Rueda-Ramírez, Diana ¹ ; Krantz, Gerald W. ² ; Palevsky, Eric ³ and Ruess, Liliane ⁴

¹Institute of Biology, Ecology Group, Humboldt-Universität zu Berlin, Philippstraße 13, 10115 Berlin, Germany.
²Integrative Biology, Oregon State University, Corvallis, OR 97331, USA.
³Newe Ya’ar Research Center, Agricultural Research Organization (ARO), Israel.
⁴Institute of Biology, Ecology Group, Humboldt-Universität zu Berlin, Philippstraße 13, 10115 Berlin, Germany.

2025 - Volume: 65 Issue: 2 pages: 417-447

ZooBank LSID: C8348491-7B0B-405E-A1F4-7358ED45CE75

Original research

Keywords

Abstract

Introduction

Macrocheles Latreille, 1829 is the largest and most diverse genus of the gamasine mite family Macrochelidae, comprising more than 300 described species of the approximately 500 species presently recognized in the family (Azevedo et al. 2015; Beaulieu et al. 2011; Emberson 2010). The great preponderance of known Macrocheles species exploit temporary, often insular organic substrates such as animal or plant dung or the nests of small mammals, birds, or social insects, feeding largely on nematodes, enchytraeid worms and microarthropods that they encounter there. When their ephemeral habitat deteriorates or desiccates, most of these species utilize phoretic attachment of adult females to co-occurring flying insects to carry them to fresh substrates (Krantz 1998; Krantz 2009; Lindquist, Krantz and Walter 2009).

The great diversity displayed by Macrocheles has led to the recognition of several species groups in the genus, based largely on shared morphological traits. However, the classification of these groups remains poorly defined and incomplete. Among the first of these to be established was the glaber group, proposed by Filipponi and Pegazzano (1962) and including M. glaber (Mueller), M. perglaber Filipponi and Pegazzano, and M. scutatus (Berlese). Walter and Krantz (1986) further divided an expanded glaber group into subgroups, with the scutatus subgroup being distinguished by a reduction in overall size and a distinctive decrease in sclerotisation of the ventral shields. Later, Walter and Krantz (1992) recognized four species complexes in the scutatus subgroup, differentiating them on morphological and distributional grounds. The scutatus complex, with seven described species, is the largest of these assemblages.

Species of the glaber group, and particularly the scutatus subgroup, are known for their close association with scarabaeoid beetles and the dung they visit (Krantz 1983; Walter 1985; Walter and Krantz 1986). Members of the subgroup have received special attention as potential biological control agents against flies breeding in these habitats (Krantz 1983; Azevedo et al. 2015). Predation by glaber-group mites on free-living nematodes (FLN) is believed to promote mite fecundity in such environments (Rodriguez, Wade and Wells 1962; Filipponi and Dojmi di Delupis 1963; Ito 1973; Azevedo et al. 2017; Azevedo et al. 2018). Fertility enhancement of mites by nematodes being a part of their diet has been thought to promote more efficient levels of biological control (Rueda-Ramírez et al. 2023).

During the course of a German Research Foundation project (DFG Grant RU 780/20-1) aimed at promoting the indigenous soil food web (microorganisms, FLN, and predatory mites) for the biological control of root-knot nematodes (Meloidogyne spp.), specimens of a scutatus subgroup species of Macrocheles were frequently collected from nematode-infested vegetable field soil in Israel. Mite specimens from different collection sites were used to initiate laboratory colonies that were fed primarily on FLN species. The mites also were found to feed readily on Meloidogyne incognita (Kofoid & White, 1919) Chitwood, 1949 (Rueda-Ramírez et al. 2023), a serious nematode pest of diverse cultivated crops worldwide (Moens, Perry and Starr 2009).

Morphological examination of the mites from Israel revealed a strong resemblance to M. scutatus (Berlese 1904), but with discreet differences in female sternal shield ornamentation and in male ventral shield shape, leg armature, and cribral development. Herein, we propose and describe a new Macrocheles scutatus-subgroup species, Macrocheles karkomensis n. sp. Rueda-Ramírez & Krantz. The main objectives of this paper are to describe the new species, update the existing dichotomous key to the scutatus complex (Walter and Krantz 1992), and provide information on DNA barcoding of related taxa based on COI data.

Material and methods

Mite collection

Specimens used in this study were obtained from laboratory colonies maintained by the Ecology Group, Institute of Biology, Humboldt-Universität zu Berlin (Germany), and from the Department of Entomology, Newe-Ya′ar Research Center, Agricultural Research Organization, Raman Yishay (Israel). The colonies were initiated with females extracted from soil samples collected by Eric Palevsky from two greenhouses cultivating organic cucumber (Cucumis sativus L.), and three open-field plots of organic lettuce (Lactuca sativa L.) (Table 1).

Samples were collected with a core sampler (5 cm diameter and 5 cm depth). Soil was removed from the cores, gently mixed and then poured onto a sieve (6 cm high and 10 cm in diameter) to extract the mites using a Galen-Berlese funnel (Rueda-Ramírez et al. 2022). Extraction was carried out for ~ 8 days, starting immediately after sample collection. Mites were collected in 7 cm plastic cups (microcosm) whose floors were covered with a mixture of nine parts gypsum and one part activated charcoal. Colonies were initiated after separating and confirming that all of the specimens represented the new species. Some colonies were kept separate based on the localities in which mites were collected, while other colonies consisted of mixed populations. Each colony was maintained in plastic units as described above. Each microcosm was filled to half capacity with vermiculite, which was kept moist by daily addition of sterile water. The mites were fed with a mixture of different free-living nematode species and, on some occasions, with a mixture of all developmental stages of the oribatid mite, Carpoglyphus lactis (Linnaeus, 1758) (Astigmatina: Carpoglyphidae).

SEM examination

Mites taken directly from colonies were fixed and maintained for at least 24 hours in 70% ethanol. Subsequently, specimens were dehydrated in ethanol solutions of increasing concentration (80%, 90%, 95%, 100%), changing the concentration at least every 15 minutes. The samples were then critical point dried in a Balzers Union CPD 030, coated with gold on a Balzers Union SCD 040 sputter coaster, and visualized with a LEO 1430 scanning electron microscope (Carl Zeiss AG, Jena, Germany).

DNA barcoding

Eighteen mite specimens were sent for molecular analysis to the Canadian Centre for DNA Barcoding (CCDB). The barcode region of cytochrome c oxidase I (COI) was sequenced with a cocktail (1:1 ratio) of LepF1/LepR1 (Hebert et al. 2003) and LCO1490/HCO2198 (Folmer et al. 1994) primers using standard protocols modified to allow voucher recovery following DNA extraction (Ivanova, de Waard and Hebert 2007; Ivanova and Grainger 2007a; Ivanova and Grainger 2007b; Porco, Bedos and Deharveng 2010). DNA extracts were archived in −80 °C freezers at the Centre for Biodiversity Genomics (CBG; biodiversitygenomics.net), and the vouchered specimens were retained in 96-well microplates with 95% ethanol for subsequent morphological preparations.

COI sequences were assembled from forward and reverse chromatograms using CodonCode Aligner v. 4.2.7 and uploaded to BOLD. They were inspected for potential contamination or misidentification by examining their placement in a Neighbor-Joining tree and by querying each record against BOLD's complete reference library using the BOLD Identification Engine. To assess the COI sequences similarity between the species described here and other Macrocheles species, we followed the procedure and short phylogeny published by Knee (2017). We used the same COI sequences as Knee (2017), except that instead of seven sequences from Macrocheles willowae Knee, 2017, we used only three. Similarly, two species of Ornithonyssus (Macronyssidae), Ornithonyssus bacoti (Hirst, 1913) and O. sylviarum (Canestrini & Fanzago, 1877), were used as outgroup. In addition, we included two unidentified Macrocheles species that were recognised as closely related to the new species described here based on comparisons with the BOLD and GenBank databases (Table 2). Only sequences with verifiable morphological identity were used.

