ). II) Wing; Cu stem vein before MCu with 5‒33 setae (W11). III) Genitalia; gonostylus process short to rather short GspR 0.14‒0.20 (G22, Fig. 98 View FIGURES 98‒100 ). Figs. 37 View FIGURES 36‒39 , 98‒100. P View FIGURES 98‒100 . saeticubitus Brodin, new species

- II) Genitalia; gonostylus outer length/mid width GsmR 4.2‒6.4 (G23, Figs. 18 View FIGURES 18‒19 and 101 View FIGURES 101‒103 ), if>5.8 then II) Wing; Cu stem vein before MCu with 0‒4 setae (W11, Fig. 5 View FIGURE 5 ) and/or III) Genitalia; gonostylus process GspR>0.20 (G22, Fig. 101 View FIGURES 101‒103 )......... 20.

20. I) Genitalia; gonostylus outer length/mid width GsmR 5.3‒6.4 (G23, Figs. 18 View FIGURES 18‒19 and 101 View FIGURES 101‒103 ). II) Leg; front leg BR 3‒6.5 (L10, Fig. 9 View FIGURES 9‒10 ). III) Genitalia; gonostylus process rather short GspR 0.18‒0.24 (G22, Fig. 101 View FIGURES 101‒103 ). Figs. 7 View FIGURES 7‒8 , 15 View FIGURES 14‒15 , 38 View FIGURES 36‒39 , 101‒103. P View FIGURES 101‒103 . islandicus ( Goetghebuer, 1931)

- I) Genitalia; gonostylus outer length/mid width GsmR 4.2‒5.7 (G23, Figs. 18 View FIGURES 18‒19 and 104 View FIGURES 104‒106 ), if>5.2 then II) Leg; front leg BR <3 (L10, Fig. 9 View FIGURES 9‒10 ) and/or III) Genitalia; gonostylus process GspR <0.18 (G22, Fig. 104 View FIGURES 104‒106 )................................ 21.

21. I) Genitalia; gonostylus process short GspR 0.10‒0.16 (G22, Fig. 104 View FIGURES 104‒106 ). II) Leg; mid leg tibia length 1.05‒1.26 mm (L13). III) Genitalia; gonostylus process length/width 0.5‒0.8 (G21, Figs. 18 View FIGURES 18‒19 and 104 View FIGURES 104‒106 ). Figs. 39 View FIGURES 36‒39 , 104‒106. P View FIGURES 104‒106 . breviatus Remmert, 1953.

- I) Genitalia; gonostylus process short to medium long GspR 0.13‒0.25 (G22, Fig. 107 View FIGURES 107‒109 ), if <0.16 then II) Leg; mid leg tibia length <1.05 mm (L13) and/or III) Genitalia; gonostylus process length/width>0.8 (G21, Figs. 17 View FIGURES 16‒17 and 107 View FIGURES 107‒109 )............ 22.

22. I) Genitalia; gonostylus process rather short to medium long GspR 0.18‒0.24 (G22, Fig. 107 View FIGURES 107‒109 ). II) Genitalia; gonostylus outer length/mid width GsmR 4.2‒5.2 (G23, Figs. 18 View FIGURES 18‒19 and 107 View FIGURES 107‒109 ). III) Body; length 4.4‒5.7 mm (B1). Figs. 1 View FIGURE 1 , 3 View FIGURES 2‒3 , 13 View FIGURE 13 , 40 View FIGURES 40‒43 , 107‒109. P View FIGURES 107‒109 . culiciformis ( Linnaeus, 1767)

- I) Genitalia; gonostylus process short to rather short GspR 0.13‒0.20 (G22, Fig. 110 View FIGURES 110‒112 ). II) Genitalia; gonostylus outer length/ mid width GsmR 4.9‒5.6 (G23, Figs. 18 View FIGURES 18‒19 and 110 View FIGURES 110‒112 ). III) Body; length 2.9‒4.4 mm (B1). Figs. 2 View FIGURES 2‒3 , 18 View FIGURES 18‒19 , 41 View FIGURES 40‒43 , 110‒112. P View FIGURES 110‒112 . choreus ( Meigen, 1804)

23. I) Genitalia; gonostylus process short GspR 0.10‒0.19 (G22, Fig. 113 View FIGURES 113‒115 ). II) Genitalia; gonostylus process length/width 0.4‒0.9 (G21, Fig. 113 View FIGURES 113‒115 )...................................................................................... 24.