Sequence alignment, pairwise distance calculation, determination of the best-fit model of molecular evolution and phylogenetic reconstruction followed the procedures described by Knee (2017), with modifications only to adjust the latest versions of the software. Briefly, sequence alignments were performed based on their translation into proteins in Mesquite version 3.81 (Maddison and Maddison 2023). Pairwise distances (p-distance) were calculated using neighbour-joining (NJ) analysis with the Kimura 2-parameter (K2P) model in PAUP* v4.0a (Swofford 2003). The best-fit model of molecular evolution was determined to be GTR+I+G using MrModeltest v2.4 (Nylander 2004). Phylogenetic reconstructions of the COI sequences were performed using Bayesian inference (BI) in MrBayes v3.2.7a (Huelsenbeck and Ronquist 2001). In our case, the set of COI sequences was run for 10 million generations in two runs, producing 20002, of which 19502 were sampled for the consensus tree. The consensus tree was constructed in MrBayes and visualised with FigTree v 1.4.4 (Rambaut 2018), with the posterior probability at each node of the tree.

Mite identification and description

Morphological identification was based on examination of 30 mites, including the 18 specimens from which DNA had earlier been extracted. All of the specimens were mounted on microscope slides using Hoyer's mounting medium. Measurements are based on six females (two of which underwent DNA extraction), six males, three deutonymphs, and three protonymphs. Mites were imaged with a digital AxioCam MRc5 connected to an Axiophot Zeiss microscope. Photos and accompanying illustrations were then processed with a digital tablet using the Adobe Illustrator® program (version 21.0.0, 2017).

Measurements were taken with ZEN 2012 software (version 1.1.1.0). Measurements of each structure are given in micrometres (µm), with the average measurement for the specimens examined followed (in parentheses) by respective size ranges for variable structures.

Shield length is the maximum distance between anterior and posterior margins, whereas shield width generally refers to the maximum width. Sternal shield width is the width at the median level of coxae II. Leg lengths are based on distances between coxal bases and tarsal termini. Idiosomatic setal nomenclature is based on Lindquist and Evans (1965), leg chaetotaxy on Evans (1963; 1969), and poroidotaxy on Athias-Henriot (1975), revised and complemented by Moraza (2025). In this work, we designate two previously unnamed lyrifissures with the abbreviation ′c′ which comes from the Latin ′canalis′ meaning ′groove′ and refers to the location of the lyrifissures within the groove of the peritreme. In leg ornamentation, the term ''tubercle'' refers to a protuberance on the cuticle which bears a seta, and the term ''spur'' to a protuberance without a seta. The number of teeth indicated for cheliceral digits does not include the terminal hook.

Abbreviations of sternal shield lines are as follows:

l.ang.: linea angulata, an anterior transverse line, deeply concave medially, terminating anterolaterally at lyrifissures iv1.

l.arc.: linea arcuata, a deeply concave line whose anterior extremities approach or touch the lineae oblique anteriores (see below).

l.m.t.: linea media transversa, a median line terminating at or near the insertions of setae st2.

l.o.a.: lineae oblique anteriores, paired lines extending posterolaterally from the l.ang. through the insertions of st2.

l.o.p.: lineae oblique posteriores, paired lines extending posterolaterally from l.m.t. through lyrifissures iv2.

a.p.: areae punctatae, punctate areas along posterior margin of shield.

Voucher specimens of M. karkomensis are deposited in the Museum für Naturkunde (MfN), Berlin, Germany, the Hebrew University of Jerusalem, National Collections of Natural History (HUJ), Jerusalem, Israel and Oregon State Arthropod Collection (OSAC), Corvallis, Oregon, USA. DNA vouchers are deposited in the Centre for Biodiversity Genomics (CBG), Guelph, Ontario, Canada. Type depositories of the new species are indicated in the ''Material examined'' section.

Results

Macrocheles karkomensis Rueda-Ramírez & Krantz n. sp.

ZOOBANK: A7FE0431-8346-491E-9D3A-59651C492D41

Diagnosis

Female dorsal shield with a weak procurved line posterior to z6 and s5, absent in some specimens; bearing 28 pairs of setae, all aciculate simple, except j1, which are appressed and distally pilose, j4 and z4, distally pilose and J5, weakly serrate; sternal shield longer than wide, with ill-defined faint punctate lines: l.m.t. undulate, l.arc. strongly procurved and almost touching l.o.a., l.o.p. barely visible, without ramus and free from l.m.t., l.ang. procurved, and a.p. reduced to a few small scattered punctae; epigynal shield bearing a pair of accessory sclerites and st5; opisthogaster with six pairs of setiform setae (Jv4 – 5, Zv1, Zv2, Zv4 and Zv5) on unsclerotised cuticle; with a pair of elongated metapodal plates; ventrianal shield longer than wide, punctate-reticulate. Male with cheliceral spermatodactyl curved downward, extending beyond the base of the dorsal seta and progressively tapering in diameter to a blunt tip; dorsal shield bearing 31 pairs of setae (the same as those present in the female, plus r5, R1 and R2), all aciculate simple, except j1, j4 and z4 which are distally pilose, J5 weakly serrate; sternogenital shield separate from ventrianal shield, which is peltate, slightly broader than long, trochanter IV without spurs; femur IV with three tubercles, the most proximal of which with a terminal small spiniform seta (v1); genu IV with av1 on a barely visible tubercle.

Adult female

(Figures 1 – 5, six specimens measured).

Gnathosoma — Anteromedian region of epistome (Figure 1a) with a median with a median, distally bifurcate element flanked by pair of shorter, broader flag-like elements, whose bases are joined to weakly serrate epistomatic margins. Deutosternal groove medial, longitudinal (Figure 1b), located partially in the anterior hypostome (distal to insertions of hypostomatic setae h2–3 and adjacent palptrochanteral junctions) and partially in the more posterior basis gnathosomatica (Evans & Till 1979, Bowman 2023); with six transverse rows of denticles delimited laterally by subparallel grooved borders of deutosternal field, row 1 located anteriorly on the hypostome and weakly ornamented, rows 2–6 on the basis gnathosomatica, each with 10–15 distinct denticles. Internal malae (Figure 1b, im) narrow, paired, extending anteriorly almost to tips of adjacent corniculi external margins lightly fimbriate. Corniculi (Figure 1b, co) elongate, horn-shaped, over twice as long as their basal width, each with a closely associated flanking salivary stylus (sst) originating posterior to insertion level of hypostomatic setae h1 (Figure 1b). Hypostomatic and capitular setae (h1–3, pc) smooth, aciculate, insertions of h3 mesad and adjacent to those of h2, capitular setae shorter than hypostomatics, flanking denticular row 6. Subcapitular setal measurements: h1 47 (46–48), h2 26 (23–31), h3 67 (61–73), pc 21 (19–22). Chelicerae with a long internal arthrodial brush reaching the mid-level of movable digit and a short external arthrodial brush, both arising ventrally at the articulation of the movable digit. Fixed cheliceral digit 69 (66–71) long, with an apical hook, a subterminal recess fitting the tip of the movable digit when closed, and where two tiny teeth are also found, with a more proximal spine-shaped pilus dentilis and a large tooth; dorsal cheliceral seta thick, dorsal lyrifissure longitudinal, internal and anterior to the dorsal seta, antiaxial lyrifissure external at the level of the arthrodial brushes; movable digit 60 (58–63) long, with an apical hook, a large, bidentate tooth flanked distally by two small proximal teeth (Figure 1c,d). Number of setae on palp trochanter–tarsus: 2-5-6-14-15, aciculate and smooth, except al of femur and al1 and al2 of palpgenu, which are stout and spine-like (Figure 1b); palptrochanter with outer seta (pv) shorter than inner seta (av); tarsal apotele 3-tined (Figure 1c).

Idiosoma — Length 654 (600–707), width 455 (363–528).

Dorsal idiosoma — (Figure 2, 3). Holodorsal shield, 590 (571–623) long and 357 (335–376) wide at level of dorsal setae r3, ovate-reticulate, reticulation less evident in sigillate region of setae z5–s6, usually with a weak procurved line posterad z6-s5, line occasionally absent. Holodorsal shield (Figure 2) bearing 28 pairs of setae, 16 pairs of lyrifissures and seven pairs of pores; shield setae smooth, aciculate except j1, which are appressed, distally flattened and pilose (Figure 3a), and j4 and z4, which are distally pilose, J5 lightly pectinate; distances j2–j3 49 (42–52), j4–j4 102 (99–107), j5–j5 54 (52–56), J5–J5 27 (26–27). Unsclerotised dorsal cuticle lateral and posterior to shield with 13 pairs of setae (r5, R1–6, UR 1–6) and lyrifissure idRp. Setal measurements are shown in Table 3.

Ventral idiosoma — (Figure 4). Tritosternum with columnar base, 33 (31–35) long and 17 (15–18) wide, with long pilose laciniae 96 (87–103), separated for ~92% total length. Sternal shield 150 (144–154) long and 121 (118–128) wide at level of coxae II, with faint punctate ornamentation (Figure 4, 15a): l.m.t. undulate, l.arc. strongly procurved and almost touching l.o.a., l.o.p. barely visible with weak ramus or without ramus and free from l.m.t., l.ang. procurved, and a.p. reduced to a few small scattered punctae; with three pairs of setae (st1–st3) and two pairs of lyrifissures (iv1, slightly posteriad and laterad st1, and iv2, posteriad and laterad st2); distances st1–st3 143 (138–151), st2–st2 104 (101–112), st3–st3 108 (103–117); shield with broad anterolateral corners extending between coxae I and II, distally abutting anterior exopodal. Setae st4 and lyrifissure iv3 on metasternal platelets. Epigynal shield bearing st5, abutting a pair of accessory sclerites, ornamented with four weak, granulate recurved lines; shield 133 (122–144) long, 131 (117–152) wide at widest level, truncate posteriorly; distance st5–st5 115 (105–124), posteriorly truncate; iv5 on unsclerotised cuticle, posterolaterad st5. Metapodal platelets elongate, somewhat irregular. Ventrianal shield subcordate, reticulate; shield 203 (189–232) long and 185 (171–197) wide at widest level; with three pairs of preanal setae (Jv1, Jv2 and Jv3), a pair of lyrifissures posteromesad Jv2, cribral glands gv3 on posterolateral shield margins and posterior to paranal setae; anal opening 26 (24–29) long and 20 (18–21) wide; cribrum narrow, confined to posterior margin of the shield. Opisthogastric unsclerotised cuticle with six pairs of setae (Jv4, Jv5, Zv1, Zv2, Zv4 and Zv5), three pairs of lyrifissures, gv2 pore posterolaterad coxa IV. Endopodal and exopodal elements as illustrated in Figure 4.

Anterior section of endopodal shield contiguous with sternal shield, with section between coxae III and IV boomerang-shaped. Exopodal shield free, fragmented into two pieces, the anterior surrounding acetabula II–III and the posterior (parapodal) an arch surrounding acetabulum IV. All ventral setae setiform. Setal measurements shown in Table 4.

Peritreme and peritrematic plate — (Figures 2a, 3 and 4). Peritreme extending anteriorly to level of dorsal lyrifissure id1, with two pairs of lyrifissures (ipc1, ipc2) on or within peritrematic groove, with ipc2 ventral at level of coxae III (Figure 4) and ipc1 anterodorsal at level of setae z2 (Figure 2a); peritrematic shield fused with dorsal shield at level of r2–r3, free and recurved posteriorly, extending no further than anterior margin of coxae III, bearing a pore (gvi) and two lyrifissures (ips and ivi) at the level of each stigma, with pore gd3 and lyrifissure ip dorsad of the peritreme and between coxae II–III.

Spermathecal apparatus — Not clearly seen in available specimens.

Legs — (Figure 5). Tarsi II–IV with claws and ambulacra; tarsus I without pretarsus, claw and ambulacrum. Leg lengths: I: 459 (427–486.2); II: 405 (390–425); III: 348 (335–365); IV: 525 (506–544). Pretarsus lengths: II: 32 (30–36); III: 31 (28–36); IV: 32 (32–33). Setation (legs I–IV): coxae: 0-0/1 0/1-0, 0-0/1 0/1-0, 0-0/1 0/1-0, 0-0/1 0/0-0; trochanters: 1-0/1 0/2-1, 1-0/1 0/2-1, 1-0/2 0/1-1, 1-1/2 0/1-0; femora: 2-3/1 2/3-2 (ad2 and ad3 thickened and distally pilose), 2-3/1 2/2-1 (ad1 thickened and distally pilose), 1-2/1 1/0-1, 1-2/1 1/0-1 (ad2 and pd1 thickened and distally pilose); genua: 2-3/1 2/1-2, 2-3/1 2/1-2, 1-2/1 2/0-1 (pd2 small spine-like), 1-2/1 2/0-0; tibiae: 2-3/2 2/1-2 (pd2 thickened and distally pilose), 2-2/1 2/1-2, 1-1/1 2/1-1, 1-1/1 2/1-1; tarsi: not counted, 18 (distal setae thickened), 18 (ventral setae thickened), 18 (ventral setae thickened).

Adult male

(Figures 6 – 9, five specimens measured)

Gnathosoma — Epistome, deutosternal groove, position and shape of hypostomatic setae, internal malae, and corniculi as in adult female. Subcapitular setal measurements: h1 45 (39–49), h2 16 (14–18), h3 61 (55–64), pc 15 (13–17). Fixed cheliceral digit 49 (45–54) long, with blunt apical hook, followed proximally by three small teeth and a large, weakly bidentate tooth; spine-shaped pilus dentilis inserted slightly anteriad the large tooth; movable digit 42 (36–47) long, with a large tooth behind terminal hook; spermatodactyl 49 (45–54) long, curved upward and posteriorly, extending beyond insertion of dorsal seta, tapering progressively distally to a blunt tip; dorsal seta thick; dorsal and antiaxial lyrifissures distinct, the former with longer axis longitudinal, on internal face of digit (Figure 6). Arthrodial process of chelicera, palp chaetotaxy, and apotele as in adult female. Palpal setae aciculate and smooth, except for stout and spinose al setae on the femur and al1 and al2 on the genu.

Idiosoma — Length 501 (458–534), width 319 (281–358).

Dorsal idiosoma — (Figure 7). Holodorsal shield, 493 (450–523) long and 311 (278–340) wide, virtually covering dorsal surface of idiosoma, narrowing slightly posteriad r5, with ornamentation similar to adult female, less evident, between j3-j4-z2 and j5-j6-z5; shield with 31 pairs of setae (the same as those of female, plus r5, R1 and R2;) distances j2–j3 41 (37–43), j4–j4 87 (83–90), j5–j5 44 (41–46), J5–J5 22 (20–23). Poroidotaxy, adenotaxy and shape of setae as in adult female. Setal measurements shown in Table 3.

Ventral idiosoma — (Figure 8). Tritosternum similar to that of adult female; base 21 (18–23) long and 14 (13–14) wide, and long pilose laciniae 84 (80–89), separated for about 94% of their total length. Sternogenital shield with anterior half reticulate, with punctate-reticulate ornamentation; posterior half with granulate ornamentation and scant wavy lateral lines oriented more or less longitudinally; 217 (199–227) long and 92 (85–98) wide at the level of coxae II; with five pairs of setae (st1–3, 5; st4 absent on at least one side in most observed specimens, sometimes completely absent; figure 8b,c), with three pairs of lyrifissures (iv1–3); distances st1–st3 132 (126–135), st2–st2 80 (78–82), st3–st3 81 (75–85). Endopodal shield fused with sternogenital shield. Ventrianal shield peltate, without anterolateral excavations around coxae IV, ornamentation similar to that of anterior half of sternal shield, reticulation absent anteriad anal opening; slightly broader than longer; 171 (153–181) long and 181 (165–190) wide at widest level; with six pairs of setae (Jv1–4, Zv1 and Zv2) in addition to circumanal setae, iv5 lyrifissures at anterior corners of shield, with additional pair lyrifissures posterolaterad to Zv1 and cribral glands gv3 at shield margin, slightly anteriad level of postanal seta; anal opening 20 (18–22) long and 19 (18–19) wide. Cribrum wider than in female and with a pair of strap-like paranal extensions on either side of postanal seta. Metapodal plates elongate, irregular. Opisthogastric unsclerotised cuticle with a pair of setae (Jv5), three pairs of lyrifissures, and gv2 pore posterolaterad coxae IV. Exopodal shield surrounding acetabula II–IV. Idiosomatic setae similar to those of adult female. Setal measurements shown in Table 4.

Peritreme and peritrematic plate — (Figure 7, 8). Peritreme similar to that of female lyrifissures anterodorsally expressed ipc1 (Figure 7), and ventral ipc2 (Figure 8). Peritrematic plate fused with dorsal shield for its entire length; bearing pore (gvi) and two lyrifissures (ips and ivi) in close association with stigmata, with pore gd3 and a lyrifissure ip dorsad of peritreme and between coxae II-III.