- I) Genitalia; gonostylus process absent to very short GspR 0‒0.06 (G22, Fig. 119 View FIGURES 119‒121 ). II) Genitalia; gonostylus process length/ width 0‒0.4 (G21, Fig. 119 View FIGURES 119‒121 )............................................................................ 25.

24. I) Genitalia; gonostylus outer length/mid width GsmR 2.6‒3.3 (G23, Figs. 18 View FIGURES 18‒19 and 113 View FIGURES 113‒115 ). II) Genitalia; gonostylus outer length/ gonocoxite length 0.42‒0.50 (G25, Figs. 16 View FIGURES 16‒17 and 113 View FIGURES 113‒115 ). IIII) Genitalia; gonocoxite base width 206‒285 µm (G11, Fig. 16 View FIGURES 16‒17 ). Figs. 42 View FIGURES 40‒43 , 113‒115. P View FIGURES 113‒115 . bellus ( Loew, 1866).......................................................................

- I) Genitalia; gonostylus outer length/mid width GsmR 3.2‒3.9 (G23, Figs. 18 View FIGURES 18‒19 and 116 View FIGURES 116‒118 ). II) Genitalia; gonostylus outer length/ gonocoxite length 0.50‒0.59 (G25, Fig. 16 View FIGURES 16‒17 ). IIII) Genitalia; gonocoxite base width 178‒216 µm (G11, Fig. 16 View FIGURES 16‒17 ). Figs. 43 View FIGURES 40‒43 , 116‒118. P View FIGURES 116‒118 . nudipennis Brundin, 1947

25. I) Thorax; katepisternum with 9‒18 setae on each side (T11, Fig. 4 View FIGURE 4 ). II) Head; antenna AR 1.3‒1.7 (H2, Fig. 1 View FIGURE 1 ). III) Wing; length 1.7‒2.3 mm (W1, Fig. 5 View FIGURE 5 ). Figs. 4 View FIGURE 4 , 44 View FIGURES 44‒46 , 119‒121. P View FIGURES 119‒121 . flavifrons Edwards, 1929

- I) Thorax; katepisternum without setae (T11). II) Head; antenna AR 1.7‒2.6 (H2, Fig. 1 View FIGURE 1 ). III) Wing; length 2.1‒3.4 mm (W1, Fig. 5 View FIGURE 5 )............................................................................................. 26.

26. I) Genitalia; gonostylus outer margin smoothly curved in basal section ( Fig. 122 View FIGURES 122‒124 ). II) Leg; front leg tibia length 0.83‒1.17 mm (L1). III) Head; antenna AR 1.7‒2.3 (H2, Fig. 1 View FIGURE 1 ). Figs. 45 View FIGURES 44‒46 , 122‒124. P View FIGURES 122‒124 . lugens Kieffer, 1915

- I) Genitalia; gonostylus outer margin abruptly angled in basal section ( Fig. 125 View FIGURES 125‒127 ). II) Leg; front leg tibia length 1.21‒1.39 mm (L1). III) Head; antenna AR 2.2‒2.6 (H2, Fig. 1 View FIGURE 1 ). Figs. 10 View FIGURES 9‒10 , 46 View FIGURES 44‒46 , 125‒127. P View FIGURES 125‒127 . imicola Kieffer, 1922

Species identification helpdesk

Consulting the key above might not always result in unambiguous species identification of European Procladius . Cases where two, or more rarely even three, species are possible may occur because of overlaps in measurements of morphological characters. The key might also have a limited functionality if diagnostic species characters of a specimen studied are obscured, deformed or completely lost, such as antennae or tarsomere setae for measurement of leg BR (Tabel 2, L10).

Table 7 functions as a helpdesk for species identification if the key is not sufficient. The helpdesk contains 21 important diagnostic characters, of which two are not in the key. The fictive example in Table 8 shows how Table 7 by adopting a stepwise procedure can be used for species identification of a difficult specimen. Usually 7‒10 steps may be enough, but in a few cases it might be necessary to go through all 21 steps to reach a conclusion. Table 7 may also serve as a general quality assurance tool for reliable species identification.

A test including a hundred specimens indicates that the most problematic species separation issues are those of the “species pairs” P. choreus / P. culiciformis , P. choreus / P. islandicus , P. culiciformis / P. pruinosus , P. pruinosus / P. tenebricosus and P. imicola / P. lugens .