Legs — (Figure 9a–c). Leg lengths: I: 416 (390–439); II: 378 (350–399); III: 321 (302–338); IV: 491 (447–528). Pretarsus lengths: II: 29 (27–32); III: 25 (21–26); IV: 29 (26–31). Setae aciculate; femur II bears a thick spur ventrally between pv1 and pv2, and with setae ad1 and pd1–2 somewhat thickened and distally pilose; ad3 absent. genu and tibia II with av1 represented by small triangular spur (Figure 9a); genu II with setae ad2, ad3, pd1, and pd2 distally pilose. Podomeric fissure between basi- and telofemur of leg III (Figure 9b) with series of small, rounded ventral excrescences; ad1 of femur III, ad1, ad2, pd1 and al1 of genu III, ad1, pd1, pd2, al1 and pl1 of tibia III, and ad4 and pd4 of genu III distally pilose; v1 of femur III thickened, distally pilose (Figure 9b). Femur and telotarsus of legs IV (Figure 9c) strongly ornamented with spurs and tubercles: femur IV with large, ventral curval setiferous tubercle carrying a small spiniform setae (v1) and two smaller adjacent spurs as illustrated, ad1 and al1 slightly thickened, distally pilose, ad2 and pd1 strongly thickened, distally pilose; genu IV with av1 on barely visible tubercle and with ad1, ad2, pd1, pd2 and al1 distally pilose; tibia IV with ad1, pd1, pd2, al1 and pl1 distally pilose; tarsus IV with a rounded dorsal spur distally, ad4, pd3–4 distally pilose (Figure 9c).

Deutonymph

(Figure 10 – 11, three specimens measured).

Gnathosoma — Anteromedian region of epistome, deutosternal groove, position and shape of hypostomatic setae, internal malae, and corniculi as in adult female. Subcapitular setal measurements: h1 37 (36–38), h2 13 (11–15), h3 61 (60–63), pc 18 (18–19). Fixed cheliceral digit 45 (44–46) long, with subapical tooth, a spine-like pilus dentilis, and two adjacent teeth; movable digit 41 (40–43) long, with two large teeth medially; dorsal seta thick, dorsal and antiaxial lyrifissures distinct. Arthrodial process of chelicera, corniculus, palp chaetotaxy, and apotele similar to those of adult female. Palpal setae aciculate and smooth, except for thickened al on femur and al1 and al2 on genu.

Idiosoma — Length 639 (614–664), width 421 (418–423).

Dorsal idiosoma — (Figure 10). Dorsal shield with deep, curved lateral incisions behind insertions of s6 and approaching those of z6; shield 457 (454–460) long and 263 (262–265) wide, covering only central portion of idionotum, tapering behind S2 and with weak punctate ornamentation posteriorly. Shield setae, lyrifissures and pores as in adult female, but lyrifissure idl4 not distinguishable; distances j2–j3 35 (33–37), j4–j4 74 (71–76), j5–j5 44 (43–44), J5–J5 21. Idiosomatic setae similar to those of female. Setal measurements shown in Table 3.

Ventral idiosoma — (Figure 11). Tritosternum similar to that of adult female; base 29 (28–30) long and 15 (14–15) wide, long pilose laciniae 78 (77–78), separated for about 92% of their total length. Sternal shield lightly sclerotised, without ornamentation, tapering posteriad st3; 206 (198–214) long and 77 (73–81) wide, with four pairs of aciculate setae (st1–st4) and three pairs of lyrifissures (iv1–iv3); distances st1–3 134 (133–135), st2–st2 87 (86–87), st3–st3 85. Seta st5 and lyrifissure iv5 on unsclerotised cuticle, the latter posterolaterad st5; distance st5–st5 92 (89–95). Opisthogastric unsclerotised cuticle with eight pairs of setae (Jv1–Jv5, Zv2, Zv4 and Zv5), three pairs of lyrifissures, gv2 pore posterolaterad coxa IV and gv3 near anal shield margin, lateral and slightly posterior to postanal seta. Exopodal shield reduced to a narrow arched strip surrounding acetabulum IV (parapodal plate). Anal shield small, semicircular, lightly sclerotised and lightly reticulate, 77 (75–79) long and 73 (71–75) wide; anal opening 23 (23–24) long and 15 (14–15) wide. Cribrum wider than in female with a pair of strap-like paranal extensions on either side of postanal seta. Idiogastric setae as in adult female. Setal measurements shown in Table 4.

Peritreme and peritrematic plate — (Figure 10, 11). Peritremes extending anteriorly to level of dorsal setae z1, recurving posteriorly at level of anterior margin of coxae II; peritrematic plate lightly sclerotised, reduced to narrow strip bordering peritreme, bearing pores gp and gd3 at levels of coxae II-III respectively, and lyrifissures ipc2 at level between coxae III-IV; lyrifissure ips on peritrematic plate terminus adjacent and mesad to stigmata, gvi and ivi on unsclerotised cuticle behind stigmata.

Legs — Leg lengths: I: 379 (379–380); II: 328 (324–331); III: 288 (287–289); IV: 435 (434–436). Pretarsus lengths: II: 22 (21–22); III: 21 (21–22); IV: 22 (20–23). Setation as in adult female. Setae aciculate and smooth, except ad1, ad2 and pd1 of femora IV, ad1 and pd1 of genua IV, and al1 and pl1 of tibiae IV distally spatulate, ventral setae of tarsus II–IV thickened.

Protonymph

(Figure 12 – 13, three specimens measured).

Gnathosoma — Epistomatic margin lightly denticulate, bifurcate anteromedian extension divided only distally. Deutosternal groove, position and shape of hypostomatic setae, internal malae, and corniculi as in adult female. Setal measurements: h1 32 (31–34), h2 14 (13–15), h3 50 (49–51), pc 17 (16–17). Fixed cheliceral digit 36 (35–36) long, with two subapical teeth, at the same level of the spine-shaped pilus dentilis; movable digit 29 (28–29) long, with two teeth. Numbers of setae on palp trochanter–tarsus: 1, 4, 5, 12, 15. Arthrodial process of chelicera and apotele as in adult female. Palpal setae aciculate and smooth, except for thickened al on the femur and genu.

Idiosoma — Length 433 (428–440), width 278 (261–294).

Dorsal idiosoma — (Figure 12). With separate podonotal and opisthonotal shields. Podonotal shield 194 (191–197) long and 173 (164–182) wide; with setation similar to that of deutonymphs and adults, but without z6, s2, s6 and r2-4; with four pairs of lyrifissures (id1, id4–6), and one pair of pores (gd2); all setae aciculate simple, except j1, j4 and z4, which are distally pilose; unsclerotised cuticule laterad podonotal shield with four pairs of setae (s6, r2, r3 and r5) and a pair of lyrifissures (id2); distances j2–j3 13 (13–14), j4–j4 57 (56–59), j5–j5 42 (39–45). Opisthonotal shield 120 (114–126) long and 92 (89–94) wide, with an inconspicuous curved punctate line between setae Z4; with setation similar to that of deutonymphs and adults, but without S1 and with S2 on unclerotized cuticle, ten pairs of distinguishable lyrifissures (idm1–5, idl1 and idl3–5), and a pair of pores (gd9); all setae aciculate simple except for serrate J5 and distally pilose Z4, Z5, S4 and S5; unsclerotised cuticle laterad opisthonotal shield carries setae R1 and S2, and lyrifissures idl2 and idRp; distance J5–J5 15 (14–15). Setal measurements shown in Table 3.

Ventral idiosoma — (Figure 13). Tritosternum similar to that of adult female, base 22 (21–22) long and 12 (11–13) wide, and a long pilose laciniae 60 (57–63), separated for about 87% of their total length. Sternal shield lightly sclerotised, smooth, with the widest region at the level of iv2, tapering posteriad st3; 138 (134–142) long and 51 (47–55) wide, with three pairs of setae (st1–st3) and two pairs of lyrifissures (iv1–iv2); distances st1–3 112 (110–113), st2–st2 68 (66–71), st3–st3 60 (58–62). Seta st5 on unsclerotised cuticle; distance st5–st5 62 (61–63). Unsclerotised opisthogastric cuticle with four pairs of setae (Jv1, Jv2, Jv5, Zv2), two pairs of lyrifissures, and two pairs of pores (gv3 adjacent to posterior margin of anal shield, gv2 posteriad coxae IV). Without endopodal or exopodal sclerites. Anal shield small, semi-ovoid, 50 (49–52) long and 42 (38–45) wide; anal opening 16 (15–16) long and 11 (10–12) wide. Cribrum wider than in female, with a pair of strap-like paranal extensions on either side of postanal seta. Shape of ventral idiosomatic setae as in adult female. Setal measurements shown in Table 4.

Peritreme and peritrematic plate — (Figure 12). Peritreme anlage short, reaching mid-level of coxa III. Peritrematic shield represented by a weakly sclerotised fragment at terminus of peritreme; lyrifissure id3 in the exterior margin of anterior plate; lyrifissure id7 in exterior margin of posterior portion of peritreme; pores gp, gd3 and gdp on unsclerotised cuticle.

Legs — Leg lengths: I: 286 (279–293); II: 242 (235–248); III: 209 (201–218); IV: 303 (297–308). Pretarsus lengths: II: 18 (17–19); III: 17 (17–18); IV: 21 (21–22). Setation (legs I–IV): coxae: 0-0/1 0/1-0, 0-0/1 0/1-0, 0-0/1 0/1-0, 0-0/1 0/0-0; trochanters: 1-0/1 0/1-1, 1-0/1 0/1-1, 1-0/1 0/1-1, 1-1/1 0/1-0; femora: 2-2/1 2/1-2, 1-2/1 2/1-1, 1-2/1 1/0-0, 1-2/0 1/0-0; genua: 1-2/0 2/1-1, 1-2/0 2/0-1, 1-2/0 2/0-1, 1-2/0 2/0-0; tibiae: 1-2/1 2/1-1, 1-1/1 2/1-1, 1-1/1 2/1-1, 1-1/1 2/1-1; tarsi: not counted, 17 (distal setae thickened), 17 (ventral setae thickened), 17 (ventral setae thickened).

Material examined

Holotype: female (ZMB-Arach 55437) from a laboratory colony deposited in MfN. Paratypes: two females (ZMB-Arach 55438 – 55439) collected in Karkom (Upper Galilee – Korazim, Israel), three females (ZMB-Arach 55440 – 55442) from a laboratory colony; four males (ZMB-Arach 55443 – 55446) from a laboratory colony, three deutonymphs (ZMB-Arach 55447 – 55449) from a laboratory colony, and three protonymphs (ZMB-Arach 55450 – 55452) from a laboratory colony deposited in MfN; a female (HUJINVACA540) from Hodaya (Coastal plain, Israel) deposited in HUJ; a female (OSAC collection number 0001418396) collected in Kfar Yehoshua (Jezreel Valley, Israel) and a male (OSAC collection number 0001418397) from a laboratory colony deposited in OSAC (Type deposition detailed in table 5).

Other material: two females (HUJINVACA541 – HUJINVACA542) collected in Hodaya (Coastal plain, Israel), two females (HUJINVACA548 – HUJINVACA549) collected in Kfar Yehoshua (Jezreel Valley, Israel), four females (HUJINVACA554 – HUJINVACA557) collected in Mehola (Beit Shean Valley, Israel), and five females (HUJINVACA600 – HUJINVACA602, HUJINVACA605 and HUJINVACA606) collected in Karkom (Upper Galilee – Korazim, Israel) deposited in HUJ. A female (ZMB-Arach 55453) collected in Kfar Yehoshua (Jezreel Valley, Israel) deposited in MfN. A female (OSAC collection number 0001418398) collected in Kfar Yehoshua (Jezreel Valley, Israel) deposited in OSAC.

Specimens with genetic information and sequence data are on BOLD in the dataset DS-MACROSPN (doi: 10.5883/DS-MACROSPN) and in GenBank (Table 5).

DNA barcodes

COI DNA barcodes were recovered from 15 specimens, including three female paratypes (BOLD:AEV3149, dx.doi.org/10.5883/DS-MACROSPN). It was not possible to recover genetic material from two of the specimens, and one specimen showed signs of contamination during processing. However, the three paratype females were confirmed as belonging to the new species based on morphological characteristics.

COI Sequences of M. karkomensis n. sp. ranged in length from 332 to 658 bp and displayed an average intraspecific p-distance of 0.21% (range: 0–1.61%), probably due to the homogeneity of the specimens obtained from a laboratory colony. Comparison of these sequences to the BOLD reference library and GenBank indicated that these are the first public sequence records for M. karkomensis n. sp. The closest available match is a species from South Africa tentatively identified only as Macrochelidae, which we verified as belonging to Macrocheles, with 11.4% divergence (p-distance).

The minimal intraspecific variation observed in M. karkomensis n. sp., compared to the sizable variations among other Macrocheles species, suggests that COI-DNA barcodes effectively distinguish between Macrocheles species. The highest intraspecific distance was observed in M. willowae (max 5%), but it remained lower than the interspecific divergence among Macrocheles species (13–30%). All COI sequences of Macrocheles species analysed showed high divergence from outgroup species (22–35%).

The consensus tree derived from the BI analysis of COI sequences was relatively well-supported, with most nodes having high posterior probabilities (11 nodes with supports greater than 85% and nine nodes with 100% support). However, two nodes had lower posterior probabilities of around 50% or less (Figure 14). Expanding sampling efforts with accurate morphological identification and more biomarkers will refine our understanding of differences between species and enhance the accuracy of future phylogenetic studies.

Remarks

Macrocheles karkomensis n. sp. is similar to Macrocheles scutatus (Berlese, 1904) described from specimens collected in Europe. M. scutatus is broadly distributed in the Old World (Berlese 1904; Filipponi and Pegazzano 1962; Walter and Krantz 1992).

A comparison between the new species herein described and M. scutatus was greatly facilitated by a loan of M. scutatus specimens from Dr. Bruno Cicolani (Department of Environmental Sciences, University of L´Aquila, Italy) (Figure 15), and by reference to published descriptions (Filipponi and Pegazzano 1962; Özbek, Bal and Doğan 2015; Walter and Krantz 1992). Adult female M. scutatus differs from M. karkomensis n. sp. in having a distinct procurved line on the dorsal shield and more pronounced ornamentation on the sternal shield. Filipponi and Pegazzano (1962) stated that the procurved line is only evident medially in some specimens of M. scutatus, but it appears to be present and complete in all of the specimens that we have examined. Most of the specimens in our collections of the new species have a weak procurved line, which in some of the type series is totally absent. In M. scutatus, sternal shield sutures l.o.p. are clearly united with l.m.t. anteriorly and joined to well-developed posterolateral rami medially, whereas in M. karkomensis n. sp. the sternal sutures are mostly represented by intermittent punctae (Figures 4 and 15a) that show neither clear l.o.p/l.m.t. convergence nor the presence of evident rami.

Differences between M. karkomensis n. sp. and M. scutatus tend to be clearer in males than in females. For example, the anterolateral margins of the ventrianal shield of M. scutatus males is concave adjacent to the insertions of coxae IV (Filipponi and Pegazzano 1962), while in M. karkomensis n. sp. males, anterolateral margins of the shield are slightly convex (Figure 8). Posterior dorsal setae Z4, Z5, S4 and S5 of M. scutatus males are elongate and distally pilose (Filipponi and Pegazzano 1962, Fig. VIII, 1), while in M. karkomensis n. sp. they are slightly longer than the small, smooth marginal R setae (Figure 8). Although not illustrated in the original description of M. scutatus, the posterior margin of the male dorsal shield of M. scutatus is crenulate, but is smooth in males of M. karkomensis n. sp. Males also differ in the pattern of ventral spurs on the legs, especially on legs IV (see Figures 16c,d and Table 6). Trochanter IV of male M. scutatus has a large rounded ventral spur (Fig. 16d), while femur IV has a complex of a large curved tubercle carrying terminal seta v1 and two adjacent, underlying spurs; genu IV has a single small tubercle that carries seta av1. Trochanter IV of male M. karkomensis n. sp. (Fig. 16c) lacks a ventral spur, and seta av1 is terminal on a barely perceptible tubercle. The femur IV spur complex is similar to that of M. scutatus, but the curved tubercle carries a much smaller spiniform terminal seta than that present in M. scutatus. Finally, the cribra of M. scutatus and M. karkomensis n. sp. males are primarily marginal, but attached rows of paranal spicules may be seen in some M. scutatus specimens and in all of the male M. karkomensis n. sp. in the material at hand.