TABLE 8. Stepwise procedure to reveal species identity of a fictive Procladius male. Step 1a: Measure GspR of the fictive male, e.g. 0.20. Step 1b: Mark all species in Table 7A 1 with a GspR-interval covering 0.20. Step 2‒7: Mark agreement with fictive male e.g. with green and disagreement e.g. with red figures. Step 8‒21: Continue to Table 7B‒C if necessary for reliable species identification. Here, P. islandicus is the most likely species as it is the only one with all figures marked green.

Species Character

G22) W3) wing G23) A2) tergite G31) gonostylus W1) wing T10)

gonostylus membrane gonostylus II‒IV posterior process length, mm anepisternum

GspR setae GsmR colour divergence ° setae

Step 1 Step 2 Step 3 Step 4 Step 5 Step 6 Step 7

fictive male 0.20 3 5.5 2 10 3.1 2

P. choreus 0.13‒0.20 2‒3 4.9‒5.6 0‒1 0‒15 1.8‒2.8 0‒2

P. saeticubitus 0.14‒0.20 3 5.9‒6.9 0.5‒2.5 0‒15 2.6‒3.4 0‒3

P. gemma 0.17‒0.22 2 5.4‒5.7 2‒2.5 25‒45 3.7‒4.3 0

P. islandicus 0.18‒0.24 2.5‒3 5.3‒6.4 1‒2.5 0‒15 2.6‒3.5 0‒3

P. culiciformis 0.18‒0.25 2‒3 4.2‒5.2 0.5‒2 0‒25 2.4‒3.5 0

Species identification of other than European Procladius

Twelve species not known from Europe, but might be found there in the future, have also been studied using several characters of the 100-character analysis (Supplement 1). The most important characteristics to identify these species are presented in Table 9.

from Europe but possibly found there in the future. Characters are explained in Table 2 and illustrated in Fig. 1‒19 View FIGURE 1 View FIGURES 2‒3 View FIGURE 4 View FIGURE 5 View FIGURE 6 View FIGURES 7‒8 View FIGURES 9‒10 View FIGURES 11‒12 View FIGURE 13 View FIGURES 14‒15 View FIGURES 16‒17 View FIGURES 18‒19 .

Procladius barbatulus Sublette, 1964 View in CoL ( Sublette 1964; Roback 1971; Murray & Fittkau 1989 as P. bellus View in CoL ) found in the United States appears at couplet 22 in the key above. P. barbatulus View in CoL is similar to P. choreus View in CoL and P. culiciformis View in CoL but has a broader gonostylus process expressed as process width divided by gonostylus stem base width (1.4‒1.6 versus 0.7‒1.2) and the front leg tarsomere 4 is much longer than tarsomere 5 (ratio 1.9‒2.1 versus 1.2‒1.6).

Procladius curtus Roback, 1971 View in CoL ( Roback 1971) found in the United States appears at couplet 22 in the key. P. curtus View in CoL can be distinguished from the similar P. choreus View in CoL and P. culiciformis View in CoL by a shorter gonostylus process (GspR 0.07‒0.09 versus 0.13‒0.25) and short front leg tibia compared with the first tarsomere (LRI 0.83‒0.85 versus 0.68‒0.78).

Procladius deltaensis Roback, 1987 View in CoL ( Wrubleski & Roback 1987) found in Canada appears at couplet 24 in the key. P. deltaensis View in CoL can be separated from the similar P. bellus View in CoL and P. nudipennis View in CoL by a shorter antenna (AR 0.9‒1.1 versus 1.4‒2.1) and shorter gonostylus length in relation to the gonocoxite length (0.35‒0.39 versus 0.42‒0.59).

Procladius desis Roback, 1971 View in CoL ( Roback 1971) found in Canada and the United States will appear at couplet 9 or 14 in the key. P. desis View in CoL can be distinguished from all species of Procladius View in CoL in Europe by measurements of three gonostylus characters (GspR 0.09‒0.16, process length/width 0.3‒0.6, GsmR 4.8‒5.3) Measurement data of three studied specimens from Canada are available in Supplement 1. BIN BOLD:AAG3919 is P. desis View in CoL .