Etymology

The specific name ''karkomensis'' to ''Karkom'', one of the type localities of the new species, and ''ensis'' denoting origin. Karkom is also the Hebrew name for the flowers of Crocus, also known as saffron.

Biological observations

Macrocheles karkomensis n. sp. has been observed in the laboratory to consume nematodes of the following species: Acrobeloides bodenheimeri (Steiner, 1936) Thorne, 1937 (Rhabditida: Cephalobidae), Oscheius tipulae (Lam & Webster, 1971) (Rhabditida: Rhabditidae), Pristionchus pacificus Sommer, Letter, Kim & Sternberg, 1996 (Rhabditida: Diplogastridae), Panagrellus nepenthicola (Menzel, 1922) Goodey, 1945, Panagrellus redivivus (Linnaeus, 1767) Goodey, 1945, Panagrellus silusioides Thalolikhin, 1965 (Rhabditida: Panagrolaimidae), Meloidogyne incognita (Kofoid & White, 1919) Chitwood, 1949 (Rhabditida: Meloidogynidae) and two unidentified species of Panagrolaimus (Rhabditida: Panagrolaimidae).

Immature development in the laboratory was shortest for mites fed on A. bodenheimeri and the two Panagrolaimus species (approximately four days). Duration was slightly longer for mites fed with M. incognita (about five days). Mites failed to develop on O. tipulae beyond the protonymphal stage (Rueda-Ramírez et al. 2023). Mite fecundity was highest when feeding on A. bodenheimeri and was almost null (approximately 0.12 eggs/female) when feeding on M. incognita (Rueda-Ramírez et al. 2023). Macrocheles karkomensis n. sp. has also been observed to feed on the astigmatine oribatid Carpoglyphus lactis (Linnaeus, 1758) (Carpoglyphidae) (Rueda-Ramírez pers. obs).

Unlike most of the known species of the scutatus subgroup, no phoretic associations are known for M. karkomensis n. sp. However, females in laboratory cultures readily grasped the hairs of a fine brush when offered, suggestive of a phoretic response.

Discussion

The scutatus complex was proposed by Walter and Krantz (1992) within the scutatus subgroup of the glaber group, to include seven species. This complex comprises species with reduced ornamentation on the ventral shields, but which retain the basic glaber ornamentation. Species were separated mainly by degree of pilosity of the female idiosoma and the pattern of ornamentation of their sternal shield (Walter and Krantz 1986; Walter and Krantz 1992) (Table 6).

Our observations of M. karkomensis n. sp. and of M. scutatus show that male characteristics may play an important role in the separation and identification of species in the scutatus complex. Unfortunately, few males of the scutatus complex have been described (M. karkomensis n. sp., M. scutatus, and M. dispar). Unlike the phoretic females, most of which have been collected from scarabaeine beetle phoronts, males of M. karkomensis n. sp. and M. scutatus have been recovered only from laboratory-reared populations. The provenance of known M. dispar males (Berlese 1918) is unclear, but data in the Berlese Acaroteca in Florence suggests that at least some male dispar may have been collected along with females on the same individual beetle (Copris sp.) (Castagnoli and Pegazzano 1985).

The females of the species here described conform to the diagnosis for the scutatus complex proposed by Walter and Krantz (1992). However, knowledge of the males of the other species will allow better characterization of the diagnoses. Based on the present status of our knowledge of the males of this complex, a new characterization of recognized scutatus complex species based on females and known males is presented below (Table 6).

Updated characterization of the scutatus complex

Sternal shield typically longer than wide and with reduced ornamentation; l.arc. not deeply punctate, sometimes weak, occasionally doubled; l.m.t. distinct, l.o.p. weak, ramus often lost or weak; a.p. reduced to scattered punctae or lost. Epigynal shield varying from smooth to reticulate, with reticules formed by punctate lines. Ventrianal shield rounded-subcordate, length equal to or greater than width, evenly reticulate; postcoxal pore (gv2) distinct and adjacent to parapodal plate. Dorsal shield reticulate, with procurved line present, sometimes weak; setae j1 appressed and distally pilose, setae j4 and z4 similarly ornamented and often thickened, J5 serrate. Known males with separate sternitigenital and ventrianal shields; ventrianal shield with deeply concave or convex (subcordate or peltate) anterolateral margins adjacent to insertions of coxa IV; femur II with one large spur; leg IV heavily ornamented with several spurs: femur IV with 1–2 spurs and a tubercle, genu IV with tiny or large protuberance, and tarsus IV with a distal rounded spur.

Updated key to known scutatus complex species initially published by Walter and Krantz (1992)

1. Setae j3 distally pilose. Southern Africa
...... 2

— Setae j3 smooth
...... 3

2. Dorsal shield setae bipectinate (z1, z5 and j6 may be smooth in some specimens), postanal and opisthogastric setae similarly ornamented
...... M. agnosticus

— Most dorsal shield setae smooth (j1, j3-4, s2, r2-3 bipectinate distally); postanal and opisthogastric setae without ornamentation
...... M. subscutatus

3. L.o.p. disjunct from l.m.t. and with a weak ramus or without ramus, a.p. reduced to a few punctae, or lost
...... 4

— L.o.p. united to l.m.t. and with ramus; a.p. distinct
...... 6

4. L.arc. lost; setae j4 distally pectinate, other dorsal setae smooth. Southern Africa
...... M. neovernalis

— L. arc. present or weak; setae j4 and z4 distally pilose. Male ventrianal shield with convex anterolateral margin adjacent to the insertion of coxae IV
...... 5

5. L. arc. weak; setae r2 and r3 usually distally pilose. Femur IV of male with a spurs and a set setiferous tubercle. Asia
...... M. dispar

— L. arc. strongly procurved; setae r2 and r3 always simple. Femur IV of male with two spurs and tubercles with a small terminal spiniform seta. Israel
...... M. karkomensis n. sp.

6. Sternal shield distinctly reticulate anterior to l.m.t., l.arc. indistinct; a.p. finely punctate. Western Europe
...... M. neoscutatus

— Sternal shield with one or two l.arc., without distinct reticulation as above; a.p. coarsely punctae
...... 7

7. With a single strongly procurved l.arc. Male ventrianal shield with deeply concave anterolateral margin adjacent to the insertions of coxae IV. Presently considered cosmopolitan
...... M. scutatus

— With two transverse parallel l.arc. which are weakly procurved medially. Southern Africa
...... M. medialis

Additional observations

Based on Calugar (2008) and O. Makarova (pers. comm.), the scutatus complex appears to include additional species whose females display little or no significant morphological differences from females of M. scutatus s. str., but which may be distinguished based on male characters alone. Clearly, characterization of the complex must be considered as ongoing, eventually to be supplemented with data from additional scutatus populations worldwide and hopefully supported by comparative genetic information.

An interesting observation in the immature stages of the new species described here is the absence of seta Zv1 in the deutonymph, which only appears in the adult stage. Evans &Till (1965) observed that, at least in certain dermanyssoid Gamasina, opisthogastric setae Zv1 first appear in the deutonymphal stage. In the new species, Zv1 appears to be smaller and more anteriorly located than observed in other species of the same complex (Walter and Krantz 1992). However, we interpret its presence in adults as being associated with gv2 and ivo, suggesting the possibility of its loss in the deutonymph stage and reappearance in the adult stage. At this point, it is not possible to conclude that loss and reappearance of Zv1 is characteristic of the scutatus complex or of its subgroup, given the limited information presently available on immatures.

Variation in opisthogastric chaetotaxy of some female Macrochelidae was reported earlier by Hyatt & Emberson (1988). Similar variation was found in males of M. karkomensis n. sp. during this study, particularly on the sternogenital shield. Most of the males in our series had only one seta st4 (Figure 8a, c), while in others both st4 setae were either present or absent (Figure 8b).

Another characteristic seen in adults of M. karkomensis n. sp. is the presence of two lyrifissures on the peritreme. These structures have previously been observed in Macrocheles embersoni Azevedo, Castilho & Berto (2017) and Macrocheles neotransversus Quintero-Gutierrez and Cómbita-Heredia (2020). Based on our observations, the lyrifissures referred to as gp? and gp in the original description of M. neotransversus are homologous with the anterior and posterior peritrematic lyrifissures depicted in M. embersoni by Azevedo et al. (2017). These structures are identified in this paper as lyrifissures ipc1 and ipc2 (Figures 2a (ipc1 expressed anterodorsally) and 4 in the female, and in Figures 7a (also expressed anterodorsally) and 8a in the male). Therefore, the pore and the lyrifissure on the peritrematic shield correspond to gd3 (anterior) and ip (posterior) (Figure 17).

Although genetic information on most species of the scutatus group is not currently available, we were able to include in our genetic comparison with M. karkomensis two species that appear to belong to this group or are close to it. In light of their potential usefulness as biological control agents (Azevedo et al. 2015; Krantz 1983), further efforts should be made to expand collection of scutatus group species so that reliable morphological identifications may be correlated with genetic information extracted from collected specimens (Rueda-Ramírez et al. 2022; Young et al. 2019; Young and Hebert 2022). This in turn may lead to a better understanding of species differences and increased precision in phylogenetic, ecological, and practical application studies. We also highlight the importance of including more molecular markers to increase phylogenetic precision (Rokas and Carroll 2005) and confirm identity of specimens whose sequences are publicly available.

Acknowledgements

To Shira Gal, Newe Ya′ar Research Center, Agricultural Research Organization (ARO), Israel, for his outstanding assistance with colonies, photos and conveying biological material. To Jason Dunlop, Anja Friederichs, and Birger Neuhaus, Museum für Naturkunde, Leibniz Institute for Evolution and Biodiversity Science, Berlin, Germany, for opening a space for the first author to look through the collection, use the microscopes and work with the material. To Bruno Cicolani for the loan of slides of M. scutatus from his personal collection. To Thomas Stach and Peer Martin, Institute of Biology, Molecular Parasitology group, Humboldt Universität zu Berlin, Germany, for granting access to their scanning electron microscope, advising and helping in preparing the samples, and taking photographs. To Gilberto J. de Moraes, Department of Entomology and Acarology, Escola Superior de Agricultura Luiz de Queiroz, Universidade de São Paulo, Brazil, for answering questions, reviewing the manuscript, and providing valuable suggestions. To María L. Moraza, Universidad de Navarra, Instituto de Biodiversidad y Medioambiente, for her time and invaluable and accurate comments and suggestions on the adenotaxy and porotaxy of M. karkomensis. To Olga Makarova, Severtsov Institute of Ecology and Evolution, Russian Academy of Sciences, for important information shared, and for her enthusiasm for future cooperative research on the scutatus complex. To Wayne Knee, Agriculture and Agri-Food Canada, for kindly sending photos of his specimens and for allowing us to base our phylogenetic analysis on his previously published analysis of some Macrocheles species. Finally, to the anonymous reviewers whose comments and suggestions significantly improved the manuscript. Funding: This work was supported by German Research Foundation (DFG) [grant number RU780/20-1] (′Harnessing the soil food web for the biological control of rootknot nematodes').