Procladius jeris Roback, 1971 View in CoL ( Roback 1971) found in Alaska in the United States will appear at couplet 7 in the key. The gonostylus of P. jeris View in CoL is similar of that of P. tatrensis View in CoL (GspR 0.40‒0.42 versus 0.35‒0.43), but the body of P. jeris View in CoL is smaller (wing length 3.5‒4.5 mm versus 2.9‒3.1 mm) and the caudolateral ends of the nine tergite much more produced. The status of the species is uncertain as only one specimen is known, and the holotype is probably lost. It might be a form of P. tatrensis View in CoL .

Procladius paragretis Roback, 1971 View in CoL ( Roback 1971; Roback 1980) found on Greenland and Alaska in the United Sates will appear at couplet 7 in the key. P. paragretis View in CoL can be distinguished from the rather similar P. tatrensis View in CoL by a longer (GspR 0.46‒0.48 versus 0.35‒0.43) and more divergent gonostylus process (50‒60° versus 25‒50°), and broader gonostylus (GsmR 5.0‒5.2 versus 5.5‒6.5). The holotype has been studied within the present project, cf. measurement data in Supplement 1. The drawing of P. paragetis in Roback (1980) shows a defect gonostylus process. BIN BOLD:AAG5430 is probably P. paragretis View in CoL .

Procladius prolongatus Roback, 1971 View in CoL ( Roback 1971; Roback 1980) found in Canada, will appear at couplet 4 in the key. P. prolongatus View in CoL is easily separated from the European species with long medioapodeme by several characters including its comparatively thin, strongly diverging (70°) and moderately long gonostylus process (GspR 0.27‒0.29). The only specimen, assigned as holotype (CNCC), has been studied within the present project, cf. measurement data in Supplement 1.

Procladius reidi ( Freeman, 1955) View in CoL ( Freeman 1955; Cranston & Judd 1989; Saether & Andersen 2000), found in Guinea, Sudan and Ghana in Africa and Saudia Arabia in Asia appears at couplet 24 in the key. P. reidi View in CoL agrees with P. nudipennis View in CoL in the key but can be distinguished by the pattern of the abdominal tergites II‒VII (dark median longitudinal band versus longitudinal band absent), shorter wing (1.5‒1.6 mm versus 1.8‒2.4 mm) and shorter mid leg tibia (0.62 mm versus 0.82‒1.04 mm).

Procladius sublettei Roback, 1971 View in CoL ( Sublette 1964; Roback 1971; Roback 1980; Murray & Fittkau 1989; Saether 2010) found in the United States and Canada will appear at couplet 6 in the key. P. sublettei View in CoL is very similar to P. signatus View in CoL but lacks a basal inner lobe (inferior volsella) of the gonocoxite. It is usually smaller (wing length 2.3‒2.9 mm versus 2.8‒3.9 mm) and mostly with a lower antennal ratio (AR 1.7‒2.0 versus 1.9‒2.5 mm).

Procladius tridentus Sanyal & Hazra, 2016 View in CoL ( Sanyal & Hazra 2016) found in India, will appear at couplet 21 in the key. P. tridentus View in CoL is distinguished from all European Procladius View in CoL by its small size (wing length 1.4‒1.5 mm, body length 2.6‒2.7 mm), and from the rather similar P. choreus View in CoL also by a shorter gonostylus process (GspR 0.05‒0.07 versus 0.13‒0.20).

Procladius wilhmi Roback, 1966 View in CoL ( Roback 1966a; Roback 1971; Roback 1980) found in the United States appears at couplet 6 in the key. P. wilhmi View in CoL is easily separated from the European species that also have a long medioapodeme by several characters including its short gonostylus process (GspR 0.11‒0.14).

Procladius sp. 8 XL, collected in China is probably a new species to science. There are ten good genitalia photos in BOLD. The wing length of males is shorter than that of all studied Procladius View in CoL in Europe. P. sp. 8XL will appear at couplet 19 in the key and can be distinguished from its probably closest relative P. choreus View in CoL by its very slender gonostylus (GsmR 6.8‒8.1 versus 4.9‒5.6), short wing length (1.4‒1.7 mm versus 1.8‒2.8 mm) and gonostylus/ gonocoxite length (0.55‒0.60 versus 0.45‒0.55). BIN BOLD:ACB4698 with Procladius sp. 8 XL contains specimens from China, Japan, South Korea and easternmost Russia.