References

1. Athias-Henriot C. 1975 . Nouvelles notes sur les Amblyseiini. II. Le releve organotaxique de la face dorsale adulte (Gamasides protoadeniques, Phytoseiidae). Acarologia, 27(1): 20-29.
2. Azevedo L.H., Castilho R.C., Berto M.M., De Moraes G.J. 2017. Macrochelid mites (Mesostigmata: Macrochelidae) from Sao Paulo state, Brazil, with description of a new species of Macrocheles. Zootaxa, 4269(3): 413-426. https://doi.org/10.11646/zootaxa.4269.3.5
3. Azevedo L.H. de, Emberson R.M., Esteca F. de C.N., Moraes G.J. de 2015. Macrochelid mites (Mesostigmata: Macrochelidae) as biological control agents. In: Carrillo D., Moraes G. J. de, Peña J. E. (Eds). Prospects for biological control of plant feeding mites and other harmful organisms, Switzerland: Springer International Publishing. pp. 103-132. https://doi.org/10.1007/978-3-319-15042-0_4
4. Azevedo L.H., Ferreira M.P., Castilho R. de C., Cançado P.H.D., Moraes G.J. de 2018. Potential of Macrocheles species (Acari: Mesostigmata: Macrochelidae) as control agents of harmful flies (Diptera) and biology of Macrocheles embersoni Azevedo, Castilho and Berto on Stomoxys calcitrans (L.) and Musca domestica. Biol. Control, 123: 1-8. https://doi.org/10.1016/j.biocontrol.2018.04.013
5. Beaulieu F., Dowling A.P.G., Klompen H., Moraes G.J. de, Walter D.E. 2011. Superorder Parasitiformes Reuter, 1909. In: Zhang, Z.-Q. (Ed.) Animal biodiversity: An outline of higher-level classification and survey of taxonomic richness. Zootaxa, 3148(1): 123-128. https://doi.org/10.11646/zootaxa.3148.1.23
6. Berlese A. 1904. Acari Nuovi. Manipulus IIus. Redia, 1: 258-280.
7. Berlese A. 1910. Lista di nuove specie e nuovi generi di Acari. Redia, 6: 242-297.
8. Berlese A. 1918. Centuria quarta di Acari Nuovi. Redia Firenze, 13: 115-192.
9. Calugar A. 2008. New and rare species of mites from some forest ecosystems of the Danube delta biosphere reserve. In: Bertrand M., Kreiter S., McCoy K. D., Migeon A., Navajas M., Tixier M. S., Vial L. (Eds). Integrative Acarology. Proceedings of the 6th European Congress6th European Congress European Association of Acarologists, Montpellier, France: EURAAC. p. 167-174.
10. Castagnoli M., Pegazzano 1985. Catalogue of the Berlese Acaroteca. Firenze, Italia: Istituto Sperimentale per la Zoologia Agraria. pp. 490.
11. Emberson R.M. 2010. A reappraisal of some basal lineages of the family Macrochelidae, with the description of a new genus (Acarina: Mesostigmata). Zootaxa, 53(2501): 37-53. https://doi.org/10.11646/zootaxa.2501.1.3
12. Evans G.O. 1963. Observations on the chaetotaxy of the legs in the free-living Gamasina (Acari: Mesostigmata). Bulletin of the British Museum (Natural History). Zoology, 10(5): 275-303. https://doi.org/10.5962/bhl.part.20528
13. Evans G.O. 1969. Observations on the ontogenetic development of the chaetotaxy of the tarsi nomenclature for the chaetotaxy of the tarsi nomenclature for the chaetotaxy of tarsi II-IV of legs II-IV in the Mesostigmata (Acari). In: Evans G.O. (Ed). Proceedings of the 2nd International Congress of Acarology, 1967, Sutton Bonington, England: Akademiai Kiado. p. 195-200.
14. Evans G.O., Till W.M. 1965. Studies on the British Demanyssidae (Acari: Mesostigmata). Part I. External Morphology. Bull. Br. Mus. (Nat. Hist.) Zool., 13(8): 246-294. https://doi.org/10.5962/bhl.part.16752
15. Filipponi A., Dojmi di Delupis G. 1963. Sul regime dietetico di alcuni macrochelidi (Acari: Mesostigmata), associati in natura a muscidi di interesse sanitario. Riv. Parassitol., XXIV(4): 277-287.
16. Filipponi A., Pegazzano F. 1962. Specie Italiane del gruppo-glaber (Acarina, Mesostigmata, Macrochelidae, Macrocheles). Redia, 47: 211-238 + Plates XXI-XXIII.
17. Folmer O., Black M., Hoeh W., Lutz R., Vrijenhoek R. 1994. DNA primers for amplification of mitochondrial cytochrome c oxidase subunit I from diverse metazoan invertebrates. Mol. Mar. Biol. Biotechnol., 3: 294-299.
18. Hebert P.D.N., Cywinska A., Ball S.L., deWaard J.R. 2003. Biological identifications through DNA barcodes. Proc. R. Soc. Lond. B, 270: 313-321. https://doi.org/10.1098/rspb.2002.2218
19. Huelsenbeck J.P., Ronquist F. 2001. MRBAYES: Bayesian inference of phylogenetic trees. Bioinform., 17: 754-755. https://doi.org/10.1093/bioinformatics/17.8.754
20. Ito Y. 1973. The effects of nematode feeding on the predatory efficiency for house fly eggs and reproduction rate of Macrocheles muscaedomesticae (Acarina: Mesostigmata). Jpn. Soc. Med. Entomol. Zool., 23(3): 209-213. https://doi.org/10.7601/mez.23.209
21. Ivanova N.V., Grainger C.M. 2007a. CCDB protocols, COI amplification [Internet]. [June 3, 2024]. Guelph: Canadian Center for DNA Barcoding. Available from: ccdb.ca/site/wp-content/ uploads/2016/09/CCDB_Amplification.pdf
22. Ivanova N.V., Grainger C.M. 2007b. CCDB protocols, sequencing [Internet]. [June 3, 2024]. Guelph: Canadian Center for DNA Barcoding. Available from: ccdb.ca/site/wp-content/uploads/2016/ 09/CCDB_Sequencing.pdf
23. Ivanova N.V., de Waard J.R., Hebert P.D.N. 2007. CCDB protocols, glass fiber plate DNA extraction [Internet], [June 3, 2024]. Guelph: Canadian Center for DNA Barcoding. Available from: ccdb.ca/site/wp-content/uploads/2016/09/CCDB_DNA_Extraction.pdf
24. Knee W. 2017. New Macrocheles species (Acari, Mesostigmata, Macrochelidae) associated with burying beetles (Silphidae, Nicrophorus) in North America. ZooKeys, 721: 1-32. https://doi.org/10.3897/zookeys.721.21747
25. Krantz G.W. 1972. Macrochelidae from Hamburg (Acari, Mesostigmata), with descriptions of two new species. Ent. Mitt. Zool. Mus. Hamburg, 4(78): 263-275.
26. Krantz G.W. 1983. Mites as biological control agents of dung-breeding flies, with special reference to the Macrochelidae. In: Hoy C. W., Cunningham G. L., Knutson L. (Eds). Biological control of pests by mites: proceedings of a conference held April 5-7, 1982, Berkeley, US.: University of California, Berkeley, and APHIS-PPQ, U.S.D.A., and Agricultural Research Service, U.S.D.A. p. 91-98.
27. Krantz G.W. 1998. Reflections on the biology, morphology and ecology of the Macrochelidae. Exp. Appl. Acarol., 22: 125-137.
28. Krantz G.W. 2009. Habits and Habitats. In: Krantz G. W., Walter D. E. (Eds). A Manual of Acarology. Lubbock, Texas: Texas Tech University Press. p. 64-82.
29. Lindquist E.E., Evans G.O. 1965. Taxonomic concepts in the Ascidae, with a modified setal nomenclature for the idiosoma of the Gamasina (Acarina: Mesostigmata). Mem. Entomol. Soc. Can., 97(S47): 5-66. https://doi.org/10.4039/entm9747fv
30. Lindquist E.E., Krantz G.W., Walter D.E. 2009. Order Mesostigmata. In: Krantz G. W., Walter D. E. (Eds). A Manual of Acarology. Lubbock, Texas: Texas Tech University Press. p. 124-232.
31. Maddison W.P., Maddison D.R. 2023. Mesquite: a modular system for evolutionary analysis [Software]. Version 3.81. Available from: http://www.mesquiteproject.org.
32. Moens M., Perry R.N., Starr J.L. 2009. Meloidogyne species - a diverse group of novel and important plant parasites. In: Perry R. N., Moens M., Starr J. L. (Eds). Root-knot nematodes. Wallingford, Oxfordshire: CABI. p. 1-17. https://doi.org/10.1079/9781845934927.0001
33. Moraza M.L. 2025. Glandular and non-glandular cuticular organs on the idiosoma of Gamasina mites (Acari: Mesostigmata). Persian J. Acarol., 14(1): 151-171.
34. Nylander J.A.A. 2004 . MrModeltest v2. Version v2.4 (2018). Program distributed by the author [Software]. Evolutionary Biology Centre, Uppsala University. Available from: https://github.com/nylander/MrModeltest2.
35. Özbek H.H., Bal D.A., Doğan S. 2015. The genus Macrocheles Latreille (Acari: Mesostigmata: Macrochelidae) from Kelkit Valley (Turkey), with three newly recorded mite species. Turk. J. Zool., 39: 768-780. https://doi.org/10.3906/zoo-1409-14
36. Porco D., Bedos A., Deharveng L. 2010. Description and DNA barcoding assessment of the new species Deutonura gibbosa (Collembola: Neanuridae: Neanurinae), a common springtail of Alps and Jura. Zootaxa, 2639: 59-68. https://doi.org/10.11646/zootaxa.2639.1.6
37. Quintero-Gutiérrez E.J., Cómbita-Heredia O., Klompen H. 2020. Mesostigmatid mites associated with Oxysternon conspicillatum (Coleoptera: Scarabaeidae): a new species of Macrocheles, description of the male of M. magna, and four new records for Colombia. Int. J. Acarol., 46(7): 496-512. https://doi.org/10.1080/01647954.2020.1815838
38. Rambaut A. 2018. FigTree Version 1.4.4 [Software]. Institute of Evolutionary Biology, University of Edinburgh. Available from: https://github.com/rambaut/figtree/releases.
39. Rodriguez J.G., Wade C.F., Wells C.N. 1962. Nematodes as a natural food for Macrocheles muscaedomesticae (Acarina: Macrochelidae), a predator of the house fly egg. Ann. Entomol. Soc. Am., 55(5): 507-511. https://doi.org/10.1093/aesa/55.5.507
40. Rokas A., Carroll S.B. 2005. More Genes or More Taxa? The Relative Contribution of Gene Number and Taxon Number to Phylogenetic Accuracy. Mol. Biol. Evol., 22(5): 1337-1344. https://doi.org/10.1093/molbev/msi121
41. Rueda-Ramírez D., Carta L., Mowery J., Bauchan G., Ochoa R., Young M., Santos J.C., Palevsky E. 2022. In memory of Gary Bauchan: Integrated taxonomy of soil mites in farming systems. Syst. Appl. Acarol., 27(2): 181-208. https://doi.org/10.11158/saa.27.2.3
42. Rueda-Ramírez D., Narberhaus A., Palevsky E., Hallmann J., Ruess L. 2023. Bottom-up effects of nematode prey on soil predatory mites (Acari: Mesostigmata). Soil Biol. Biochem., 185: 109143. https://doi.org/10.1016/j.soilbio.2023.109143
43. Ryke P.A.J., Meyer M.K.P. 1958. Some parasitoid mites (Mesostigmata: Acarina) associated with Coleoptera in the Western Transvaal. J. Entomol. Soc. S. Afr., 21(1): 139-161.
44. Swofford D.L. 2003. PAUP*. Phylogenetic Analysis Using Parsimony (*and Other Methods). Version 4.0a [Software]. Sinauer Associates. Available from: https://paup.phylosolutions.com/.
45. Walter D.E. 1985. A revision of the Macrocheles glaber species group (Acari:Macrochelidae) using phylogenetic systematics. Doctor of Philosophy thesis. Corvallis, OR (USA): Oregon State University. pp. 214.
46. Walter D.E., Krantz G.W. 1986. A review of glaber-group (s-str) species of the genus Macrocheles (Acari, Macrochelidae), and a discussion of species complexes. Acarologia, 27(4): 277-294.
47. Walter D.E., Krantz G.W. 1992. A review of glaber-like species with reduced sclerotization and ventral ornamentation: The scutatus subgroup (Acari: Macrochelidae: Macrocheles). Int. J. Acarol., 18(3): 241-249. https://doi.org/10.1080/01647959208683956
48. Young M.R., Hebert P.D.N. 2022. Unearthing soil arthropod diversity through DNA metabarcoding. PeerJ, 10: e12845. https://doi.org/10.7717/peerj.12845
49. Young M.R., Moraza M.L., Ueckermann E., Heylen D., Baardsen L.F., Lima-Barbero J.F., Gal S., Gavish-Regev E., Gottlieb Y., Roy L., Recht E., El Adouzi M., Palevsky E. 2019. Linking morphological and molecular taxonomy for the identification of poultry house, soil, and nest dwelling mites in the Western Palearctic. Sci Rep, 9(1): 5784. https://doi.org/10.1038/s41598-019-41958-9

instructions