Geographical distribution and ecology

The literature search identified about 3 560 papers with adequate information on Procladius . The search included comprehensive information in databases, particularly BOLD ( Ratnasingham et al. 2024). Loans and contacts with museums, scientists and hobby collectors provided additional unpublished geographical and ecological information, as also information on slide labels, alcohol sample notes, and in computer files.

Procladius is reported from 135 of the about 200 countries or autonomous regions of the world ( Table 10). Canada, the United States, Finland, England, Scotland, Norway and Spain had most localities with findings. Records of Procladius could not be found from six ( Afghanistan, Libya, Mali, Mauretania, Mozambique and Venezuela) of the 42 countries with land area exceeding 640 000 km 2. There are very few studies of Chironomidae species in these countries.

Recent findings of Procladius are from New Zealand ( Krosch et al. 2022), New Caledonia ( Moubayed-Breil et al. 2021) and Vanuatu ( Strandberg et al. 2023). The possibly most isolated findings are from the Azores ( Murray et al. 2004; Raposeiro et al. 2009) in the Atlantic Ocean and Réunion ( Freeman 1958) in the Indian Ocean.

The northernmost findings of a Procladius species are from 82°N latitude on Ellesmere Island in Canada ( Oliver 1963) and the southernmost findings seems to be from 46°S in southern Argentina ( Massaferro et al. 2014).

The geographical range of Procladius choreus stands out among the European Procladius species with reports from 58 countries or autonomous regions ( Table 11), mostly from Europe including the remote Azores, but also a wide geographical belt stretching from the Canary Islands, to Egypt, Iran, India, Mongolia, Japan and the United States. P. choreus is the only of the European Procladius with quality assured findings in Africa. Other European species with wide geographical range covering Europe, Asia and North America are P. culiciformis and P. ferrugineus reported from 37 and 30 countries or autonomous regions, respectively. As much as 20 of the 27 European species of Procladius are also found in Asia and/or North America.

Table 11 furthermore shows that six of the Procladius species of Europe reach above 2 000 meters altitude. P. tatrensis has the highest record of 2 670 m and might reach 2 800 m in the Alps ( Heiri & Lotter 2008). Worldwide it is surpassed by P. brevipetiolatus which reaches altitude 3 500 m in Uganda ( Eggermont & Verschuren 2007) and an unknown species of Procladius at 4 730 m in Himalayan China ( Echeverría-Galindo et al. 2023). None of the European Procladius species is only confined to high altitude. All, but for P. fimbriatus with lowest record at 410 m, occur at altitudes of less than 80 m.

The coldest conditions with findings of Procladius are on Ellesmere Island of northernmost Canada ( Oliver 1963) and the New Siberian Islands of Russia ( Lundström 1915; Krasheninnikov 2013; Nazarova et al. 2015) with mean annual temperature of about ‒14 to ‒18 °C. Coldest conditions known for Procladius in Europe is ‒8 °C mean annual temperature prevailing during the 1920s on the Svalbard islands of northernmost Norway ( Edwards 1924; Stur & Ekrem 2020).

As many as 8 of the 27 identified species of Procladius in Europe have been found in areas with annual mean annual temperature as low as ‒10 to ‒18 °C ( Table 11). P. clavus ( Roback 1971) , P. exilis and P. frigidus ( Lundström 1915; Oliver 1963; Saether 2004) reach the coldest records. Four other species of Procladius ( P. desis , P. jeris , P. paragretis and P. prolongatus ), all from North America, have also been found at sites with mean annual temperature from ‒10 °C or colder. This means that Procladius , with 12 species, is one of the most species-rich genera of aquatic insects in extremely cold conditions, such as in Canada ( Oliver 1963; Danks 1992; Pentinsaari et al. 2020), Greenland ( Edwards 1931; Böcher et al. 2015), Russia ( Lundström 1915; Saether 2004; Coulson et al. 2014) and Alaska of the United States ( Butler et al. 1980; Butler 1982; Lougheed et al. 2011). All species in Europe can thrive in areas with a mean annual temperature below +6 °C and are thus able to withstand ice cover on lakes or smaller water bodies.

Warmest conditions with records of Procladius worldwide are in Niger ( Alhou et al. 2012) and the United Arab Emirates ( Reeves & Epler 2016) with mean annual temperature about +29 °C. Table 11 shows that only P. choreus of the European species reach mean annual temperature above +20 °C, with the highest record at +25 °C in Saudi Arabia ( Cranston & Judd 1989).

Six species of Procladius species found in Europe have temperature ranges of more than 20 °C, with the most extended range noted for P. lugens ( Table 11). All species but four have ranges of 10‒20 °C. Among these, three species, P. fimbriatus , P. gemma and P. saeticubitus , might be rare as they have been recorded from less than eleven localities each.

Most reports of Procladius species are from lentic freshwater bodies of 0.01 km 2 or more, thus lakes and reservoirs. Records are rather frequent from other standing freshwater such as springs, ponds, puddles, marshes, swamps, bogs and human constructions such as rice fields, sewage water treatment plants, sewage ponds and rainwater barrels. Larvae of Procladius also frequently inhabit salty conditions such as estuaries, brackish sea water and even salt lakes with salinity higher than that of the sea. They are, however, not known from true marine environments.

All 27 European Procladius species have been found in lakes and reservoirs, most of them also in other kinds of water ( Table 12). Brackish and slightly marine water have representatives of nine species of European Procladius , while moderately fast running water only three.

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All 25 species with water depth notes have been found in shallow water between 0 to 2 m, while findings beyond 100 m are only reported for P. lugubris and P. culiciformis , of which the latter has a probable record from 206 m in Lake Vierwaldstätter in Switzerland ( Zschokke 1905).

Table 12 furthermore shows that most Procladius species, as most other species of Chironomidae , have a wide adaptability to different trophic levels in lakes and reservoirs encompassing oligotrophic, mesotrophic and eutrophic conditions ( Saether 1979; Ruse 2010). No Procladius species is, however, noted for all five categories of trophic conditions. Five species which mainly or solely inhabit lakes in northern Europe are only found in ultraoligotrophic or oligotrophic conditions, thus with low levels of available nutrient for primary production.

Species of Procladius have proven useful as indicators of environmental problems caused by human activity, including eutrophication ( Zinchenko 1992; Ruse 2015; Takamura et al. 2021), contamination by heavy metals or toxic organic compounds ( Pettigrove 1989; Warwick 1991; Aliyev et al. 2013), climate-change effects ( Engels et al. 2019; Rigterink et al. 2022, Brodin & Hellberg 2023) or biological diversity changes ( Alhou et al. 2012; Bista et al. 2017; Gadawski et al. 2022). Indicator usefulness of Procladius is however often hampered by the fact that identification could only be achieved to the genus level.

Species of Procladius are generally not among those reported to cause trouble for humans, e.g. because of mass-development or allergy problems. An unidentified species of Procladius is suspected to be a vector of cholera ( Maheshwari et al. 2010).

The species; systematics, distribution and ecology

The 27 European species are presented in alphabetical order. After the heading with the species name and author follows in chronological order a list of Latin names, including synonyms, with pertinent information on literature, countries or autonomous regions with findings, life stages and available keys, illustrations and photos. At the end of the list, a question mark in front of the name indicates questionable synonym.

Material examined is presented in the following way and order; 1) in alphabetic order, country or autonomous region in capital letters, 2) numbers and life stage, 3) within parentheses (type-status if it is a type, if another name is assigned to the specimen, code of four letters of the museum or institution where the specimen is deposited) ( Table 13), 4) site with WGS84 geographic coordinates, 5) altitude in meters above sea level abbreviated m a.s.l., 6) mean annual temperature abbreviated m.a.t. of the locality of the year of sampling, 7) date of sampling, 8) leg. collector name, 9) within brackets [barcode code]. Several specimens, sometimes also types, are in private collections and don’t have a museum or institution code.

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Diagnostic characters: Reference to figures of the species and couplet of appearance in the key above are given. The most important morphological information to distinguish the species in question from those most similar ones is highlighted. Notes on taxonomy and the existence of barcodes are also provided. The helpdesk above contains twenty-one characters useful to distinguish the species from other species. Supplement 1 serves as a comprehensive description of the species based on one hundred characters.

Geographical distribution and ecology: A synthesis based on important information from published literature and other sources including unpublished works and information only on slides, pins or of alcohol samples is given. Most of the information is also available in Table 11 and 12. Only information from sources where the species in question can be reliably identified is considered.

References: This part contains a list of published scientific literature or other works with important information about the species in question, provided that species identification can be regarded as reliable, e.g. done be an expert on Chironomidae taxonomy. The species in question is identified with another name or only as Procladius sp. in several works, but the species identity is revealed by drawings, photos, specimen loans or correspondence by email.